β-catenin Search Results


  • Logo
  • About
  • News
  • Press Release
  • Team
  • Advisors
  • Partners
  • Contact
  • Bioz Stars
  • Bioz vStars
  • 99
    Cell Signaling Technology Inc β catenin
    α-SYN is required for the maintenance, but not the generation, of adult subependymal NSCs. a , Representative SEZ whole-mount preparations of 2 month mice immunostained for γ-tubulin (blue), <t>β-catenin</t> (red), and GFAP (green). White dashed lines indicate examples of pinwheels. b , Pinwheel and B1 cell density per mm 2 at the anterodorsal (plain) and posterodorsal (striped) SEZ regions. Snca knock-out mice display lower density of pinwheels and B1 cells than wild-type littermates. c , Representative immunofluorescent detection of GFAP (red) and Ki67 (green) in the SEZ of 2 month wild-type ( Snca +/+ ) and mutant ( Snca −/− ) mice. White arrowhead points at GFAP + Ki67 + cells. d , Graphs showing the mean proportion (± SEM) of GFAP + cells relative to total number of cells (on the left axis) and GFAP + cells that are in cell cycle (Ki67 + ) or express S100β in the SEZ of Snca +/+ and Snca −/− mice at 2 months (on the right axis). e , Representative immunofluorescent detection of GFAP (red) and Ki67 (green) in the SEZ of postnatal day 7 mice. White arrowheads point at GFAP + Ki67 + cells. DAPI was used for nuclear staining. f , FACS analysis of qNSCs and aNSCs within the Lin − /CD24 −/low /GLAST + /CD9 high population discriminated by EGFR expression, showing a decrease in aNSCs in mutant mice. g , FACS analysis for CD9 in the Lin − /CD24 −/low /GLAST + population showing an apparent increase in niche CD9 low astrocytes in mutant mice. h , Graph representing the levels of CD9 expression in SEZ NSCs and in striatal astrocytes of wild-type mice and in cells of the mutant SEZ (orange line). * p
    β Catenin, supplied by Cell Signaling Technology Inc, used in various techniques. Bioz Stars score: 99/100, based on 7322 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
    https://www.bioz.com/result/β catenin/product/Cell Signaling Technology Inc
    Average 99 stars, based on 7322 article reviews
    Price from $9.99 to $1999.99
    β catenin - by Bioz Stars, 2020-03
    99/100 stars
      Buy from Supplier

    99
    Abcam β catenin
    The functional role of miR-139 in XIST regulating <t>β-catenin</t> and ECM proteins ( A ) miR-139 expression in response to XIST knockdown in MLFCs was determined by using real-time PCR. ( B ) miR-139 mimics or miR-139 inhibitor was transfected into MLFCs to achieve ectopic miR-139 expression or miR-139 inhibition, as verified by real-time PCR assays. ( C ) The expression levels of XIST in response to ectopic miR-139 expression or miR-139 inhibition in MLFCs was determined by using real-time PCR assays. ( D ) and ( E ) After co-processing sh-XIST and miR-139 inhibition in MLFCs, β-catenin protein levels (total, nucleus, cytoplasm) were determined by using Western blot assays. ( F ) After co-processing sh-XIST and miR-139 inhibition in MLFCs, Collagen I and α-SMA protein levels were determined by using Western blot assays. The data are showed as mean± SD of three independent experiments. * P
    β Catenin, supplied by Abcam, used in various techniques. Bioz Stars score: 99/100, based on 3235 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
    https://www.bioz.com/result/β catenin/product/Abcam
    Average 99 stars, based on 3235 article reviews
    Price from $9.99 to $1999.99
    β catenin - by Bioz Stars, 2020-03
    99/100 stars
      Buy from Supplier

    99
    Becton Dickinson β catenin
    GSK3β phosphorylates Dzip1. (A and B) Dzip1 interacts with GSK3β. Endogenous GSK3β was immunoprecipitated by Dzip1 but not IgG (A), and endogenous Dzip1 was immunoprecipitated with GFP-GSK3β in HEK 293T cells (B). (C and D) Dzip1 is co-localized with GSK3β at the basal body. G0-phase NIH 3T3 cells expressing GFP-GSK3β were immunostained for Dzip1 and AcTub (C), or cells expressing BFP-Centrin2 were immunostained with GSK3β and Dzip1 (D). Scale bar: 5 μm. (E) GSK3β binds Dzip1 in a kinase-substrate interaction manner. Wild-type (WT) GFP-GSK3β and the mutants S9A, K85R, and R96A were each co-expressed with Myc-Dzip1 in G0-phase HEK 293T cells, and treated with the CK1 inhibitor D4476 or the CK2 inhibitor CX4945. Note that treatment with CX4945 but not D4476 led to a significant decrease in the extent of the up-shifted Dzip1 bands, although the binding of Dzip1 to the GFP-GSK3β variants showed no difference. The extent of the up-shifting of the Dzip1 bands was decreased in K85R-expressing cells. (F) Phosphorylation of Dzip1 is coordinated with GSK3β activation. The kinase activity of GSK3β was negatively correlated with serum stimulation in NIH 3T3 cells. Note that the up-shifted bands (arrowheads) of Dzip1 became evident after serum depletion for 24–48 h, and disappeared after serum restimulation. γ-Tubulin was set as a loading control. (G) GSK3β phosphorylates Dzip1 in vivo. In resting mouse embryo fibroblast (MEFs) treated versus not treated with GSK3 and CK2 inhibitors, the Dzip1 bands were up-shifted less in GSK3- and CK2-inhibited cells. The protein levels of total <t>β-Catenin</t> and GSK3β were steady, but the phosphorylated (S33/37/T41) β-Catenin specifically disappeared from GSK3-inhibited cells. α-Tubulin was set as a loading control. (H) GSK3β phosphorylates Dzip1 in vitro. Auto-phosphorylation of GSK3β (55 kD), and the phosphorylated bands of the middle (28 kD), C-terminus (36 kD), and N- terminus (50 kD) of Dzip1 are shown (left panel). Coomassie blue staining of the gel shows the loaded amounts of Dzip1 fragments (right panel). Note that the S520A mutation resulted in decreased phosphorylation of Dzip1 by GSK3β. (I) Inhibition of GSK3 by BIO causes loss of phospho-S520 in Dzip1.
    β Catenin, supplied by Becton Dickinson, used in various techniques. Bioz Stars score: 99/100, based on 5305 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
    https://www.bioz.com/result/β catenin/product/Becton Dickinson
    Average 99 stars, based on 5305 article reviews
    Price from $9.99 to $1999.99
    β catenin - by Bioz Stars, 2020-03
    99/100 stars
      Buy from Supplier

    99
    Proteintech β catenin
    a  qRT-PCR analysis of chromatin modifiers’ fold changes after the inhibition of Wnt signaling. rECA109 and rKyse150 cells were treated with iCRT14 (25 μM for 24 h) and analyzed for expression of chromatin modifiers using qRT-PCR.  b  qRT-PCR analysis of HMGB1 and CHD4. 24 h after treatment of WNT1 protein, qRT-PCR was performed to analyze the expression of HMGB1 and CHD4. Mean ± SD,  N  = 3, * P
    β Catenin, supplied by Proteintech, used in various techniques. Bioz Stars score: 99/100, based on 361 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
    https://www.bioz.com/result/β catenin/product/Proteintech
    Average 99 stars, based on 361 article reviews
    Price from $9.99 to $1999.99
    β catenin - by Bioz Stars, 2020-03
    99/100 stars
      Buy from Supplier

    93
    SLIT2 LTD β catenin
    SLIT/ROBO1 signaling regulates the subcellular localization of <t>β-catenin</t>
    β Catenin, supplied by SLIT2 LTD, used in various techniques. Bioz Stars score: 93/100, based on 106 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
    https://www.bioz.com/result/β catenin/product/SLIT2 LTD
    Average 93 stars, based on 106 article reviews
    Price from $9.99 to $1999.99
    β catenin - by Bioz Stars, 2020-03
    93/100 stars
      Buy from Supplier

    99
    Thermo Fisher β catenin
    TRAIL-TZD <t>regulatesβ-catenin</t> expression via a post-translational mechanism independent of GSK3β. A , Huh-7 cells were treated with DMSO or the TRAIL-TZD combination in the presence of 20 μg/ml cycloheximide ( CHX, lanes 3–12 ). Cells were harvested at 0, 2, 4, 6, and 8 h after treatment followed by Western blot analysis with the indicated antibodies. The Huh-7 cells treated with either DMSO or the TRAIL-TZD combination in the absence of cycloheximide were included in lanes 1 and 2 , respectively, as positive controls to show TRAIL-TZD effects on <t>β-catenin.</t> B , Huh-7 cells were treated with the TRAIL-TZD combination for 24 h in the presence (+) or absence (-) of a pretreatment with the GSK3β inhibitor AR-A014418. At the end of the incubation total cell extracts were prepared and Western blot analysis was performed with the indicated antibodies. C , wild type ( WT +/+ ) or GSK3β knock-out ( KO -/- ) MEFs were incubated with either DMSO or the TRAIL-TZD combination. Total cell extracts were prepared at the indicated time points followed by Western blot analysis with the antibodies shown. D , Huh-7 cells were treated as in B followed by apoptosis assay utilizing the cell death detection ELISA PLUS kit. The data represent the mean ± S.D. of two independent experiments. E , phase-contrast microscopic pictures showing apoptotic morphology of Huh-7 cells following treatment with either DMSO or the TRAIL-TZD combination in the presence (+) or absence (-) of AR-A014418.
    β Catenin, supplied by Thermo Fisher, used in various techniques. Bioz Stars score: 99/100, based on 2181 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
    https://www.bioz.com/result/β catenin/product/Thermo Fisher
    Average 99 stars, based on 2181 article reviews
    Price from $9.99 to $1999.99
    β catenin - by Bioz Stars, 2020-03
    99/100 stars
      Buy from Supplier

    96
    Agilent technologies β catenin
    Consecutive sections with foci of (upper left) intense FHL2 expression with concomitant (upper right) reduced E-cadherin expression and (lower) appearance of nuclear expression of  β -catenin (arrows) in areas of EMT. Note the lower FHL2 expression as well as the stronger E-cadherin expression and the absence of nuclear  β -catenin expression in adjacent well-differentiated tumour glands (** magnification × 400).
    β Catenin, supplied by Agilent technologies, used in various techniques. Bioz Stars score: 96/100, based on 393 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
    https://www.bioz.com/result/β catenin/product/Agilent technologies
    Average 96 stars, based on 393 article reviews
    Price from $9.99 to $1999.99
    β catenin - by Bioz Stars, 2020-03
    96/100 stars
      Buy from Supplier

    92
    Johnson & Johnson β catenin
    Cad6B CTF2s are stabilized through <t>β-catenin</t> binding. (A) Cad6B CTF2 mutants were generated through alanine substitutions within the β-catenin minimal binding domain and at key adjacent phosphorylation sites. Black bars represent the ratio of coimmunoprecipitated β-catenin to immunoprecipitated (IP) mutant CTF2 levels normalized to the wild-type (WT) ratio (set at 100%); gray bars represent the input levels of mutant CTF2 protein (normalized to β-actin levels) relative to WT CTF2 input levels (set at 100%). Means that share letter superscripts are not significantly different (P
    β Catenin, supplied by Johnson & Johnson, used in various techniques. Bioz Stars score: 92/100, based on 19 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
    https://www.bioz.com/result/β catenin/product/Johnson & Johnson
    Average 92 stars, based on 19 article reviews
    Price from $9.99 to $1999.99
    β catenin - by Bioz Stars, 2020-03
    92/100 stars
      Buy from Supplier

    99
    Santa Cruz Biotechnology β catenin
    FoxO1 knockdown promotes migration, invasion potential, and EMT in OSCC cells. (A) FoxO1 expression was detected by qRT-PCR and Western blot after transfected with FoxO1 shRNA in OSCC cells. (B) Images of the wound closure of monolayer Cal-27 and Tca8113 cells with FoxO1 knockdown at the time point of 0 and 24 h are presented on the left. Quantitative results are illustrated on the right. (C) The effect of FoxO1 knockdown on OSCC cells invasion were determined by Transwell assay with Matrigel, and the representative images are on the left. Quantitative results are illustrated on the right. (D) The effects of Cal-27-FoxO1 knockdown on expressions of EMT markers, E-cadherin, N-cadherin, <t>β-catenin,</t> and Vimentin, were measured using qRT-PCR and Western blot. And the effects of Cal-27-sh-2-STAT3/sh-1-FoxO1 on expressions of EMT markers, E-cadherin, N-cadherin, β-catenin, and Vimentin, were measured using qRT-PCR compared with those of sh-2-STAT3/scramble group. β-actin was used as a loading control. (E) Flow cytometry was used to examine the percentage of apoptotic cells in Cal-27 cells with FoxO1 knockdown and scramble control cells. All assays were carried out in triplicate. Results were shown as means ± SD. * P
    β Catenin, supplied by Santa Cruz Biotechnology, used in various techniques. Bioz Stars score: 99/100, based on 4329 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
    https://www.bioz.com/result/β catenin/product/Santa Cruz Biotechnology
    Average 99 stars, based on 4329 article reviews
    Price from $9.99 to $1999.99
    β catenin - by Bioz Stars, 2020-03
    99/100 stars
      Buy from Supplier

    99
    Horizon Discovery β catenin
    PolyP-70-mediated expression of cyclin D1 in the absence and presence of siRNA or specific inhibitors for signaling molecules. (A) The efficiency of gene knockdown of <t>β-catenin</t> ( > 75%) was determined 48h post transfection by Western-blotting using a specific antibody. The (B) PolyP-mediated up-regulation of cyclin D1 was monitored after siRNA knockdown of β-catenin in EA.hy926 endothelial cells. (C–D) PolyP-mediated overexpression of cyclin D1 in the absence and presence of increasing concentrations of two different Wnt signaling inhibitors (iCRT3 and PNU-74654). (E) PolyP-mediated up-regulation of cyclin D1 in the absence and presence of increasing concentrations of PI3K inhibitor (wortmannin). (F) PolyP-mediated overexpression of cyclin D1 in the absence and presence of increasing concentrations of AKT inhibitor (AKT inhibitor VIII). (G) PolyP-mediated overexpression of cyclin D1 in the absence and presence of increasing concentrations of intracellular calcium chelator (BAPTA-AM). (H) PolyP-mediated up-regulation of cyclin D1 in the absence and presence of increasing concentrations of PLC inhibitor (U-73122). (I) PolyP-mediated overexpression of cyclin D1 in the absence and presence of increasing concentrations of PKC inhibitor (BIS). (J) EA.hy926 cells were transiently transfected with control siRNA or siRNA for ERK1/2 and the efficiency of gene knockdown ( > 75%) was determined 48h post transfection by Western-blotting using a specific antibody. (K) The same as J except that polyP-mediated overexpression of cyclin D1 was monitored after siRNA knockdown of ERK1/2. (L) PolyP-mediated overexpression of cyclin D1 in the absence and presence of increasing concentrations of ERK inhibitor (PD-98059). The results are shown as mean ± standard deviation of 3 different experiments. *P
    β Catenin, supplied by Horizon Discovery, used in various techniques. Bioz Stars score: 99/100, based on 269 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
    https://www.bioz.com/result/β catenin/product/Horizon Discovery
    Average 99 stars, based on 269 article reviews
    Price from $9.99 to $1999.99
    β catenin - by Bioz Stars, 2020-03
    99/100 stars
      Buy from Supplier

    92
    Biomol GmbH β catenin
    Figure 5 Immunoblots for <t>β‐catenin</t> on Triton‐soluble and Triton‐insoluble fractions of HT29 cells cocultured with Helicobacter pylori . A reduction in band intensity is seen for the Triton‐insoluble (membrane‐bound)
    β Catenin, supplied by Biomol GmbH, used in various techniques. Bioz Stars score: 92/100, based on 23 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
    https://www.bioz.com/result/β catenin/product/Biomol GmbH
    Average 92 stars, based on 23 article reviews
    Price from $9.99 to $1999.99
    β catenin - by Bioz Stars, 2020-03
    92/100 stars
      Buy from Supplier

    99
    Cell Marque β catenin
    Immunohistochemistry of human CU-ACC2 tumor and CU-ACC2 PDX. A . and B . The left columns shows CU-ACC2 human tumor sample and the right two column is from CU-ACC2 PDX. A . The immunochemistry stains include H E, SF1, α-inhibin, Melan-A, Ki-67 and B . <t>β-catenin,</t> p53 and MSH2 C . Immunocytochemistry for Ki67 and SF1 (right columns) for CU-ACC2 cells (DAPI left column).
    β Catenin, supplied by Cell Marque, used in various techniques. Bioz Stars score: 99/100, based on 106 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
    https://www.bioz.com/result/β catenin/product/Cell Marque
    Average 99 stars, based on 106 article reviews
    Price from $9.99 to $1999.99
    β catenin - by Bioz Stars, 2020-03
    99/100 stars
      Buy from Supplier

    99
    GeneTex β catenin
    SJ26 suppressed cell migration in H1299, MCF7 and Hep3B 2.17 cells A. SJ26 suppressed <t>Wnt1/β-catenin-signaling</t> pathway in H1299, MCF7 and Hep3B 2.17 cells. Cells were treated with 10 μM SJ26 and the total cell extracts were prepared. Immunoblotting analysis was conducted using antibodies against Wnt1, β-catenin, or α-tubulin. B. H1299, MCF7 or Hep3B 2.17 cells were incubated with 10 μM of SJ26 and were subjected to trans-well assay analysis. The extent of cell migration across the wells was recorded after 6 hours. The value of migration rate in DMSO-treated cells was defined as 1. Asterisks indicate p
    β Catenin, supplied by GeneTex, used in various techniques. Bioz Stars score: 99/100, based on 69 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
    https://www.bioz.com/result/β catenin/product/GeneTex
    Average 99 stars, based on 69 article reviews
    Price from $9.99 to $1999.99
    β catenin - by Bioz Stars, 2020-03
    99/100 stars
      Buy from Supplier

    99
    Genechem β catenin
    EVI1 regulated the <t>Wnt/β-catenin</t> signaling pathway in NPC. ( a ) EVI1 downregulation inhibited β-catenin expression in the total cell lysate and the nuclear fraction. In contrast, forced expression of EVI1 enhanced β-catenin expression in the total cell lysate and the nuclear fraction. ( b ) The Wnt/β-catenin activity was repressed when EVI1 was knocked down as revealed by a TOP/FOP Flash reporter assay (left panel). EVI1 elevated Wnt/β-catenin activity when EVI1 was overexpressed (right panel). ( c ) The expression levels of β-catenin downstream targets AXIN2 and c-myc were downregulated in sh-EVI1 cells and were upregulated in LV-EVI1 cells. ( d ) The β-catenin promoter region contains one putative EVI1-binding site. ( e ) A ChIP assay demonstrated that EVI1 bound to the β-catenin promoter. ( f ) EVI1 positively regulated the β-catenin promoter activity
    β Catenin, supplied by Genechem, used in various techniques. Bioz Stars score: 99/100, based on 68 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
    https://www.bioz.com/result/β catenin/product/Genechem
    Average 99 stars, based on 68 article reviews
    Price from $9.99 to $1999.99
    β catenin - by Bioz Stars, 2020-03
    99/100 stars
      Buy from Supplier

    98
    Addgene inc β catenin
    Rhein inhibits <t>β‐catenin</t> expression in tumour xenografts. ( A ) Representative tumour tissue sections (100 × magnification) stained for β‐catenin by immunohistochemistry. ( B ) Tumour lysates were subjected to Western blot analysis of β‐catenin. The blots were subjected to densitometric analysis and relative quantification. ** P
    β Catenin, supplied by Addgene inc, used in various techniques. Bioz Stars score: 98/100, based on 267 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
    https://www.bioz.com/result/β catenin/product/Addgene inc
    Average 98 stars, based on 267 article reviews
    Price from $9.99 to $1999.99
    β catenin - by Bioz Stars, 2020-03
    98/100 stars
      Buy from Supplier

    91
    Assay Designs Inc β catenin
    Partial rescue of the cell fate change in Osr2-IresCre;Smad4 fl/fl dental mesenchyme using an exogenous WNT pathway inhibitor in vitro. ( A-C′ ) Immunofluorescence of <t>β-catenin</t> (red) in Smad4 fl/fl dental mesenchymal cells after adenoviral
    β Catenin, supplied by Assay Designs Inc, used in various techniques. Bioz Stars score: 91/100, based on 11 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
    https://www.bioz.com/result/β catenin/product/Assay Designs Inc
    Average 91 stars, based on 11 article reviews
    Price from $9.99 to $1999.99
    β catenin - by Bioz Stars, 2020-03
    91/100 stars
      Buy from Supplier

    98
    VANGL2 LTD β catenin
    Decreased <t>β-catenin</t> and Vangl2 levels are observed in Atmin Gpg6/Gpg6 E13.5 embryonic kidneys compared with WT littermates. Kidney sections from WT ( A and C ) and Atmin Gpg6/Gpg6 ( B and D ) embryos were immunostained for β-catenin (A and B) and Vangl2 (C and D). β-Catenin immunostaining was significantly reduced in Atmin Gpg6/Gpg6 (B) compared with WT (A). Vangl2 immunostaining was also dramatically decreased and apical enrichment was altered in Atmin Gpg6/Gpg6 (D) compared with WT (C). Overlays of DAPI and Vangl2 localization emphasize the loss of apical enrichment of Vangl2 protein in Atmin Gpg6/Gpg6 (D inset) versus WT (C inset; n = 4). All images are maximum projections of confocal z stacks taken at 0.4 µm intervals. Images were taken in identical conditions and post-acquisition manipulations were identical. Western blotting revealed a 0.4-fold decrease ( I ) in β-catenin ( P
    β Catenin, supplied by VANGL2 LTD, used in various techniques. Bioz Stars score: 98/100, based on 153 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
    https://www.bioz.com/result/β catenin/product/VANGL2 LTD
    Average 98 stars, based on 153 article reviews
    Price from $9.99 to $1999.99
    β catenin - by Bioz Stars, 2020-03
    98/100 stars
      Buy from Supplier

    99
    GenePharma Company β catenin
    SPAG5 regulates SCARA5 expression through the <t>wnt/β-catenin</t> pathway in HCC cells. a , Western blot and qRT-PCR analyses were used to detect SCARA5 expression in Huh7 cells stably transfected with the shNC or the shβ-catenin plasmid. * p
    β Catenin, supplied by GenePharma Company, used in various techniques. Bioz Stars score: 99/100, based on 143 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
    https://www.bioz.com/result/β catenin/product/GenePharma Company
    Average 99 stars, based on 143 article reviews
    Price from $9.99 to $1999.99
    β catenin - by Bioz Stars, 2020-03
    99/100 stars
      Buy from Supplier

    99
    Millipore β catenin
    Nuclear staining associated with mAb 8E7 is observed in <t>β-catenin</t> null cells . A B : Mouse L cell fibroblasts +/- Wnt3a A . Immunofluorescence staining of total (TL-) β-catenin (mAb clone 14, BD Biosciences) and ABC in mouse L cells stably expressing Wnt3a (right) or an empty vector (left). Note prominent nuclear staining in both Wnt3a expressing and non-expressing cells using 8E7 mAb. Exposure times for TL-β-catenin and ABC images are 1313 and 1439 milliseconds, respectively. B . Corresponding immuno-blot. Note that 8E7 detects non-specific band at ~160 kDa (*). C D : WT and β-catenin/plakoglobin null F9 teratocarcinoma cells immunostained with antibodies that recognize TL- and ABC. C . The split panel (upper right) shows the presence of β-catenin/plakoglobin null F9 cells on the coverslip (phase) that are not stained with the TL-β-catenin antibody, but show a diffuse cytoplasmic/nuclear stain with mAb 8E7 (lower right). Inset shows the GAM secondary antibody control. Exposure times for TL-β-catenin and ABC images are 228 and 453 milliseconds, respectively. D . Corresponding immunoblot shows that 8E7 detects a non-specific band at ~160 kDa in F9 cells (*). It is unclear whether this non-specific band contributes to background nuclear staining.
    β Catenin, supplied by Millipore, used in various techniques. Bioz Stars score: 99/100, based on 2365 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
    https://www.bioz.com/result/β catenin/product/Millipore
    Average 99 stars, based on 2365 article reviews
    Price from $9.99 to $1999.99
    β catenin - by Bioz Stars, 2020-03
    99/100 stars
      Buy from Supplier

    96
    BioLegend β catenin
    Analysis of intracellular components of Wnt pathway displayed downregulation of Wnt canonical pathway regulators in sFRP4 OE. There was a decrease in <t>β-catenin</t> as shown by immunocytochemical staining of β-catenin (red) in sFRP4 OE cells while there was an increase in nuclear β-catenin in sFRP4 SI cells (scale bar = 10 μm) ( A ). Western blot analysis showed a decrease in β-catenin and increase of GSK3β and APAF1 in sFRP4 OE cells ( B ), FURA-2AM analysis by flow cytometry showed an increase in intracellular calcium level in sFRP4 OE compared to sFRP4 SI cells ( C ).
    β Catenin, supplied by BioLegend, used in various techniques. Bioz Stars score: 96/100, based on 19 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
    https://www.bioz.com/result/β catenin/product/BioLegend
    Average 96 stars, based on 19 article reviews
    Price from $9.99 to $1999.99
    β catenin - by Bioz Stars, 2020-03
    96/100 stars
      Buy from Supplier

    92
    Pharmingen β catenin
    The interaction of Merlin with both <t>β-catenin</t> and Par3 is required for the establishment of functional junctions (A) IVTT-produced Merlin or Merlin 18-595 was mixed with either GST-β-catenin (top) or Myc-Par3 (middle). Complexes were isolated using either GST beads (top panel) or an anti-Myc antibody (middle panel) and immunoblotted with anti-GST, -Myc, or -Nf2 antibodies. Merlin 18-595 can readily bind Myc-Par3 but not GST-β-catenin in vitro . 10% of the input is shown in the bottom panel. (B) PAM212 keratinocytes were transfected with Myc-Par3, Myc-Par3 511-1266 or Myc-Par3 1-373 and cultured in calcium-containing media for the indicated times. Immunoprecipitation of Par3 followed by immunoblotting with Myc-, AJ- or TJ-specific antibodies revealed that full-length Myc-Par3 and Myc-Par3 511-1266 can associate with Merlin and with AJ and TJ proteins. In contrast, Myc-Par3 1-373 does not associate with Merlin or with AJ and TJ proteins. Note that Myc-Par3 511-1266 and Myc-Par3 1-373 are produced from a Par3 splice variant that does not contain the aPKC-binding site. NRS, normal rabbit serum. (C) TER was measured across calcium-stimulated PAM212 keratinocyte monolayers that stably express Merlin, Merlin 18-595 , or Myc-Par3 1-373 . Note that both Merlin 18-595 and Myc-Par3 1-373 dominantly interfered with the establishment of TER. Values = mean +/− SD. (D–G) Primary wild-type (D and F) or K14-Cre;Nf2 lox/lox (E and G) keratinocytes were stimulated with calcium-containing media for 8 hours, and either processed for indirect immunofluorescence (Fix 1, D and E) or subject to a more stringent in situ extraction (Fix 2, F and G). Immunostaining revealed normal localization of Par3 (green) at cell-cell boundaries in K14-Cre;Nf2 lox/lox .
    β Catenin, supplied by Pharmingen, used in various techniques. Bioz Stars score: 92/100, based on 12 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
    https://www.bioz.com/result/β catenin/product/Pharmingen
    Average 92 stars, based on 12 article reviews
    Price from $9.99 to $1999.99
    β catenin - by Bioz Stars, 2020-03
    92/100 stars
      Buy from Supplier

    96
    Leica Biosystems β catenin
    Expressions of signaling molecules of Wnt pathway, Wnt5b (A and B) and <t>β-catenin</t> (C and D). The expression of Wnt5b is observed in both tumor cells and CAFs, at the tumor surface (A) and the invasive margin (B). The increased expression and nuclear
    β Catenin, supplied by Leica Biosystems, used in various techniques. Bioz Stars score: 96/100, based on 17 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
    https://www.bioz.com/result/β catenin/product/Leica Biosystems
    Average 96 stars, based on 17 article reviews
    Price from $9.99 to $1999.99
    β catenin - by Bioz Stars, 2020-03
    96/100 stars
      Buy from Supplier

    96
    Epitomics β catenin
    Differentiation. HE staining of mammary tumors (A–B) and pulmonary metastases (C) from Apc +/1572T mice shows typical mammary glandular architecture and squamous differentiation. (D–F) Luminal epithelial differentiation as shown by cytokeratin 8 (Ck8) IHC staining. (G–I) Myoepithelial differentiation revealed by IHC staining with the Sma antibody. (J–L) IHC analysis with antibodies directed against cytokeratin 14 (Ck14) confirm the presence of squamous differentiation (hair follicle and skin cellular types). (M–O) <t>β-catenin</t> IHC analysis shows heterogeneous subcellular localization and intracellular accumulation with fewer cells characterized by positive nuclear staining. The results shown in this figure were confirmed in 12 independent primary tumors.
    β Catenin, supplied by Epitomics, used in various techniques. Bioz Stars score: 96/100, based on 116 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
    https://www.bioz.com/result/β catenin/product/Epitomics
    Average 96 stars, based on 116 article reviews
    Price from $9.99 to $1999.99
    β catenin - by Bioz Stars, 2020-03
    96/100 stars
      Buy from Supplier

    99
    Merck & Co β catenin
    GTP-Rac1 and pAKT did not participate in <t>TCF/β-catenin</t> signaling pathway after SLP-2 inhibition in NSCLC cells. a Western blot analyses for the expressions of pAkt in A549, H460, H838, and H157 cells treated with either AdSLP2i or AdCtrl at m.o.i. of 100. b Downregulation of SLP-2 increased the GTP-Rac1 proteins in A549 and H157 cells, but decreased the GTP-Rac1 proteins in H838 and H460 cells as determined by pull-down assay. Total cell lysates were subjected to the pull-down assay for GTP-Rac1 activities. Total cell lysates were analyzed for Rac1 expression as a loading control. The ratio of GTP-Rac1/total Rac1 in cells was analyzed by densitometry of the blot
    β Catenin, supplied by Merck & Co, used in various techniques. Bioz Stars score: 99/100, based on 14 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
    https://www.bioz.com/result/β catenin/product/Merck & Co
    Average 99 stars, based on 14 article reviews
    Price from $9.99 to $1999.99
    β catenin - by Bioz Stars, 2020-03
    99/100 stars
      Buy from Supplier

    97
    Novocastra β catenin
    Model of Egfr activity requirements during intestinal tumorigenesis. Red dots represent nuclear <t>β-catenin-positive</t> cells. Solid green lines indicate evidence for Egfr activity during establishment and adenocarcinoma expansion. Dashed green lines indicate that a requirement for Egfr activity during adenoma expansion, progression, and invasion has yet to be demonstrated conclusively.
    β Catenin, supplied by Novocastra, used in various techniques. Bioz Stars score: 97/100, based on 82 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
    https://www.bioz.com/result/β catenin/product/Novocastra
    Average 97 stars, based on 82 article reviews
    Price from $9.99 to $1999.99
    β catenin - by Bioz Stars, 2020-03
    97/100 stars
      Buy from Supplier

    99
    Merck KGaA β catenin
    Analysis of Akt and <t>Wnt/β-catenin</t> signaling in Tcl1- deficient and -overexpressing ES cells. (A) Western blot analysis of GSK, Akt, and β-catenin in wild-type (WT), Tcl1 −/− (KO) #2 and #4, Tcl1 −/− (CAG- Tcl1 ) #1 and #3, and Tcl1 −/− (CAG-EGFP) #5 ES cells. (B) Western blot analysis of active β-catenin, Oct3/4, and HSP90 in the cytoplasmic (C) and nuclear (N) fractions of wild-type (WT), Tcl1 −/− (KO) #2 and #4, and Tcl1 −/− (CAG- Tcl1 ) #10 and #1 ES cells. Tcl1 −/− (CAG- Tcl1 ) #10 and #1 were derived from Tcl1 −/− (KO) #2 and #4, respectively. Proper fractionation was confirmed by western blotting of Oct3/4 and HSP90, which localize to the nucleus and cytoplasm, respectively. Be8cause active β-catenin levels in the nuclear fractions were much lower than those in the cytoplasmic fractions, active β-catenin in the nuclear fractions was detected by approximately two-fold longer exposure compared with that in the cytoplasmic fractions. (C) TOPflash assay. P values of wild-type ES cells (WT) compared with Tcl1 −/− (CAG- Tcl1 ) #1 and #3 ES cells were less than 0.01. P values of Tcl1 −/− (KO) #4 and #5 ES cells compared with Tcl1 −/− (CAG- Tcl1 ) #1 and #3 ES cells were less than 0.02.
    β Catenin, supplied by Merck KGaA, used in various techniques. Bioz Stars score: 99/100, based on 55 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
    https://www.bioz.com/result/β catenin/product/Merck KGaA
    Average 99 stars, based on 55 article reviews
    Price from $9.99 to $1999.99
    β catenin - by Bioz Stars, 2020-03
    99/100 stars
      Buy from Supplier

    99
    Beyotime β catenin
    Effects of PA on the expression of E-cadherin, <t>β-catenin</t> and COX-2 in HO-8910 cells. After treatment with or without different concentrations of PA, corresponding proteins were determined at 48 h.
    β Catenin, supplied by Beyotime, used in various techniques. Bioz Stars score: 99/100, based on 102 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
    https://www.bioz.com/result/β catenin/product/Beyotime
    Average 99 stars, based on 102 article reviews
    Price from $9.99 to $1999.99
    β catenin - by Bioz Stars, 2020-03
    99/100 stars
      Buy from Supplier

    94
    Jackson Immuno β catenin
    Effects of PA on the expression of E-cadherin, <t>β-catenin</t> and COX-2 in HO-8910 cells. After treatment with or without different concentrations of PA, corresponding proteins were determined at 48 h.
    β Catenin, supplied by Jackson Immuno, used in various techniques. Bioz Stars score: 94/100, based on 29 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
    https://www.bioz.com/result/β catenin/product/Jackson Immuno
    Average 94 stars, based on 29 article reviews
    Price from $9.99 to $1999.99
    β catenin - by Bioz Stars, 2020-03
    94/100 stars
      Buy from Supplier

    97
    Sangon Biotech β catenin
    Up-regulation of <t>β-catenin</t> mediates 5-HT-induced PPAR γ reduction
    β Catenin, supplied by Sangon Biotech, used in various techniques. Bioz Stars score: 97/100, based on 31 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
    https://www.bioz.com/result/β catenin/product/Sangon Biotech
    Average 97 stars, based on 31 article reviews
    Price from $9.99 to $1999.99
    β catenin - by Bioz Stars, 2020-03
    97/100 stars
      Buy from Supplier

    99
    Ventana Medical β catenin
    Nodal LAM’s immunohistochemical expression of HMB45 is typically focal with a granular or gritty quality that fills just a portion of the cytoplasm (A, 400X). <t>β-catenin</t> consistently strongly and diffusely labels the cytoplasm of nodal
    β Catenin, supplied by Ventana Medical, used in various techniques. Bioz Stars score: 99/100, based on 95 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
    https://www.bioz.com/result/β catenin/product/Ventana Medical
    Average 99 stars, based on 95 article reviews
    Price from $9.99 to $1999.99
    β catenin - by Bioz Stars, 2020-03
    99/100 stars
      Buy from Supplier

    97
    Enzo Biochem β catenin
    Effect of galantamine and/or vildagliptin on <t>Wnt/β-catenin</t> pathway. Effect of different doses of galantamine (Galan 2.5, 5 10 mg/kg), vildagliptin (Vilda 3, 10 30 mg/kg) and their combination (Galan5 + Vilda 30) on the hepatic and muscular contents of p-GSK-3β (A, B) and β-catenin (C, D) in n-STZ diabetic rats. Drugs were gavaged orally for four weeks. Values are means of 10 rats ± S.E.M as compared with normal control (*), diabetic control (#), Galan5 (η), Galan10 (ɸ) and Vilda30 (Ψ)-treated groups (one-way ANOVA followed by Tukey post hoc test) at P
    β Catenin, supplied by Enzo Biochem, used in various techniques. Bioz Stars score: 97/100, based on 30 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
    https://www.bioz.com/result/β catenin/product/Enzo Biochem
    Average 97 stars, based on 30 article reviews
    Price from $9.99 to $1999.99
    β catenin - by Bioz Stars, 2020-03
    97/100 stars
      Buy from Supplier

    92
    Vector Biolabs β catenin
    D-DT-mediated <t>β-catenin</t> stabilization contributes to, but is not wholly responsible for, D-DT-dependent COX-2 expression A, Cells were plated in 6-well plates and were transfected the following day with pCDNA3.1/D-DT or pCDNA3.1/β-catenin
    β Catenin, supplied by Vector Biolabs, used in various techniques. Bioz Stars score: 92/100, based on 21 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
    https://www.bioz.com/result/β catenin/product/Vector Biolabs
    Average 92 stars, based on 21 article reviews
    Price from $9.99 to $1999.99
    β catenin - by Bioz Stars, 2020-03
    92/100 stars
      Buy from Supplier

    Image Search Results


    α-SYN is required for the maintenance, but not the generation, of adult subependymal NSCs. a , Representative SEZ whole-mount preparations of 2 month mice immunostained for γ-tubulin (blue), β-catenin (red), and GFAP (green). White dashed lines indicate examples of pinwheels. b , Pinwheel and B1 cell density per mm 2 at the anterodorsal (plain) and posterodorsal (striped) SEZ regions. Snca knock-out mice display lower density of pinwheels and B1 cells than wild-type littermates. c , Representative immunofluorescent detection of GFAP (red) and Ki67 (green) in the SEZ of 2 month wild-type ( Snca +/+ ) and mutant ( Snca −/− ) mice. White arrowhead points at GFAP + Ki67 + cells. d , Graphs showing the mean proportion (± SEM) of GFAP + cells relative to total number of cells (on the left axis) and GFAP + cells that are in cell cycle (Ki67 + ) or express S100β in the SEZ of Snca +/+ and Snca −/− mice at 2 months (on the right axis). e , Representative immunofluorescent detection of GFAP (red) and Ki67 (green) in the SEZ of postnatal day 7 mice. White arrowheads point at GFAP + Ki67 + cells. DAPI was used for nuclear staining. f , FACS analysis of qNSCs and aNSCs within the Lin − /CD24 −/low /GLAST + /CD9 high population discriminated by EGFR expression, showing a decrease in aNSCs in mutant mice. g , FACS analysis for CD9 in the Lin − /CD24 −/low /GLAST + population showing an apparent increase in niche CD9 low astrocytes in mutant mice. h , Graph representing the levels of CD9 expression in SEZ NSCs and in striatal astrocytes of wild-type mice and in cells of the mutant SEZ (orange line). * p

    Journal: The Journal of Neuroscience

    Article Title: Synaptic Regulator α-Synuclein in Dopaminergic Fibers Is Essentially Required for the Maintenance of Subependymal Neural Stem Cells

    doi: 10.1523/JNEUROSCI.2276-17.2017

    Figure Lengend Snippet: α-SYN is required for the maintenance, but not the generation, of adult subependymal NSCs. a , Representative SEZ whole-mount preparations of 2 month mice immunostained for γ-tubulin (blue), β-catenin (red), and GFAP (green). White dashed lines indicate examples of pinwheels. b , Pinwheel and B1 cell density per mm 2 at the anterodorsal (plain) and posterodorsal (striped) SEZ regions. Snca knock-out mice display lower density of pinwheels and B1 cells than wild-type littermates. c , Representative immunofluorescent detection of GFAP (red) and Ki67 (green) in the SEZ of 2 month wild-type ( Snca +/+ ) and mutant ( Snca −/− ) mice. White arrowhead points at GFAP + Ki67 + cells. d , Graphs showing the mean proportion (± SEM) of GFAP + cells relative to total number of cells (on the left axis) and GFAP + cells that are in cell cycle (Ki67 + ) or express S100β in the SEZ of Snca +/+ and Snca −/− mice at 2 months (on the right axis). e , Representative immunofluorescent detection of GFAP (red) and Ki67 (green) in the SEZ of postnatal day 7 mice. White arrowheads point at GFAP + Ki67 + cells. DAPI was used for nuclear staining. f , FACS analysis of qNSCs and aNSCs within the Lin − /CD24 −/low /GLAST + /CD9 high population discriminated by EGFR expression, showing a decrease in aNSCs in mutant mice. g , FACS analysis for CD9 in the Lin − /CD24 −/low /GLAST + population showing an apparent increase in niche CD9 low astrocytes in mutant mice. h , Graph representing the levels of CD9 expression in SEZ NSCs and in striatal astrocytes of wild-type mice and in cells of the mutant SEZ (orange line). * p

    Article Snippet: After blocking, sections were incubated in rabbit antibodies to α-SYN (1:1200, AbD Serotec), Ki67 (1:150, Abcam), β-catenin (1:100, Cell Signaling Technology), TH (1:600, Pel-Freez), β-III-tubulin (1:175, Sigma-Aldrich), S100β (1:100, Dako), or epidermal growth factor receptor (EGFR) (1:100, Cell Signaling Technology), mouse antibodies to S100β (1:500, Sigma-Aldrich), β-III-tubulin (1:200, Covance), Ki67 (1:50, Novocastra), α-SYN (1:1200, BD Biosciences), Ascl1 (1:150, BD Biosciences), Neu-N (1:100, Millipore), TH (1:500, Sigma-Aldrich), or human α-SYN (1:1350, Abcam, ab27766), rat antibodies to BrdU (1:800, Abcam), goat antibodies to Sox2 (1:150, R & D Systems), γ-tubulin (1:150, Santa Cruz Biotechnology), and chicken antibodies to GFAP (1:800, Millipore), alone or in different combinations for 24–48 h at 4°C.

    Techniques: Mouse Assay, Knock-Out, Mutagenesis, Staining, FACS, Expressing

    α-SYN is present in fibers supplying the SEZ. a , Immunofluorescent detection of α-SYN (green) and β-III-tubulin (red) in coronal sections through the anterior (a) and posterior (p) half of the RMS. White arrowheads point at doubly positive cells present only in the aRMS. DAPI was used as a counterstain. b , Immunofluorescent detection of α-SYN (red) and β-catenin (green) to label cell membranes in the SEZ in Snca +/+ and Snca −/− mice as a specificity control. White arrowheads point at puncta within the SEZ region. c , Synaptic distribution of α-SYN (red) observed by colocalization with synaptophysin + (green). d , Synaptic colocalization of α-SYN (green) with TH + (red). There is characteristic punctate staining of nerve terminals and the lack of staining in SEZ cells in the high-power micrographs (right) of the area indicated by the square. e , Low-power confocal micrographs showing the immunofluorescent detection of α-SYN (green), GFAP (red), and Sox2 (blue) and high-power images of the areas indicated by the white squares. α-SYN + terminals are closely adjacent to GFAP + Sox2 + cells (indicated by white arrowheads). f , Serial confocal sections through a SEZ whole mount at different z levels from the ventricular surface (z + 0) show the staining for β-catenin (blue), GFAP (red), and α-SYN (green). Dotted yellow lines indicate two pinwheels. Different GFAP + B1 cells traced in the z -axis from the ventricular surface with α-SYN + puncta closely apposed (white arrowheads) as shown at a higher magnification of the dotted white lined square. DAPI (blue) was used for nuclear staining. Scale bars: a , 20 μm; b–f , 10 μm.

    Journal: The Journal of Neuroscience

    Article Title: Synaptic Regulator α-Synuclein in Dopaminergic Fibers Is Essentially Required for the Maintenance of Subependymal Neural Stem Cells

    doi: 10.1523/JNEUROSCI.2276-17.2017

    Figure Lengend Snippet: α-SYN is present in fibers supplying the SEZ. a , Immunofluorescent detection of α-SYN (green) and β-III-tubulin (red) in coronal sections through the anterior (a) and posterior (p) half of the RMS. White arrowheads point at doubly positive cells present only in the aRMS. DAPI was used as a counterstain. b , Immunofluorescent detection of α-SYN (red) and β-catenin (green) to label cell membranes in the SEZ in Snca +/+ and Snca −/− mice as a specificity control. White arrowheads point at puncta within the SEZ region. c , Synaptic distribution of α-SYN (red) observed by colocalization with synaptophysin + (green). d , Synaptic colocalization of α-SYN (green) with TH + (red). There is characteristic punctate staining of nerve terminals and the lack of staining in SEZ cells in the high-power micrographs (right) of the area indicated by the square. e , Low-power confocal micrographs showing the immunofluorescent detection of α-SYN (green), GFAP (red), and Sox2 (blue) and high-power images of the areas indicated by the white squares. α-SYN + terminals are closely adjacent to GFAP + Sox2 + cells (indicated by white arrowheads). f , Serial confocal sections through a SEZ whole mount at different z levels from the ventricular surface (z + 0) show the staining for β-catenin (blue), GFAP (red), and α-SYN (green). Dotted yellow lines indicate two pinwheels. Different GFAP + B1 cells traced in the z -axis from the ventricular surface with α-SYN + puncta closely apposed (white arrowheads) as shown at a higher magnification of the dotted white lined square. DAPI (blue) was used for nuclear staining. Scale bars: a , 20 μm; b–f , 10 μm.

    Article Snippet: After blocking, sections were incubated in rabbit antibodies to α-SYN (1:1200, AbD Serotec), Ki67 (1:150, Abcam), β-catenin (1:100, Cell Signaling Technology), TH (1:600, Pel-Freez), β-III-tubulin (1:175, Sigma-Aldrich), S100β (1:100, Dako), or epidermal growth factor receptor (EGFR) (1:100, Cell Signaling Technology), mouse antibodies to S100β (1:500, Sigma-Aldrich), β-III-tubulin (1:200, Covance), Ki67 (1:50, Novocastra), α-SYN (1:1200, BD Biosciences), Ascl1 (1:150, BD Biosciences), Neu-N (1:100, Millipore), TH (1:500, Sigma-Aldrich), or human α-SYN (1:1350, Abcam, ab27766), rat antibodies to BrdU (1:800, Abcam), goat antibodies to Sox2 (1:150, R & D Systems), γ-tubulin (1:150, Santa Cruz Biotechnology), and chicken antibodies to GFAP (1:800, Millipore), alone or in different combinations for 24–48 h at 4°C.

    Techniques: Mouse Assay, Staining

    No effect of β-catenin deletion on baseline behaviors a, Social interaction (SI) in control, non-stressed animals (t 8 =0.840, P > 0.05, two tailed t-test, n=5/group). b, total distance traveled in arena (t 8 =0.251, P > 0.05, two tailed t-test, n=5/group). c, average velocity (t 8 =0.251, P > 0.05, two tailed t-test, n=5/group). Data is presented as mean and SEM. All data shown are representative of at least two experiments.

    Journal: Nature

    Article Title: β-catenin mediates behavioral resilience through Dicer1/microRNA regulation

    doi: 10.1038/nature13976

    Figure Lengend Snippet: No effect of β-catenin deletion on baseline behaviors a, Social interaction (SI) in control, non-stressed animals (t 8 =0.840, P > 0.05, two tailed t-test, n=5/group). b, total distance traveled in arena (t 8 =0.251, P > 0.05, two tailed t-test, n=5/group). c, average velocity (t 8 =0.251, P > 0.05, two tailed t-test, n=5/group). Data is presented as mean and SEM. All data shown are representative of at least two experiments.

    Article Snippet: The beads were centrifuged, and the supernatant was transferred to fresh tubes, where they were incubated with 5 µl of β-catenin antibody (Cell Signaling #9581) for one hour before 50 µl of a homogeneous protein G-agarose suspension was added and then incubated overnight at 4°C on a rotator.

    Techniques: Two Tailed Test

    β-catenin ChIP-seq in NAc 48 hours post CSDS a, qChIP validation of β-catenin ChIP (* P

    Journal: Nature

    Article Title: β-catenin mediates behavioral resilience through Dicer1/microRNA regulation

    doi: 10.1038/nature13976

    Figure Lengend Snippet: β-catenin ChIP-seq in NAc 48 hours post CSDS a, qChIP validation of β-catenin ChIP (* P

    Article Snippet: The beads were centrifuged, and the supernatant was transferred to fresh tubes, where they were incubated with 5 µl of β-catenin antibody (Cell Signaling #9581) for one hour before 50 µl of a homogeneous protein G-agarose suspension was added and then incubated overnight at 4°C on a rotator.

    Techniques: Chromatin Immunoprecipitation

    Validation of HSV-β-catenin a, β-catenin mRNA levels following HSV-β-catenin vs. HSV-GFP injection into NAc (t 4 =2.240, *P

    Journal: Nature

    Article Title: β-catenin mediates behavioral resilience through Dicer1/microRNA regulation

    doi: 10.1038/nature13976

    Figure Lengend Snippet: Validation of HSV-β-catenin a, β-catenin mRNA levels following HSV-β-catenin vs. HSV-GFP injection into NAc (t 4 =2.240, *P

    Article Snippet: The beads were centrifuged, and the supernatant was transferred to fresh tubes, where they were incubated with 5 µl of β-catenin antibody (Cell Signaling #9581) for one hour before 50 µl of a homogeneous protein G-agarose suspension was added and then incubated overnight at 4°C on a rotator.

    Techniques: Injection

    Other β-catenin manipulations a, Schematic of Cre-dependent HSV-lox-stop (LS1L)-β-catenin cassette. b, Validation of β-catenin knock down in the NAc of floxed β-catenin mice (t 7 =5.620, ***P

    Journal: Nature

    Article Title: β-catenin mediates behavioral resilience through Dicer1/microRNA regulation

    doi: 10.1038/nature13976

    Figure Lengend Snippet: Other β-catenin manipulations a, Schematic of Cre-dependent HSV-lox-stop (LS1L)-β-catenin cassette. b, Validation of β-catenin knock down in the NAc of floxed β-catenin mice (t 7 =5.620, ***P

    Article Snippet: The beads were centrifuged, and the supernatant was transferred to fresh tubes, where they were incubated with 5 µl of β-catenin antibody (Cell Signaling #9581) for one hour before 50 µl of a homogeneous protein G-agarose suspension was added and then incubated overnight at 4°C on a rotator.

    Techniques: Mouse Assay

    Regulation of β-catenin signaling in human depression and mouse CSDS a, mRNA from human NAc (Axin2: **P

    Journal: Nature

    Article Title: β-catenin mediates behavioral resilience through Dicer1/microRNA regulation

    doi: 10.1038/nature13976

    Figure Lengend Snippet: Regulation of β-catenin signaling in human depression and mouse CSDS a, mRNA from human NAc (Axin2: **P

    Article Snippet: The beads were centrifuged, and the supernatant was transferred to fresh tubes, where they were incubated with 5 µl of β-catenin antibody (Cell Signaling #9581) for one hour before 50 µl of a homogeneous protein G-agarose suspension was added and then incubated overnight at 4°C on a rotator.

    Techniques:

    Ingenuity pathway analysis (IPA) identifies a network of genes that show upregulated β-catenin binding at promoter regions in the NAc of resilient vs. susceptible mice Nodes represent differentially regulated genes, with green meaning up in resilient vs. susceptible and red meaning down in resilient vs. susceptible. The blue arrows indicate that the direction of regulation is consistent with IPA’s prediction of an upregulated β-catenin network in resilience; for example, a blue arrow means that a target gene that would be expected to be upregulated by β-catenin is in fact upregulated in this list. In contrast, yellow arrows indicate that the regulation observed is inconsistent with expectations, while gray arrows indicate lack of pre-existing data to formulate expectations of β-catenin action. Left panel shows mostly expected regulation of the β-catenin network (i.e., upregulation) in resilience; right panel shows non-specific changes occurring in a randomly chosen signal transducer and activator of transcription-4 (STAT4) network.

    Journal: Nature

    Article Title: β-catenin mediates behavioral resilience through Dicer1/microRNA regulation

    doi: 10.1038/nature13976

    Figure Lengend Snippet: Ingenuity pathway analysis (IPA) identifies a network of genes that show upregulated β-catenin binding at promoter regions in the NAc of resilient vs. susceptible mice Nodes represent differentially regulated genes, with green meaning up in resilient vs. susceptible and red meaning down in resilient vs. susceptible. The blue arrows indicate that the direction of regulation is consistent with IPA’s prediction of an upregulated β-catenin network in resilience; for example, a blue arrow means that a target gene that would be expected to be upregulated by β-catenin is in fact upregulated in this list. In contrast, yellow arrows indicate that the regulation observed is inconsistent with expectations, while gray arrows indicate lack of pre-existing data to formulate expectations of β-catenin action. Left panel shows mostly expected regulation of the β-catenin network (i.e., upregulation) in resilience; right panel shows non-specific changes occurring in a randomly chosen signal transducer and activator of transcription-4 (STAT4) network.

    Article Snippet: The beads were centrifuged, and the supernatant was transferred to fresh tubes, where they were incubated with 5 µl of β-catenin antibody (Cell Signaling #9581) for one hour before 50 µl of a homogeneous protein G-agarose suspension was added and then incubated overnight at 4°C on a rotator.

    Techniques: Indirect Immunoperoxidase Assay, Binding Assay, Mouse Assay

    Dicer1 bridges β-catenin and miRNA regulation in CSDS a, β-catenin ChIP-seq enrichment around the Dicer1 TSS. b, Effect of NAc Dicer1 knockdown (HSV-Cre) in sub-threshold defeat with HSV-GFP as control (**P

    Journal: Nature

    Article Title: β-catenin mediates behavioral resilience through Dicer1/microRNA regulation

    doi: 10.1038/nature13976

    Figure Lengend Snippet: Dicer1 bridges β-catenin and miRNA regulation in CSDS a, β-catenin ChIP-seq enrichment around the Dicer1 TSS. b, Effect of NAc Dicer1 knockdown (HSV-Cre) in sub-threshold defeat with HSV-GFP as control (**P

    Article Snippet: The beads were centrifuged, and the supernatant was transferred to fresh tubes, where they were incubated with 5 µl of β-catenin antibody (Cell Signaling #9581) for one hour before 50 µl of a homogeneous protein G-agarose suspension was added and then incubated overnight at 4°C on a rotator.

    Techniques: Chromatin Immunoprecipitation

    Repeated optogenetic burst stimulation of VTA cell bodies has no effect on canonical β-catenin signaling in NAc Experiment was performed as in Figure 2 with the exception of the optic fiber, which was placed above VTA for cell body stimulation (P > 0.05, two tailed t-test, n = 8/group). Data is presented as mean and SEM. Data is from one experiment.

    Journal: Nature

    Article Title: β-catenin mediates behavioral resilience through Dicer1/microRNA regulation

    doi: 10.1038/nature13976

    Figure Lengend Snippet: Repeated optogenetic burst stimulation of VTA cell bodies has no effect on canonical β-catenin signaling in NAc Experiment was performed as in Figure 2 with the exception of the optic fiber, which was placed above VTA for cell body stimulation (P > 0.05, two tailed t-test, n = 8/group). Data is presented as mean and SEM. Data is from one experiment.

    Article Snippet: The beads were centrifuged, and the supernatant was transferred to fresh tubes, where they were incubated with 5 µl of β-catenin antibody (Cell Signaling #9581) for one hour before 50 µl of a homogeneous protein G-agarose suspension was added and then incubated overnight at 4°C on a rotator.

    Techniques: Two Tailed Test

    β-catenin in NAc mediates pro-resilient, antidepressant, and anxiolytic responses a, IHC illustrating viral transgene expression mediated by HSV-β-catenin with coronal cartoon of NAc highlighted. b, Pro-resilient effect of HSV-β-catenin on social interaction after ASD (*P

    Journal: Nature

    Article Title: β-catenin mediates behavioral resilience through Dicer1/microRNA regulation

    doi: 10.1038/nature13976

    Figure Lengend Snippet: β-catenin in NAc mediates pro-resilient, antidepressant, and anxiolytic responses a, IHC illustrating viral transgene expression mediated by HSV-β-catenin with coronal cartoon of NAc highlighted. b, Pro-resilient effect of HSV-β-catenin on social interaction after ASD (*P

    Article Snippet: The beads were centrifuged, and the supernatant was transferred to fresh tubes, where they were incubated with 5 µl of β-catenin antibody (Cell Signaling #9581) for one hour before 50 µl of a homogeneous protein G-agarose suspension was added and then incubated overnight at 4°C on a rotator.

    Techniques: Immunohistochemistry, Expressing

    Regulation of β-catenin signaling in human depression and after CSDS in mice a, Axin2 expression is suppressed in both medicated and unmedicated depressed patients, both groups of which were clinically depressed at their time of death (F 2,13 =7.425, P

    Journal: Nature

    Article Title: β-catenin mediates behavioral resilience through Dicer1/microRNA regulation

    doi: 10.1038/nature13976

    Figure Lengend Snippet: Regulation of β-catenin signaling in human depression and after CSDS in mice a, Axin2 expression is suppressed in both medicated and unmedicated depressed patients, both groups of which were clinically depressed at their time of death (F 2,13 =7.425, P

    Article Snippet: The beads were centrifuged, and the supernatant was transferred to fresh tubes, where they were incubated with 5 µl of β-catenin antibody (Cell Signaling #9581) for one hour before 50 µl of a homogeneous protein G-agarose suspension was added and then incubated overnight at 4°C on a rotator.

    Techniques: Mouse Assay, Expressing

    Knockdown of Axin can counteract the growth inhibition induced by NAMPT depletion via Wnt/β-catenin signaling activation. a Western blot analysis of whole cell lysates showed the protein levels of Axin from LoVo cells exposed to 2% DMSO, FK866 (10 nM), or NMN (100 μM) + FK866 (10 nM) for 2 days. The data are presented as the mean ± SD of three independent experiments. Student’s t-test was used for statistical analysis. b Western blot analysis of whole cell lysates showed the protein levels of Axin from HCT116 cells exposed to 2% DMSO, FK866 (10 nM), or NMN (100 μM) + FK866 (10 nM) for 2 days. The data are presented as the mean ± SD of three independent experiments. Student’s t-test was used for statistical analysis. c The expression of Axin in LoVo cells with shNAMPT or shCtrl was analyzed by western blotting. The data are presented as the mean ± SD of three independent experiments. Student’s t-test was used for statistical analysis. d The efficiency of Axin knockdown in LoVo cells was determined by western blot analysis. The data are presented as the mean ± SD of three independent experiments. Student’s t-test was used for statistical analysis. The expressions of β-catenin from whole cell lysates ( e ) and cytoplasmic lysates ( f ) of LoVo cells with shAxin + 2% DMSO, shCtrl + 2% DMSO, shAxin + FK866, or shCtrl +FK866 were analyzed by western blotting. Student’s t-test was used for statistical analysis. g The viability of LoVo cells with shCtrl + 2% DMSO, shCtrl + FK866, shAxin + 2% DMSO, or shAxin + FK866 was analyzed using a CCK-8 assay. Student’s t-test was used for statistical analysis. h Overall survival (OS) analysis of CRC patients based on Axin expression was analyzed by Kaplan-Meier plotter. * P

    Journal: Cell Communication and Signaling : CCS

    Article Title: Targeting the NAD+ salvage pathway suppresses APC mutation-driven colorectal cancer growth and Wnt/β-catenin signaling via increasing Axin level

    doi: 10.1186/s12964-020-0513-5

    Figure Lengend Snippet: Knockdown of Axin can counteract the growth inhibition induced by NAMPT depletion via Wnt/β-catenin signaling activation. a Western blot analysis of whole cell lysates showed the protein levels of Axin from LoVo cells exposed to 2% DMSO, FK866 (10 nM), or NMN (100 μM) + FK866 (10 nM) for 2 days. The data are presented as the mean ± SD of three independent experiments. Student’s t-test was used for statistical analysis. b Western blot analysis of whole cell lysates showed the protein levels of Axin from HCT116 cells exposed to 2% DMSO, FK866 (10 nM), or NMN (100 μM) + FK866 (10 nM) for 2 days. The data are presented as the mean ± SD of three independent experiments. Student’s t-test was used for statistical analysis. c The expression of Axin in LoVo cells with shNAMPT or shCtrl was analyzed by western blotting. The data are presented as the mean ± SD of three independent experiments. Student’s t-test was used for statistical analysis. d The efficiency of Axin knockdown in LoVo cells was determined by western blot analysis. The data are presented as the mean ± SD of three independent experiments. Student’s t-test was used for statistical analysis. The expressions of β-catenin from whole cell lysates ( e ) and cytoplasmic lysates ( f ) of LoVo cells with shAxin + 2% DMSO, shCtrl + 2% DMSO, shAxin + FK866, or shCtrl +FK866 were analyzed by western blotting. Student’s t-test was used for statistical analysis. g The viability of LoVo cells with shCtrl + 2% DMSO, shCtrl + FK866, shAxin + 2% DMSO, or shAxin + FK866 was analyzed using a CCK-8 assay. Student’s t-test was used for statistical analysis. h Overall survival (OS) analysis of CRC patients based on Axin expression was analyzed by Kaplan-Meier plotter. * P

    Article Snippet: The primary antibodies used for western blot include the following: anti-NAMPT (Cell Signaling Technology, # 61122), anti-GAPDH (Cell Signaling Technology, # 5174), anti-β-catenin (Cell Signaling Technology, # 8480), anti-cyclin D1 (Cell Signaling Technology, # 2978), and anti-Axin (Cell Signaling Technology, # 2087).

    Techniques: Inhibition, Activation Assay, Western Blot, Expressing, CCK-8 Assay

    Model of NAMPT’s role in the proliferation of human CRC cells through the regulation of Wnt/β-catenin signaling. A high level of NAMPT (left panel) yields more NAD + and induces downregulation of Axin and the nuclear translocation of β-catenin, thereby leading to the activation of Wnt/β-catenin signaling and the increased growth of CRC cells. However, when NAMPT is inhibited by genetic or pharmacological methods (right panel), the NAD + level is reduced and Axin is upregulated. Then, β-catenin is degraded, and the Wnt/β-catenin pathway is suppressed, resulting in the inhibition of cancer cell proliferation

    Journal: Cell Communication and Signaling : CCS

    Article Title: Targeting the NAD+ salvage pathway suppresses APC mutation-driven colorectal cancer growth and Wnt/β-catenin signaling via increasing Axin level

    doi: 10.1186/s12964-020-0513-5

    Figure Lengend Snippet: Model of NAMPT’s role in the proliferation of human CRC cells through the regulation of Wnt/β-catenin signaling. A high level of NAMPT (left panel) yields more NAD + and induces downregulation of Axin and the nuclear translocation of β-catenin, thereby leading to the activation of Wnt/β-catenin signaling and the increased growth of CRC cells. However, when NAMPT is inhibited by genetic or pharmacological methods (right panel), the NAD + level is reduced and Axin is upregulated. Then, β-catenin is degraded, and the Wnt/β-catenin pathway is suppressed, resulting in the inhibition of cancer cell proliferation

    Article Snippet: The primary antibodies used for western blot include the following: anti-NAMPT (Cell Signaling Technology, # 61122), anti-GAPDH (Cell Signaling Technology, # 5174), anti-β-catenin (Cell Signaling Technology, # 8480), anti-cyclin D1 (Cell Signaling Technology, # 2978), and anti-Axin (Cell Signaling Technology, # 2087).

    Techniques: Translocation Assay, Activation Assay, Inhibition

    Modulation of the Wnt/β-catenin signaling pathway caused by NAMPT inhibition are NMN-dependent. a TOPFlash assay of LoVo or HCT116 cells treated with 2% DMSO, FK866 (10 nM), or NMN (100 μM) + FK866 (10 nM) for 2 days. The data are presented as the mean ± SD of three independent experiments. Student’s t-test was used for statistical analysis. b The expression of β-catenin and cyclin D1 in LoVo cells with shCtrl + 2% DMSO, shNAMPT + 2% DMSO, or shNAMPT + NMN (100 μM) was analyzed by western blotting. The data are presented as the mean ± SD of three independent experiments. Student’s t-test was used for statistical analysis. c Western blot analysis of whole cell lysates from LoVo cells exposed to 2% DMSO, FK866 (10 nM), or NMN (100 μM) + FK866 (10 nM) for 2 days showed the levels of β-catenin and cyclin D1. The data are presented as the mean ± SD of three independent experiments. Student’s t-test was used for statistical analysis. d Western blot analysis of whole cell lysates from HCT116 cells exposed to 2% DMSO, FK866 (10 nM), or NMN (100 μM) + FK866 (10 nM) for 2 days showed the levels of β-catenin and cyclin D1. The data are presented as the mean ± SD of three independent experiments. Student’s t-test was used for statistical analysis. e The cytoplasmic and nuclear expressions of β-catenin in LoVo cells treated with 2% DMSO, FK866 (10 nM), or NMN (100 μM) + FK866 (10 nM) for 2 days were analyzed by western blotting. The data are presented as the mean ± SD of three independent experiments. Student’s t-test was used for statistical analysis. f The cytoplasmic and nuclear expression of β-catenin in LoVo cells with shCtrl + 2% DMSO, shNAMPT + 2% DMSO, or shNAMPT + NMN (100 μM) was analyzed by western blotting. The data are presented as the mean ± SD of three independent experiments. Student’s t-test was used for statistical analysis. g The expression of β-catenin and cyclin D1 in LoVo cells with shCtrl + 2% DMSO, shNAMPT + 2% DMSO, shCtrl + CHIR99021 (100 ng/mL), or shNAMPT + CHIR99021 (100 ng/mL) was analyzed by western blotting. The data are presented as the mean ± SD of three independent experiments. Student’s t-test was used for statistical analysis. * P

    Journal: Cell Communication and Signaling : CCS

    Article Title: Targeting the NAD+ salvage pathway suppresses APC mutation-driven colorectal cancer growth and Wnt/β-catenin signaling via increasing Axin level

    doi: 10.1186/s12964-020-0513-5

    Figure Lengend Snippet: Modulation of the Wnt/β-catenin signaling pathway caused by NAMPT inhibition are NMN-dependent. a TOPFlash assay of LoVo or HCT116 cells treated with 2% DMSO, FK866 (10 nM), or NMN (100 μM) + FK866 (10 nM) for 2 days. The data are presented as the mean ± SD of three independent experiments. Student’s t-test was used for statistical analysis. b The expression of β-catenin and cyclin D1 in LoVo cells with shCtrl + 2% DMSO, shNAMPT + 2% DMSO, or shNAMPT + NMN (100 μM) was analyzed by western blotting. The data are presented as the mean ± SD of three independent experiments. Student’s t-test was used for statistical analysis. c Western blot analysis of whole cell lysates from LoVo cells exposed to 2% DMSO, FK866 (10 nM), or NMN (100 μM) + FK866 (10 nM) for 2 days showed the levels of β-catenin and cyclin D1. The data are presented as the mean ± SD of three independent experiments. Student’s t-test was used for statistical analysis. d Western blot analysis of whole cell lysates from HCT116 cells exposed to 2% DMSO, FK866 (10 nM), or NMN (100 μM) + FK866 (10 nM) for 2 days showed the levels of β-catenin and cyclin D1. The data are presented as the mean ± SD of three independent experiments. Student’s t-test was used for statistical analysis. e The cytoplasmic and nuclear expressions of β-catenin in LoVo cells treated with 2% DMSO, FK866 (10 nM), or NMN (100 μM) + FK866 (10 nM) for 2 days were analyzed by western blotting. The data are presented as the mean ± SD of three independent experiments. Student’s t-test was used for statistical analysis. f The cytoplasmic and nuclear expression of β-catenin in LoVo cells with shCtrl + 2% DMSO, shNAMPT + 2% DMSO, or shNAMPT + NMN (100 μM) was analyzed by western blotting. The data are presented as the mean ± SD of three independent experiments. Student’s t-test was used for statistical analysis. g The expression of β-catenin and cyclin D1 in LoVo cells with shCtrl + 2% DMSO, shNAMPT + 2% DMSO, shCtrl + CHIR99021 (100 ng/mL), or shNAMPT + CHIR99021 (100 ng/mL) was analyzed by western blotting. The data are presented as the mean ± SD of three independent experiments. Student’s t-test was used for statistical analysis. * P

    Article Snippet: The primary antibodies used for western blot include the following: anti-NAMPT (Cell Signaling Technology, # 61122), anti-GAPDH (Cell Signaling Technology, # 5174), anti-β-catenin (Cell Signaling Technology, # 8480), anti-cyclin D1 (Cell Signaling Technology, # 2978), and anti-Axin (Cell Signaling Technology, # 2087).

    Techniques: Inhibition, TOPFlash assay, Expressing, Western Blot

    miR-223 targets the Mef2 c-β-catenin pathway during breast cancer cell invasion . (A) miR-223 transfection inhibited Mef2c gene expression. HEK-293T cells were co-transfected with miR-223 and the Mef2 c 3'-UTR-directed luciferase reporter (luciferase-Mef2c; Mef2c 3'-UTR cloned into the pMIR-REPORTER). Luciferase activities were measured as described in Figure 2. Cells transfected with miR-223 and luciferase vector, miR-NC and luciferase-Mef2c, or miR-223 and luciferase-miR-223C (miR-223 complementary sequence cloned in the 3'-UTR of luciferase reporter) were used as controls. Data are averages of triplicates from three independent experiments. * p

    Journal: Molecular Cancer

    Article Title: Microvesicles secreted by macrophages shuttle invasion-potentiating microRNAs into breast cancer cells

    doi: 10.1186/1476-4598-10-117

    Figure Lengend Snippet: miR-223 targets the Mef2 c-β-catenin pathway during breast cancer cell invasion . (A) miR-223 transfection inhibited Mef2c gene expression. HEK-293T cells were co-transfected with miR-223 and the Mef2 c 3'-UTR-directed luciferase reporter (luciferase-Mef2c; Mef2c 3'-UTR cloned into the pMIR-REPORTER). Luciferase activities were measured as described in Figure 2. Cells transfected with miR-223 and luciferase vector, miR-NC and luciferase-Mef2c, or miR-223 and luciferase-miR-223C (miR-223 complementary sequence cloned in the 3'-UTR of luciferase reporter) were used as controls. Data are averages of triplicates from three independent experiments. * p

    Article Snippet: Confocal microscopy Cells prepared on coverslips were fixed in 4% PFA, treated with 0.3% Triton X-100, blocked with 5% BSA and incubated with an anti-β-catenin antibody (1:200, Cell Signaling Technology, #9587) overnight at 4°C.

    Techniques: Transfection, Expressing, Luciferase, Clone Assay, Plasmid Preparation, Sequencing

    β-catenin and pSmad2 coimmunoprecipitation in murine lungs following bleomycin injury.

    Journal:

    Article Title: Epithelial cell ?3?1 integrin links ?-catenin and Smad signaling to promote myofibroblast formation and pulmonary fibrosis

    doi: 10.1172/JCI36940

    Figure Lengend Snippet: β-catenin and pSmad2 coimmunoprecipitation in murine lungs following bleomycin injury.

    Article Snippet: Polyclonal β-catenin antibody was from Cell Signaling.

    Techniques:

    α3 integrin regulates association between β-catenin and pSmad2 ex vivo.

    Journal:

    Article Title: Epithelial cell ?3?1 integrin links ?-catenin and Smad signaling to promote myofibroblast formation and pulmonary fibrosis

    doi: 10.1172/JCI36940

    Figure Lengend Snippet: α3 integrin regulates association between β-catenin and pSmad2 ex vivo.

    Article Snippet: Polyclonal β-catenin antibody was from Cell Signaling.

    Techniques: Ex Vivo

    pY654–β-catenin/pSmad2 complexes in IPF lungs.

    Journal:

    Article Title: Epithelial cell ?3?1 integrin links ?-catenin and Smad signaling to promote myofibroblast formation and pulmonary fibrosis

    doi: 10.1172/JCI36940

    Figure Lengend Snippet: pY654–β-catenin/pSmad2 complexes in IPF lungs.

    Article Snippet: Polyclonal β-catenin antibody was from Cell Signaling.

    Techniques:

    Expression of β-catenin is upregulated in endometriosis. ( A ) HE staining of a normal endometrium. An endometrial gland and endometrial stromal cells are shown. ( B ) HE staining of endometriosis. ( C ) A representative image of immunohistochemical staining of a normal endometrium with anti-human β-catenin. ( D ) A representative image of immunohistochemical staining of endometriosis with anti-human β-catenin. ( E,F ) Significant upregulation of β-catenin protein expression in ECSC compared with NESC is shown by western blotting. n = 5, *p

    Journal: Scientific Reports

    Article Title: β-catenin signaling inhibitors ICG-001 and C-82 improve fibrosis in preclinical models of endometriosis

    doi: 10.1038/s41598-019-56302-4

    Figure Lengend Snippet: Expression of β-catenin is upregulated in endometriosis. ( A ) HE staining of a normal endometrium. An endometrial gland and endometrial stromal cells are shown. ( B ) HE staining of endometriosis. ( C ) A representative image of immunohistochemical staining of a normal endometrium with anti-human β-catenin. ( D ) A representative image of immunohistochemical staining of endometriosis with anti-human β-catenin. ( E,F ) Significant upregulation of β-catenin protein expression in ECSC compared with NESC is shown by western blotting. n = 5, *p

    Article Snippet: Antibodies against β-catenin (Cell Signaling, MA, USA), α-SMA (Gene Tex, Irvine, California, USA), and GAPDH (FUJIFILM Wako Chemical Corporation, Osaka, Japan) were used.

    Techniques: Expressing, Staining, Immunohistochemistry, Western Blot

    NF1-associated and sporadic MPNST cell lines show a decrease in cell viability, soft agar colony formation, and xenograft tumor growth when Wnt/β-catenin signaling is down-regulated

    Journal: Cancer discovery

    Article Title: Canonical Wnt/?-catenin Signaling Drives Human Schwann Cell Transformation, Progression, and Tumor Maintenance

    doi: 10.1158/2159-8290.CD-13-0081

    Figure Lengend Snippet: NF1-associated and sporadic MPNST cell lines show a decrease in cell viability, soft agar colony formation, and xenograft tumor growth when Wnt/β-catenin signaling is down-regulated

    Article Snippet: Cells were incubated in β-catenin primary antibody (1:100, Cell Signaling) at 4 degrees Celsius overnight, followed by 1 hour room temperature incubation in anti-rabbit AlexaFluor 488 secondary antibody (Invitrogen).

    Techniques:

    Human tissue microarray shows a subset of human neurofibromas and MPNSTs have activated Wnt/β-catenin signaling

    Journal: Cancer discovery

    Article Title: Canonical Wnt/?-catenin Signaling Drives Human Schwann Cell Transformation, Progression, and Tumor Maintenance

    doi: 10.1158/2159-8290.CD-13-0081

    Figure Lengend Snippet: Human tissue microarray shows a subset of human neurofibromas and MPNSTs have activated Wnt/β-catenin signaling

    Article Snippet: Cells were incubated in β-catenin primary antibody (1:100, Cell Signaling) at 4 degrees Celsius overnight, followed by 1 hour room temperature incubation in anti-rabbit AlexaFluor 488 secondary antibody (Invitrogen).

    Techniques: Microarray

    Immortalized human Schwann cells show an increase in transformed properties when Wnt/β-catenin signaling is activated

    Journal: Cancer discovery

    Article Title: Canonical Wnt/?-catenin Signaling Drives Human Schwann Cell Transformation, Progression, and Tumor Maintenance

    doi: 10.1158/2159-8290.CD-13-0081

    Figure Lengend Snippet: Immortalized human Schwann cells show an increase in transformed properties when Wnt/β-catenin signaling is activated

    Article Snippet: Cells were incubated in β-catenin primary antibody (1:100, Cell Signaling) at 4 degrees Celsius overnight, followed by 1 hour room temperature incubation in anti-rabbit AlexaFluor 488 secondary antibody (Invitrogen).

    Techniques: Transformation Assay

    The secreted Wnt/β-catenin activator RSPO2 is highly expressed in a subset of human Schwann cell tumors and can be detected as a fusion transcript with the upstream EIF3E gene

    Journal: Cancer discovery

    Article Title: Canonical Wnt/?-catenin Signaling Drives Human Schwann Cell Transformation, Progression, and Tumor Maintenance

    doi: 10.1158/2159-8290.CD-13-0081

    Figure Lengend Snippet: The secreted Wnt/β-catenin activator RSPO2 is highly expressed in a subset of human Schwann cell tumors and can be detected as a fusion transcript with the upstream EIF3E gene

    Article Snippet: Cells were incubated in β-catenin primary antibody (1:100, Cell Signaling) at 4 degrees Celsius overnight, followed by 1 hour room temperature incubation in anti-rabbit AlexaFluor 488 secondary antibody (Invitrogen).

    Techniques:

    Murine peripheral nerve tumors and human MPNST cell lines show an increase in nuclear β-catenin and Wnt pathway outputs

    Journal: Cancer discovery

    Article Title: Canonical Wnt/?-catenin Signaling Drives Human Schwann Cell Transformation, Progression, and Tumor Maintenance

    doi: 10.1158/2159-8290.CD-13-0081

    Figure Lengend Snippet: Murine peripheral nerve tumors and human MPNST cell lines show an increase in nuclear β-catenin and Wnt pathway outputs

    Article Snippet: Cells were incubated in β-catenin primary antibody (1:100, Cell Signaling) at 4 degrees Celsius overnight, followed by 1 hour room temperature incubation in anti-rabbit AlexaFluor 488 secondary antibody (Invitrogen).

    Techniques:

    Effect of miR-144/451 cluster onexpression of possible key proteins. a Expression of β-catenin. b Expression of c-Myc and phosphorylated cdc2. c Expression of MAPK/ERK pathway related proteins. d Expression of p53 and caspase3. e Expression of MMP9

    Journal: Cancer Cell International

    Article Title: miR-144/451 cluster plays an oncogenic role in esophageal cancer by inhibiting cell invasion

    doi: 10.1186/s12935-018-0679-8

    Figure Lengend Snippet: Effect of miR-144/451 cluster onexpression of possible key proteins. a Expression of β-catenin. b Expression of c-Myc and phosphorylated cdc2. c Expression of MAPK/ERK pathway related proteins. d Expression of p53 and caspase3. e Expression of MMP9

    Article Snippet: Antibodies and reagents PTEN (138G6) Rabbit Monoclonal antibody, Phospho-cdc2 (Tyr15) antibody, Phospho-p44/42 MAPK (ERK1/2) (Thr202/Tyr204) Rabbit Monoclonal antibody, p44/42 MAPK (Erk1/2) Rabbit Monoclonal antibody, Phospho-β-Catenin (Ser33/37/Thr41) antibody, Total-β-Catenin antibody, Phospho-c-Myc (Ser62) Rabbit Monoclonal antibody, p53 Rabbit Monoclonal antibody, Non-phospho (Active) β-Catenin (Ser33/37/Thr41) Rabbit Monoclonal antibody were obtained from Cell Signaling Technology (CST).

    Techniques: Expressing

    Upregulation of β-catenin protein levels and signaling in newborn Col2a1-Smurf2 chondrocytes through reduction of GSK-3β. (A–D) Examination of β-catenin levels in newborn WT (A, C) and Col2a1-Smurf2 (B, D) tibia by immunofluorescence

    Journal: Experimental cell research

    Article Title: Smurf2 Induces Degradation of GSK-3? and Upregulates ?-Catenin in Chondrocytes: A Potential Mechanism for Smurf2-Induced Degeneration of Articular Cartilage

    doi: 10.1016/j.yexcr.2009.05.019

    Figure Lengend Snippet: Upregulation of β-catenin protein levels and signaling in newborn Col2a1-Smurf2 chondrocytes through reduction of GSK-3β. (A–D) Examination of β-catenin levels in newborn WT (A, C) and Col2a1-Smurf2 (B, D) tibia by immunofluorescence

    Article Snippet: The antibodies used included primary anti-β-catenin (Cell Signaling Technology, Inc.), primary anti-type II collagen (Millpore), and secondary FITC conjugated donkey anti–rabbit (H & L) (Rockland Inc).

    Techniques: Immunofluorescence

    Upregulation of β-catenin protein levels in Col2a1-Smurf2 articular cartilage. (A–D) Examination of β-catenin levels in 2.5 week-old WT (A, C) and Col2a1-Smurf2 (B, D) femoral articular cartilage by immunofluorescence (A, B) and

    Journal: Experimental cell research

    Article Title: Smurf2 Induces Degradation of GSK-3? and Upregulates ?-Catenin in Chondrocytes: A Potential Mechanism for Smurf2-Induced Degeneration of Articular Cartilage

    doi: 10.1016/j.yexcr.2009.05.019

    Figure Lengend Snippet: Upregulation of β-catenin protein levels in Col2a1-Smurf2 articular cartilage. (A–D) Examination of β-catenin levels in 2.5 week-old WT (A, C) and Col2a1-Smurf2 (B, D) femoral articular cartilage by immunofluorescence (A, B) and

    Article Snippet: The antibodies used included primary anti-β-catenin (Cell Signaling Technology, Inc.), primary anti-type II collagen (Millpore), and secondary FITC conjugated donkey anti–rabbit (H & L) (Rockland Inc).

    Techniques: Immunofluorescence

    Beta-catenin is critically involved in the development of resistance to BRAFi. a) 451Lu-TetOn-shCTNNB1 cells were used for chronic treatment with 2 μM vemurafenib (1 μM during the first week) either in the presence (green and red curves) or absence (grey and black curves) of doxycyline (1 μg/ml doxycycline) and cell numbers were weekly counted by automated cell counting and viability assessment (CASY). Knockdown of β-catenin increased the time by approx. 2.5 fold until cell numbers exceeded the cell numbers of the maximum cell number in the first week. Morphology of the treated cells is shown after four weeks of cultivation with the indicated four different treatments (scale bar is 200 μm). b) The significantly (p

    Journal: EBioMedicine

    Article Title: A Nexus Consisting of Beta-Catenin and Stat3 Attenuates BRAF Inhibitor Efficacy and Mediates Acquired Resistance to Vemurafenib

    doi: 10.1016/j.ebiom.2016.04.037

    Figure Lengend Snippet: Beta-catenin is critically involved in the development of resistance to BRAFi. a) 451Lu-TetOn-shCTNNB1 cells were used for chronic treatment with 2 μM vemurafenib (1 μM during the first week) either in the presence (green and red curves) or absence (grey and black curves) of doxycyline (1 μg/ml doxycycline) and cell numbers were weekly counted by automated cell counting and viability assessment (CASY). Knockdown of β-catenin increased the time by approx. 2.5 fold until cell numbers exceeded the cell numbers of the maximum cell number in the first week. Morphology of the treated cells is shown after four weeks of cultivation with the indicated four different treatments (scale bar is 200 μm). b) The significantly (p

    Article Snippet: Immunohistochemistry staining of clinical FFPE specimens was performed with a β-catenin specific antibody (Cell Signaling #9562) diluted 1:50 in PBS containing 0.3% Triton-X100 and 1% BSA.

    Techniques: Cell Counting

    Beta-catenin IHC of a tissue microarray containing 270 melanoma biopsies. The slide was stained using a β-catenin specific antibody (Cell Signaling #9562 1:100). Staining intensities (IHC score) were judged by two experimenters from 0 (= absent) to 3(= strong) and grouped according tumor thickness and metastases. Kaplan Meier analysis was done using primary tumor data.

    Journal: EBioMedicine

    Article Title: A Nexus Consisting of Beta-Catenin and Stat3 Attenuates BRAF Inhibitor Efficacy and Mediates Acquired Resistance to Vemurafenib

    doi: 10.1016/j.ebiom.2016.04.037

    Figure Lengend Snippet: Beta-catenin IHC of a tissue microarray containing 270 melanoma biopsies. The slide was stained using a β-catenin specific antibody (Cell Signaling #9562 1:100). Staining intensities (IHC score) were judged by two experimenters from 0 (= absent) to 3(= strong) and grouped according tumor thickness and metastases. Kaplan Meier analysis was done using primary tumor data.

    Article Snippet: Immunohistochemistry staining of clinical FFPE specimens was performed with a β-catenin specific antibody (Cell Signaling #9562) diluted 1:50 in PBS containing 0.3% Triton-X100 and 1% BSA.

    Techniques: Immunohistochemistry, Microarray, Staining

    Accumulated β-catenin in BRAFi resistant melanoma cell lines acts independent of the canonical Wnt signaling pathway and the TCF/LEF factors. a) TOPflash luciferase reporter assays were done to measure the transcriptional activity of TCF/LEF complexes. Sensitive (black bars) and resistant (red bars) melanoma cell lines were transfected with the reporter construct plus CMV-renilla luciferase as a normalization control and treated for 24 h with the indicated concentrations of vemurafenib. For the induction of the full signaling activity a pre-treatment with 15 mM LiCl was performed. Firefly luciferase activity was normalized to renilla activity. Mean values and standard deviations of six samples are presented. Multiple t-tests with Holm–Šídák correction were used to compare data of sensitive and resistant samples and p

    Journal: EBioMedicine

    Article Title: A Nexus Consisting of Beta-Catenin and Stat3 Attenuates BRAF Inhibitor Efficacy and Mediates Acquired Resistance to Vemurafenib

    doi: 10.1016/j.ebiom.2016.04.037

    Figure Lengend Snippet: Accumulated β-catenin in BRAFi resistant melanoma cell lines acts independent of the canonical Wnt signaling pathway and the TCF/LEF factors. a) TOPflash luciferase reporter assays were done to measure the transcriptional activity of TCF/LEF complexes. Sensitive (black bars) and resistant (red bars) melanoma cell lines were transfected with the reporter construct plus CMV-renilla luciferase as a normalization control and treated for 24 h with the indicated concentrations of vemurafenib. For the induction of the full signaling activity a pre-treatment with 15 mM LiCl was performed. Firefly luciferase activity was normalized to renilla activity. Mean values and standard deviations of six samples are presented. Multiple t-tests with Holm–Šídák correction were used to compare data of sensitive and resistant samples and p

    Article Snippet: Immunohistochemistry staining of clinical FFPE specimens was performed with a β-catenin specific antibody (Cell Signaling #9562) diluted 1:50 in PBS containing 0.3% Triton-X100 and 1% BSA.

    Techniques: Luciferase, Activity Assay, Transfection, Construct

    Interaction of β-catenin with Stat3 occurs preferentially in BRAFi resistant melanoma cells. a) Co-immunoprecipitation experiments were done to confirm an interaction of β-catenin with Stat3. Lysates were prepared after crosslinking of the proteins with 0.4% buffered formaldehyde using the sensitive and resistant versions of 451Lu and Mel1617 cells. Precipitation was performed with a β-catenin or a Stat3 specific antibody and detected by immunoblot. Semi-quantification by densitometric analysis was done by calculating the ratios (bound fraction:input fraction) followed by normalization to the sensitive cells (see numbers below the blots and table). b) Immunoblot analysis for Stat3 expression and phosphorylation (Ser727 and Tyr705) in the sensitive and resistant 451Lu, Mel1617 and A375 cell lines. The same lysates as in Fig. 1 B were used. Beta-actin protein levels served as loading controls. c) Firefly reporter assay for Stat3 specific transciption was used to detect the nuclear Stat3 signaling activity in the sensitive and resistant melanoma cells of 451Lu, Mel1617 and A375. LiCl (15 mM for 24 h) treatment was used to induce β-catenin accumulation. Firefly signals were normalized to CMV-renilla control signals. Mean values +/− SD of sixtuplicates are shown. Multiple t-tests with Holm–Šídák correction were used to compare data of sensitive and resistant samples and p

    Journal: EBioMedicine

    Article Title: A Nexus Consisting of Beta-Catenin and Stat3 Attenuates BRAF Inhibitor Efficacy and Mediates Acquired Resistance to Vemurafenib

    doi: 10.1016/j.ebiom.2016.04.037

    Figure Lengend Snippet: Interaction of β-catenin with Stat3 occurs preferentially in BRAFi resistant melanoma cells. a) Co-immunoprecipitation experiments were done to confirm an interaction of β-catenin with Stat3. Lysates were prepared after crosslinking of the proteins with 0.4% buffered formaldehyde using the sensitive and resistant versions of 451Lu and Mel1617 cells. Precipitation was performed with a β-catenin or a Stat3 specific antibody and detected by immunoblot. Semi-quantification by densitometric analysis was done by calculating the ratios (bound fraction:input fraction) followed by normalization to the sensitive cells (see numbers below the blots and table). b) Immunoblot analysis for Stat3 expression and phosphorylation (Ser727 and Tyr705) in the sensitive and resistant 451Lu, Mel1617 and A375 cell lines. The same lysates as in Fig. 1 B were used. Beta-actin protein levels served as loading controls. c) Firefly reporter assay for Stat3 specific transciption was used to detect the nuclear Stat3 signaling activity in the sensitive and resistant melanoma cells of 451Lu, Mel1617 and A375. LiCl (15 mM for 24 h) treatment was used to induce β-catenin accumulation. Firefly signals were normalized to CMV-renilla control signals. Mean values +/− SD of sixtuplicates are shown. Multiple t-tests with Holm–Šídák correction were used to compare data of sensitive and resistant samples and p

    Article Snippet: Immunohistochemistry staining of clinical FFPE specimens was performed with a β-catenin specific antibody (Cell Signaling #9562) diluted 1:50 in PBS containing 0.3% Triton-X100 and 1% BSA.

    Techniques: Immunoprecipitation, Expressing, Reporter Assay, Activity Assay

    Knockdown of β-catenin synergizes with vemurafenib for the inhibition of tumor growth of BRAFi resistant melanomas. a) Xenograft experiment using Tet-inducible 451Lu-R cells for s.c. injection (1 × 10 6 cells) into SHO mice. After formation of 100 mm 3 tumor nodules upon daily i.p. injections with vemurafenib (25 mg/kg) the mice were randomized into four treatment groups: i) vemurafenib treatment (black), ii) untreated (grey), iii) shRNA induction by doxycycline (1 mg/ml) in the drinking water ad libitum and (green) iv) combination (red) (n = 5 per group). The tumor growth was daily monitored by caliper measurements and normalized to the size at day 0 of treatment. Multiple t tests were used to determine the data points of the combination group with p

    Journal: EBioMedicine

    Article Title: A Nexus Consisting of Beta-Catenin and Stat3 Attenuates BRAF Inhibitor Efficacy and Mediates Acquired Resistance to Vemurafenib

    doi: 10.1016/j.ebiom.2016.04.037

    Figure Lengend Snippet: Knockdown of β-catenin synergizes with vemurafenib for the inhibition of tumor growth of BRAFi resistant melanomas. a) Xenograft experiment using Tet-inducible 451Lu-R cells for s.c. injection (1 × 10 6 cells) into SHO mice. After formation of 100 mm 3 tumor nodules upon daily i.p. injections with vemurafenib (25 mg/kg) the mice were randomized into four treatment groups: i) vemurafenib treatment (black), ii) untreated (grey), iii) shRNA induction by doxycycline (1 mg/ml) in the drinking water ad libitum and (green) iv) combination (red) (n = 5 per group). The tumor growth was daily monitored by caliper measurements and normalized to the size at day 0 of treatment. Multiple t tests were used to determine the data points of the combination group with p

    Article Snippet: Immunohistochemistry staining of clinical FFPE specimens was performed with a β-catenin specific antibody (Cell Signaling #9562) diluted 1:50 in PBS containing 0.3% Triton-X100 and 1% BSA.

    Techniques: Inhibition, Injection, Mouse Assay, shRNA

    Beta-catenin expression levels increase in BRAFi resistant melanomas cells compared to the sensitive parentals. a) Immunohistochemical staining for β-catenin of clinical specimens before and after the acquisition of resistance to BRAFi. Beta-catenin expression levels are shown in red (Fast Red substrate) with hematoxylin counter staining (scale bar is 100 μm). b) Immunoblots of whole cell lysates from sensitive and resistant pairs of the melanoma cell lines 451Lu, Mel1617, A375 and SKMel19 showing the expression levels of β-catenin and (phospho) Erk1/2. Semiquantitative analysis was performed by building the ratios of (β-catenin:β-actin) and (p-ERk1/2:Erk1/2) and normalization to the sensitive cells.

    Journal: EBioMedicine

    Article Title: A Nexus Consisting of Beta-Catenin and Stat3 Attenuates BRAF Inhibitor Efficacy and Mediates Acquired Resistance to Vemurafenib

    doi: 10.1016/j.ebiom.2016.04.037

    Figure Lengend Snippet: Beta-catenin expression levels increase in BRAFi resistant melanomas cells compared to the sensitive parentals. a) Immunohistochemical staining for β-catenin of clinical specimens before and after the acquisition of resistance to BRAFi. Beta-catenin expression levels are shown in red (Fast Red substrate) with hematoxylin counter staining (scale bar is 100 μm). b) Immunoblots of whole cell lysates from sensitive and resistant pairs of the melanoma cell lines 451Lu, Mel1617, A375 and SKMel19 showing the expression levels of β-catenin and (phospho) Erk1/2. Semiquantitative analysis was performed by building the ratios of (β-catenin:β-actin) and (p-ERk1/2:Erk1/2) and normalization to the sensitive cells.

    Article Snippet: Immunohistochemistry staining of clinical FFPE specimens was performed with a β-catenin specific antibody (Cell Signaling #9562) diluted 1:50 in PBS containing 0.3% Triton-X100 and 1% BSA.

    Techniques: Expressing, Immunohistochemistry, Staining, Western Blot

    Beta-catenin is directly involved in the efficacy of the response to BRAFi and suppresses growth inhibition a) Cell viability assay (MUH) of melanoma cell lines pre-treated with 7.5 mM LiCl for 24 h for stabilization of β-catenin. Immunoblots show the stabilization of β-catenin after treatment of 451Lu cells with LiCl. After the pre-treatment, cells were treated with increasing concentrations of vemurafenib for 72 h before the assessment of cell viability. Black symbols represent sensitive control cells and red symbols represent LiCl pre-treated cells. Signals were normalized to the control cells without vemurafenib treatment. Mean +/ SD values of six replicates are shown. Multiple t-tests with Holm–Šídák correction were used to compare data points of the two curves and p

    Journal: EBioMedicine

    Article Title: A Nexus Consisting of Beta-Catenin and Stat3 Attenuates BRAF Inhibitor Efficacy and Mediates Acquired Resistance to Vemurafenib

    doi: 10.1016/j.ebiom.2016.04.037

    Figure Lengend Snippet: Beta-catenin is directly involved in the efficacy of the response to BRAFi and suppresses growth inhibition a) Cell viability assay (MUH) of melanoma cell lines pre-treated with 7.5 mM LiCl for 24 h for stabilization of β-catenin. Immunoblots show the stabilization of β-catenin after treatment of 451Lu cells with LiCl. After the pre-treatment, cells were treated with increasing concentrations of vemurafenib for 72 h before the assessment of cell viability. Black symbols represent sensitive control cells and red symbols represent LiCl pre-treated cells. Signals were normalized to the control cells without vemurafenib treatment. Mean +/ SD values of six replicates are shown. Multiple t-tests with Holm–Šídák correction were used to compare data points of the two curves and p

    Article Snippet: Immunohistochemistry staining of clinical FFPE specimens was performed with a β-catenin specific antibody (Cell Signaling #9562) diluted 1:50 in PBS containing 0.3% Triton-X100 and 1% BSA.

    Techniques: Inhibition, Viability Assay, Western Blot

    Novel Interaction partners of β-catenin in BRAFi resistant melanoma cells. a) Mel1617-S and -R cells were used for preparing nuclear enriched lysates in RIPA buffer before performing an immunoprecipitation with 500 μg of lysate and β-catenin specific antibody. LC-MS/MS analysis was performed using the co-precipitated and trypsin digested proteins. In Mel1617-R cells highly enriched and nuclear localized candidates for interaction with β-catenin were selected using DAVID and the raw intensity values plotted to compare the interaction in sensitive (black) versus resistant (red) Mel1617 melanoma cells. b) An in silico interactome analysis of β-catenin and Stat3 interaction partners was carried out using prePPI 58 . 148 putative shared interaction partners of β-catenin and Stat3 (Venn diagram) were found with a (prePPI-) p-value > 0.5 and sorted according their p-value products. A direct interaction of β-catenin with Stat3 was calculated with the probability p = 0.83.

    Journal: EBioMedicine

    Article Title: A Nexus Consisting of Beta-Catenin and Stat3 Attenuates BRAF Inhibitor Efficacy and Mediates Acquired Resistance to Vemurafenib

    doi: 10.1016/j.ebiom.2016.04.037

    Figure Lengend Snippet: Novel Interaction partners of β-catenin in BRAFi resistant melanoma cells. a) Mel1617-S and -R cells were used for preparing nuclear enriched lysates in RIPA buffer before performing an immunoprecipitation with 500 μg of lysate and β-catenin specific antibody. LC-MS/MS analysis was performed using the co-precipitated and trypsin digested proteins. In Mel1617-R cells highly enriched and nuclear localized candidates for interaction with β-catenin were selected using DAVID and the raw intensity values plotted to compare the interaction in sensitive (black) versus resistant (red) Mel1617 melanoma cells. b) An in silico interactome analysis of β-catenin and Stat3 interaction partners was carried out using prePPI 58 . 148 putative shared interaction partners of β-catenin and Stat3 (Venn diagram) were found with a (prePPI-) p-value > 0.5 and sorted according their p-value products. A direct interaction of β-catenin with Stat3 was calculated with the probability p = 0.83.

    Article Snippet: Immunohistochemistry staining of clinical FFPE specimens was performed with a β-catenin specific antibody (Cell Signaling #9562) diluted 1:50 in PBS containing 0.3% Triton-X100 and 1% BSA.

    Techniques: Immunoprecipitation, Liquid Chromatography with Mass Spectroscopy, Mass Spectrometry, In Silico

    Nuclear localized beta-catenin in BRAFi resistant melanomas cells compared to the sensitive parentals. a) Immunofluorescence staining for β-catenin (blue) and nuclear YOPRO-1 staining (green) with confocal microscopy for expression and localization analysis of β-catenin (white scale bars represent 50 μm). b) Immunoblot of cytosolic and nuclear extracts showing the increased nuclear localization of β-catenin in the resistant cells of the cell lines 451Lu, Mel1617 and A375. Semiquantitation was performed densitometrically by using the ratios of [β-catenin:β-actin] for cytosolic and [β-catenin:LaminB] for nuclear fractions of the sensitive and resistant cell line pairs. All ratios were normalized to the sensitive parental cell line to compare the sensitive to the corresponding resistant cell line.

    Journal: EBioMedicine

    Article Title: A Nexus Consisting of Beta-Catenin and Stat3 Attenuates BRAF Inhibitor Efficacy and Mediates Acquired Resistance to Vemurafenib

    doi: 10.1016/j.ebiom.2016.04.037

    Figure Lengend Snippet: Nuclear localized beta-catenin in BRAFi resistant melanomas cells compared to the sensitive parentals. a) Immunofluorescence staining for β-catenin (blue) and nuclear YOPRO-1 staining (green) with confocal microscopy for expression and localization analysis of β-catenin (white scale bars represent 50 μm). b) Immunoblot of cytosolic and nuclear extracts showing the increased nuclear localization of β-catenin in the resistant cells of the cell lines 451Lu, Mel1617 and A375. Semiquantitation was performed densitometrically by using the ratios of [β-catenin:β-actin] for cytosolic and [β-catenin:LaminB] for nuclear fractions of the sensitive and resistant cell line pairs. All ratios were normalized to the sensitive parental cell line to compare the sensitive to the corresponding resistant cell line.

    Article Snippet: Immunohistochemistry staining of clinical FFPE specimens was performed with a β-catenin specific antibody (Cell Signaling #9562) diluted 1:50 in PBS containing 0.3% Triton-X100 and 1% BSA.

    Techniques: Immunofluorescence, Staining, Confocal Microscopy, Expressing

    Wnt signaling is not majorly involved in resistance mechanisms to BRAFi. a) Bead-based GST-pull-down assay of “free” β-catenin using GST-tagged TCF4. Samples were prepared in triplicates after cultivation under the indicated conditions for 24 h. Signals were normalized to the total amount of β-catenin measured by a bead-based sandwich immunoassay and compared to the untreated controls. b) Heatmap of mRNA expression analysis using the Illumina beadchip-based microarray dataset GSE50509 dealing with 21 pairs of samples from melanoma patients treated with BRAFi 33 . Fold changes in gene expression of BRAFi treated and matched pre-treatment tumors were calculated for each patient using log2 transformed and RMA normalized signal intensities. Exemplarily the gene sets M223 (canonical Wnt signaling) and M31 (non-canonical Wnt signaling) from the Gene set enrichment analysis (GSEA) tool from the Broad Institute is shown which shows nuclear β-catenin signaling and target gene transcription. c) Cell viability assay of cells pre-treated with Wnt3a (100 ng/ml) before treatment with vemurafenib for 72 h reveals no effect of Wnt3a stimulation. After the pre-treatment, cells were re-stimulated with fresh Wnt3a when the BRAFi treatment was started. Black data represent unstimulated cells, red data show Wnt3a treated cells. d) Sensitive 451Lu-S, Mel1617-S and A375-S cells were pre-treated with Wnt5a (100ng/ml) and re-stimulated after 24 h. Vemurafenib treatment was applied for 72 h before measurement of the cell viability (MUH). No differences in susceptibility to BRAFi were found after treatment with Wnt3a or Wnt5a. e) Three different Gluc-reporter systems (AP1-Gluc, CRE-Gluc, NFAT-Gluc) were used to assay the non-canonical Wnt signaling activity in the sensitive and resistant versions of 451Lu, Mel1617 and A375 melanoma cells. After transfection with the reporter plasmids cells were treated for 24 h with 15 mM LiCl for stabilizing β-catenin. Secreted gaussia luciferase activity was measured and normalized to cytoplasmic firefly luciferase activity from a co-transfected SV40-fluc control plasmid. Assays were performed in sixtuplicates. Mean values and standard deviations of sixtuplicates are presented. Multiple t-tests with Holm–Šídák correction were used to compare data of sensitive and resistant samples and p

    Journal: EBioMedicine

    Article Title: A Nexus Consisting of Beta-Catenin and Stat3 Attenuates BRAF Inhibitor Efficacy and Mediates Acquired Resistance to Vemurafenib

    doi: 10.1016/j.ebiom.2016.04.037

    Figure Lengend Snippet: Wnt signaling is not majorly involved in resistance mechanisms to BRAFi. a) Bead-based GST-pull-down assay of “free” β-catenin using GST-tagged TCF4. Samples were prepared in triplicates after cultivation under the indicated conditions for 24 h. Signals were normalized to the total amount of β-catenin measured by a bead-based sandwich immunoassay and compared to the untreated controls. b) Heatmap of mRNA expression analysis using the Illumina beadchip-based microarray dataset GSE50509 dealing with 21 pairs of samples from melanoma patients treated with BRAFi 33 . Fold changes in gene expression of BRAFi treated and matched pre-treatment tumors were calculated for each patient using log2 transformed and RMA normalized signal intensities. Exemplarily the gene sets M223 (canonical Wnt signaling) and M31 (non-canonical Wnt signaling) from the Gene set enrichment analysis (GSEA) tool from the Broad Institute is shown which shows nuclear β-catenin signaling and target gene transcription. c) Cell viability assay of cells pre-treated with Wnt3a (100 ng/ml) before treatment with vemurafenib for 72 h reveals no effect of Wnt3a stimulation. After the pre-treatment, cells were re-stimulated with fresh Wnt3a when the BRAFi treatment was started. Black data represent unstimulated cells, red data show Wnt3a treated cells. d) Sensitive 451Lu-S, Mel1617-S and A375-S cells were pre-treated with Wnt5a (100ng/ml) and re-stimulated after 24 h. Vemurafenib treatment was applied for 72 h before measurement of the cell viability (MUH). No differences in susceptibility to BRAFi were found after treatment with Wnt3a or Wnt5a. e) Three different Gluc-reporter systems (AP1-Gluc, CRE-Gluc, NFAT-Gluc) were used to assay the non-canonical Wnt signaling activity in the sensitive and resistant versions of 451Lu, Mel1617 and A375 melanoma cells. After transfection with the reporter plasmids cells were treated for 24 h with 15 mM LiCl for stabilizing β-catenin. Secreted gaussia luciferase activity was measured and normalized to cytoplasmic firefly luciferase activity from a co-transfected SV40-fluc control plasmid. Assays were performed in sixtuplicates. Mean values and standard deviations of sixtuplicates are presented. Multiple t-tests with Holm–Šídák correction were used to compare data of sensitive and resistant samples and p

    Article Snippet: Immunohistochemistry staining of clinical FFPE specimens was performed with a β-catenin specific antibody (Cell Signaling #9562) diluted 1:50 in PBS containing 0.3% Triton-X100 and 1% BSA.

    Techniques: Pull Down Assay, Expressing, Microarray, Transformation Assay, Viability Assay, Activity Assay, Transfection, Luciferase, Plasmid Preparation

    Knockdown of β-catenin enhances the BRAFi mediated induction of apoptosis and senescence. a) Cell cycle analysis of Tet-inducible 451LuTet-S (upper panel) and 451LuTet-R (lower panel) cells after treatment with 5 or 10 μM vemurafenib for 72 h. Doxycycline pre-treatment for 24 h (right bar diagrams) was used to induce β-catenin specific shRNA. Representative FACS histograms of the cell cycle analysis comparing vehicle (DMSO) treated cells with vemurafenib (10 μM) are shown. Cell cycle distributions represent the result of three biologic replicates (mean +/− SD). b) SA-associated β-galactosidase staining after 72 h of vemurafenib treatment of the above mentioned Tet-inducible cells (scale bars represent 200 μm). Clear blue cells were counted and normalized to the total number of cells using four different microscopic pictures (top row) per treatment. Mean numbers +/− SD are shown from three independent biological replicates (bottom row). ANOVA analyis was performed using Tukey's multiple comparisons test (asterisks).

    Journal: EBioMedicine

    Article Title: A Nexus Consisting of Beta-Catenin and Stat3 Attenuates BRAF Inhibitor Efficacy and Mediates Acquired Resistance to Vemurafenib

    doi: 10.1016/j.ebiom.2016.04.037

    Figure Lengend Snippet: Knockdown of β-catenin enhances the BRAFi mediated induction of apoptosis and senescence. a) Cell cycle analysis of Tet-inducible 451LuTet-S (upper panel) and 451LuTet-R (lower panel) cells after treatment with 5 or 10 μM vemurafenib for 72 h. Doxycycline pre-treatment for 24 h (right bar diagrams) was used to induce β-catenin specific shRNA. Representative FACS histograms of the cell cycle analysis comparing vehicle (DMSO) treated cells with vemurafenib (10 μM) are shown. Cell cycle distributions represent the result of three biologic replicates (mean +/− SD). b) SA-associated β-galactosidase staining after 72 h of vemurafenib treatment of the above mentioned Tet-inducible cells (scale bars represent 200 μm). Clear blue cells were counted and normalized to the total number of cells using four different microscopic pictures (top row) per treatment. Mean numbers +/− SD are shown from three independent biological replicates (bottom row). ANOVA analyis was performed using Tukey's multiple comparisons test (asterisks).

    Article Snippet: Immunohistochemistry staining of clinical FFPE specimens was performed with a β-catenin specific antibody (Cell Signaling #9562) diluted 1:50 in PBS containing 0.3% Triton-X100 and 1% BSA.

    Techniques: Cell Cycle Assay, shRNA, FACS, Staining

    Increased resistance of melanoma cell lines after chronic treatment with 2 μM PLX4032 (vemurafenib). a) Cell viability assays (MUH) were performed after 72 h of treatment with increasing concentrations of vemurafenib. Black data points show the parental sensitive cells, red data points show the corresponding resistant cells. Assays were performed in sixtuplicates and data were normalized to the untreated control, mean values +/− SD are shown with the calculated IC 50 values vemurafenib. b) Cell cycle analysis of the sensitive and resistant melanoma cell lines using propidium iodide staining. c) Mutation analysis of hotspot mutations in the four cell line pairs Mel167, 451Lu, A375 and SKMel19 for BRAF, NRAS, CTNNB1 and MEK1. d) Western blot to detect the expression levels of BRAF in the four cell lines together with truncated splice variants of BRAF. e) Transcript expression (real-time qPCR) of genes known to be involved in resistance mechanisms and target genes of Wnt-/β-catenin signaling (MITF and TYR) and Stat3 (IL6). f) Immunoblots corresponding to Fig. 1 with additional signaling proteins. g) Immunoblots showing the expression of regulators of β-catenin expression (APC, Sfrp1, Axin-1) in the four cell line pairs.

    Journal: EBioMedicine

    Article Title: A Nexus Consisting of Beta-Catenin and Stat3 Attenuates BRAF Inhibitor Efficacy and Mediates Acquired Resistance to Vemurafenib

    doi: 10.1016/j.ebiom.2016.04.037

    Figure Lengend Snippet: Increased resistance of melanoma cell lines after chronic treatment with 2 μM PLX4032 (vemurafenib). a) Cell viability assays (MUH) were performed after 72 h of treatment with increasing concentrations of vemurafenib. Black data points show the parental sensitive cells, red data points show the corresponding resistant cells. Assays were performed in sixtuplicates and data were normalized to the untreated control, mean values +/− SD are shown with the calculated IC 50 values vemurafenib. b) Cell cycle analysis of the sensitive and resistant melanoma cell lines using propidium iodide staining. c) Mutation analysis of hotspot mutations in the four cell line pairs Mel167, 451Lu, A375 and SKMel19 for BRAF, NRAS, CTNNB1 and MEK1. d) Western blot to detect the expression levels of BRAF in the four cell lines together with truncated splice variants of BRAF. e) Transcript expression (real-time qPCR) of genes known to be involved in resistance mechanisms and target genes of Wnt-/β-catenin signaling (MITF and TYR) and Stat3 (IL6). f) Immunoblots corresponding to Fig. 1 with additional signaling proteins. g) Immunoblots showing the expression of regulators of β-catenin expression (APC, Sfrp1, Axin-1) in the four cell line pairs.

    Article Snippet: Immunohistochemistry staining of clinical FFPE specimens was performed with a β-catenin specific antibody (Cell Signaling #9562) diluted 1:50 in PBS containing 0.3% Triton-X100 and 1% BSA.

    Techniques: Cell Cycle Assay, Staining, Mutagenesis, Western Blot, Expressing, Real-time Polymerase Chain Reaction

    Stat3 and β-catenin levels cooperatively mediate resistance to BRAFi in melanoma cells. a) Stat3 was overexpressed in sensitive A375 and SKMel19 cells using a lentivirus (LV-STAT3). After selection the stable overexpression was tested by western blot and the cells were used for cell viability testings (MUH) after 72 h of treatment with vemurafenib. Signals were normalized to the control cells without vemurafenib treatment. Mean +/− SD values of six replicates are shown. Multiple t-tests with Holm–Šídák correction were used to compare data points of the two curves and p

    Journal: EBioMedicine

    Article Title: A Nexus Consisting of Beta-Catenin and Stat3 Attenuates BRAF Inhibitor Efficacy and Mediates Acquired Resistance to Vemurafenib

    doi: 10.1016/j.ebiom.2016.04.037

    Figure Lengend Snippet: Stat3 and β-catenin levels cooperatively mediate resistance to BRAFi in melanoma cells. a) Stat3 was overexpressed in sensitive A375 and SKMel19 cells using a lentivirus (LV-STAT3). After selection the stable overexpression was tested by western blot and the cells were used for cell viability testings (MUH) after 72 h of treatment with vemurafenib. Signals were normalized to the control cells without vemurafenib treatment. Mean +/− SD values of six replicates are shown. Multiple t-tests with Holm–Šídák correction were used to compare data points of the two curves and p

    Article Snippet: Immunohistochemistry staining of clinical FFPE specimens was performed with a β-catenin specific antibody (Cell Signaling #9562) diluted 1:50 in PBS containing 0.3% Triton-X100 and 1% BSA.

    Techniques: Selection, Over Expression, Western Blot

    Proposed model of regulation of PME-1 expression by PI3K-GSK-3β signaling. Activation of PI3K signaling results in phosphorylation of GSK-3β and inhibition its activity in phosphorylating β-catenin. β-catenin translocates to nucleus and act as co-activator with LEF1/TCF to promote PME-1 expression, which catalyzes the demethylation of PP2Ac.

    Journal: Aging (Albany NY)

    Article Title: Glycogen synthase kinase-3β suppresses the expression of protein phosphatase methylesterase-1 through β-catenin

    doi: 10.18632/aging.102413

    Figure Lengend Snippet: Proposed model of regulation of PME-1 expression by PI3K-GSK-3β signaling. Activation of PI3K signaling results in phosphorylation of GSK-3β and inhibition its activity in phosphorylating β-catenin. β-catenin translocates to nucleus and act as co-activator with LEF1/TCF to promote PME-1 expression, which catalyzes the demethylation of PP2Ac.

    Article Snippet: Polyclonal anti-β-catenin, anti-Non-pS-β-catenin (dephosphorylated at S33/S37/T41), anti-GSK-3β and anti-pS9-GSK-3β were purchased from Cell Signaling Technology (Danvers, MA, USA).

    Techniques: Expressing, Activation Assay, Inhibition, Activity Assay, Activated Clotting Time Assay

    The promoter of human PME-1 contains two putative LEF1/TCF elements. ( A ) Human PME-1 promoter region has two potential LEF1/TCF like elements (red). The promoter (-1000 to +389) of human PME-1 was analyzed by MatInspector software. Cis -elements are labeled with different color. There are two potential LEF1/TCF like elements. Other cis -elements are NFAT (nuclear factor of activated T cells), E2F (transcription factor family including E2F- and DP-like subunits), c-myb (Cellular homologue of avian myeloblastosis virus oncogene). ( B ) The SH-SY5Y cells were overexpressed with β-catenin tagged with HA. Monoclonal anti-HA were used to immunoprecipitate β-catenin and co-immunoprecipitated LEF1/TCF cis-elements were amplified by PCR using primers specific to LEF1/TCF cis -elements 1 and 2. ( C ) Schematic diagram of PME-1 promoter and its luciferase reporter plasmid. Human PME-1 promoter was inserted into pGL4.10 containing luciferase reporter gene to generate pGL4/PME-1 -1000 . ( D ) pGL4/PME-1 -1000 and pGL4.10 showed in panel C were transfected into HEK-293T or SH-SY5Y cells. The Photinus pyralis luciferase activity and Renilla reniformis luciferase activity were measured subsequently and the Photinus pyralis luciferase activity was normalized with Renilla reniformis luciferase activity. Data are presented as mean ± SD (n=3); *P

    Journal: Aging (Albany NY)

    Article Title: Glycogen synthase kinase-3β suppresses the expression of protein phosphatase methylesterase-1 through β-catenin

    doi: 10.18632/aging.102413

    Figure Lengend Snippet: The promoter of human PME-1 contains two putative LEF1/TCF elements. ( A ) Human PME-1 promoter region has two potential LEF1/TCF like elements (red). The promoter (-1000 to +389) of human PME-1 was analyzed by MatInspector software. Cis -elements are labeled with different color. There are two potential LEF1/TCF like elements. Other cis -elements are NFAT (nuclear factor of activated T cells), E2F (transcription factor family including E2F- and DP-like subunits), c-myb (Cellular homologue of avian myeloblastosis virus oncogene). ( B ) The SH-SY5Y cells were overexpressed with β-catenin tagged with HA. Monoclonal anti-HA were used to immunoprecipitate β-catenin and co-immunoprecipitated LEF1/TCF cis-elements were amplified by PCR using primers specific to LEF1/TCF cis -elements 1 and 2. ( C ) Schematic diagram of PME-1 promoter and its luciferase reporter plasmid. Human PME-1 promoter was inserted into pGL4.10 containing luciferase reporter gene to generate pGL4/PME-1 -1000 . ( D ) pGL4/PME-1 -1000 and pGL4.10 showed in panel C were transfected into HEK-293T or SH-SY5Y cells. The Photinus pyralis luciferase activity and Renilla reniformis luciferase activity were measured subsequently and the Photinus pyralis luciferase activity was normalized with Renilla reniformis luciferase activity. Data are presented as mean ± SD (n=3); *P

    Article Snippet: Polyclonal anti-β-catenin, anti-Non-pS-β-catenin (dephosphorylated at S33/S37/T41), anti-GSK-3β and anti-pS9-GSK-3β were purchased from Cell Signaling Technology (Danvers, MA, USA).

    Techniques: Software, Labeling, Immunoprecipitation, Amplification, Polymerase Chain Reaction, Luciferase, Plasmid Preparation, Transfection, Activity Assay

    PME-1 expression is regulated by GSK-3β/β-catenin pathway. ( A , B ) HEK-293T cells were transiently transfected with GSK-3β. Levels of PME-1, β-catenin, GSK-3β, and Non-pS-β-catenin (dephosphorylated β-catenin at Ser33, Ser37 and Thr41) were analyzed by Western blots and normalized with GAPDH or corresponding proteins ( B ). ( C , D ) HA-tagged GSK-3β was overexpressed in HeLa cells and immuno-stained by polyclonal rabbit anti-β-catenin or mouse monoclonal anti-HA (GSK-3β) followed by florescence labeled anti-rabbit (green) or anti-mouse (red) second antibodies, respectively ( C ). The fluorescence levels of the nucleus and the cytoplasm were measured by IMAGE J and nucleus/cytoplasm ratio of β-catenin was analyzed ( D ). ( E ) pGL4/PME-1 -1000 were co-transfected with GSK-3β or siGSK-3β in HEK-293T cells. The luciferase activity was measured. ( F ) HEK-293T cells were co-transfected with β-catenin and/or GSK-3β with pGL4/PME-1 -1000 . The luciferase activity was measured. *: compared with control (Con). : compared with β-catenin. ( G ) The luciferase activity as measured. *: compared with control (Con), #: compared with β-catenin. Data are presented as mean ± SD (n=3), *P

    Journal: Aging (Albany NY)

    Article Title: Glycogen synthase kinase-3β suppresses the expression of protein phosphatase methylesterase-1 through β-catenin

    doi: 10.18632/aging.102413

    Figure Lengend Snippet: PME-1 expression is regulated by GSK-3β/β-catenin pathway. ( A , B ) HEK-293T cells were transiently transfected with GSK-3β. Levels of PME-1, β-catenin, GSK-3β, and Non-pS-β-catenin (dephosphorylated β-catenin at Ser33, Ser37 and Thr41) were analyzed by Western blots and normalized with GAPDH or corresponding proteins ( B ). ( C , D ) HA-tagged GSK-3β was overexpressed in HeLa cells and immuno-stained by polyclonal rabbit anti-β-catenin or mouse monoclonal anti-HA (GSK-3β) followed by florescence labeled anti-rabbit (green) or anti-mouse (red) second antibodies, respectively ( C ). The fluorescence levels of the nucleus and the cytoplasm were measured by IMAGE J and nucleus/cytoplasm ratio of β-catenin was analyzed ( D ). ( E ) pGL4/PME-1 -1000 were co-transfected with GSK-3β or siGSK-3β in HEK-293T cells. The luciferase activity was measured. ( F ) HEK-293T cells were co-transfected with β-catenin and/or GSK-3β with pGL4/PME-1 -1000 . The luciferase activity was measured. *: compared with control (Con). : compared with β-catenin. ( G ) The luciferase activity as measured. *: compared with control (Con), #: compared with β-catenin. Data are presented as mean ± SD (n=3), *P

    Article Snippet: Polyclonal anti-β-catenin, anti-Non-pS-β-catenin (dephosphorylated at S33/S37/T41), anti-GSK-3β and anti-pS9-GSK-3β were purchased from Cell Signaling Technology (Danvers, MA, USA).

    Techniques: Expressing, Transfection, Western Blot, Staining, Labeling, Fluorescence, Luciferase, Activity Assay

    β-catenin enhances PME-1 expression. ( A ) β-Catenin or siβ-catenin was transfected together with pGL4/PME-1 -350 and pRL-TK into HEK-293T cells, and then the PME-1 expression was determined through luciferase activity. ( B ) HEK-293T cells were co-transfected with pGL4/PME-1s, β-catenin and pRL-TK. The luciferase activity was measured. ( C – E ) HEK-293T cells were transfected with β-catenin or siβ-catenin and analyzed for PME-1 and β-catenin by Western blots ( C , D ) or qPCR ( E ). Quantification ( E ) of the Western blots are presented as mean ± SD. *P

    Journal: Aging (Albany NY)

    Article Title: Glycogen synthase kinase-3β suppresses the expression of protein phosphatase methylesterase-1 through β-catenin

    doi: 10.18632/aging.102413

    Figure Lengend Snippet: β-catenin enhances PME-1 expression. ( A ) β-Catenin or siβ-catenin was transfected together with pGL4/PME-1 -350 and pRL-TK into HEK-293T cells, and then the PME-1 expression was determined through luciferase activity. ( B ) HEK-293T cells were co-transfected with pGL4/PME-1s, β-catenin and pRL-TK. The luciferase activity was measured. ( C – E ) HEK-293T cells were transfected with β-catenin or siβ-catenin and analyzed for PME-1 and β-catenin by Western blots ( C , D ) or qPCR ( E ). Quantification ( E ) of the Western blots are presented as mean ± SD. *P

    Article Snippet: Polyclonal anti-β-catenin, anti-Non-pS-β-catenin (dephosphorylated at S33/S37/T41), anti-GSK-3β and anti-pS9-GSK-3β were purchased from Cell Signaling Technology (Danvers, MA, USA).

    Techniques: Expressing, Transfection, Luciferase, Activity Assay, Western Blot, Real-time Polymerase Chain Reaction

    PI3K/AKT signaling upregulates the PME-1 expression through GSK-3β/β-catenin. ( A – C ) HEK-293T cells were transfected with β-catenin and cultured without Fetal bovine serum (FBS, as control, con) or with 2% FBS for 48 hr. The cell lysates were analyzed for total GSK-3β, β-catenin, PME-1 and the phosphorylation of β-catenin and GSK-3β by Western blots ( A ). Levels of phosphorylated β-catenin and GSK-3β were normalized with corresponding proteins ( B ). The mRNA level of PME-1 was measured by qPCR and normalized with GAPDH ( C ). The protein level of PME-1 was quantified from panel A and normalized with GAPDH. ( D ) SH-SY5Y cells were treated with 100 nM insulin for 0.5 hr or 18 hr and analyzed by Western blots developed with indicated antibodies. ( E , F ) Primary cortical neurons were treated with 100 nM insulin for 18 hr. The levels of β-catenin and PME-1 were analyzed by Western blots ( E ) and normalized with GAPDH ( F ). ( G ) Primary cortical neurons were cultured and treated with the indicated concentration LY294002 for 4.5 hr. The PME-1 mRNA level was measured by qPCR and normalized with GAPDH. Data are presented as mean ± SD (n=3); *P

    Journal: Aging (Albany NY)

    Article Title: Glycogen synthase kinase-3β suppresses the expression of protein phosphatase methylesterase-1 through β-catenin

    doi: 10.18632/aging.102413

    Figure Lengend Snippet: PI3K/AKT signaling upregulates the PME-1 expression through GSK-3β/β-catenin. ( A – C ) HEK-293T cells were transfected with β-catenin and cultured without Fetal bovine serum (FBS, as control, con) or with 2% FBS for 48 hr. The cell lysates were analyzed for total GSK-3β, β-catenin, PME-1 and the phosphorylation of β-catenin and GSK-3β by Western blots ( A ). Levels of phosphorylated β-catenin and GSK-3β were normalized with corresponding proteins ( B ). The mRNA level of PME-1 was measured by qPCR and normalized with GAPDH ( C ). The protein level of PME-1 was quantified from panel A and normalized with GAPDH. ( D ) SH-SY5Y cells were treated with 100 nM insulin for 0.5 hr or 18 hr and analyzed by Western blots developed with indicated antibodies. ( E , F ) Primary cortical neurons were treated with 100 nM insulin for 18 hr. The levels of β-catenin and PME-1 were analyzed by Western blots ( E ) and normalized with GAPDH ( F ). ( G ) Primary cortical neurons were cultured and treated with the indicated concentration LY294002 for 4.5 hr. The PME-1 mRNA level was measured by qPCR and normalized with GAPDH. Data are presented as mean ± SD (n=3); *P

    Article Snippet: Polyclonal anti-β-catenin, anti-Non-pS-β-catenin (dephosphorylated at S33/S37/T41), anti-GSK-3β and anti-pS9-GSK-3β were purchased from Cell Signaling Technology (Danvers, MA, USA).

    Techniques: Expressing, Transfection, Cell Culture, Western Blot, Real-time Polymerase Chain Reaction, Concentration Assay

    Inhibition of GSK-3β with ARA014418 enhances the interaction of β-catenin with LEF1/TCF elements and up-regulates PME-1 expression in cultured cells and in vivo. ( A ) SH-SY5Y cells were collected after ARA014418 (20μM) treatment for 4.5 hr, for ChIP assay using antibody to β-catenin. The two LEF1/TCFs were amplified by PCR with their specific primers. ( B ) Primary cortical neurons from embryonic day 18 SD rat were cultured and treated with the indicated concentration ARA014418 for 4.5 hr. The PME-1 mRNA level was measured by qPCR and normalized with GAPDH. ( C – E ) ARA014418 (5 mM 2 μl/mouse) was intracerebroventricularly injected into hTau transgenic mice for 48 hr. The cortex was homogenized and analyzed by Western blots developed with the indicated antibodies ( C ) or qPCR for PME mRNA ( E ). GAPDH was included as a loading control. Levels of phosphorylated β-catenin and GSK-3β were normalized with corresponding proteins ( D ). Data are presented as mean ± SD (n=4 or 5), *P

    Journal: Aging (Albany NY)

    Article Title: Glycogen synthase kinase-3β suppresses the expression of protein phosphatase methylesterase-1 through β-catenin

    doi: 10.18632/aging.102413

    Figure Lengend Snippet: Inhibition of GSK-3β with ARA014418 enhances the interaction of β-catenin with LEF1/TCF elements and up-regulates PME-1 expression in cultured cells and in vivo. ( A ) SH-SY5Y cells were collected after ARA014418 (20μM) treatment for 4.5 hr, for ChIP assay using antibody to β-catenin. The two LEF1/TCFs were amplified by PCR with their specific primers. ( B ) Primary cortical neurons from embryonic day 18 SD rat were cultured and treated with the indicated concentration ARA014418 for 4.5 hr. The PME-1 mRNA level was measured by qPCR and normalized with GAPDH. ( C – E ) ARA014418 (5 mM 2 μl/mouse) was intracerebroventricularly injected into hTau transgenic mice for 48 hr. The cortex was homogenized and analyzed by Western blots developed with the indicated antibodies ( C ) or qPCR for PME mRNA ( E ). GAPDH was included as a loading control. Levels of phosphorylated β-catenin and GSK-3β were normalized with corresponding proteins ( D ). Data are presented as mean ± SD (n=4 or 5), *P

    Article Snippet: Polyclonal anti-β-catenin, anti-Non-pS-β-catenin (dephosphorylated at S33/S37/T41), anti-GSK-3β and anti-pS9-GSK-3β were purchased from Cell Signaling Technology (Danvers, MA, USA).

    Techniques: Inhibition, Expressing, Cell Culture, In Vivo, Chromatin Immunoprecipitation, Amplification, Polymerase Chain Reaction, Concentration Assay, Real-time Polymerase Chain Reaction, Injection, Transgenic Assay, Mouse Assay, Western Blot

    WNT/β-catenin signaling is crucial for tongue muscle development. (A) β-Galactosidase staining of newborn Myog-Cre:R26R mice. Scale bar: 1 mm. (B) Gross picture of newborn wild-type (WT) control and Ctnnb1 conditional null (cKO)

    Journal: Development (Cambridge, England)

    Article Title: WNT/β-catenin signaling plays a crucial role in myoblast fusion through regulation of nephrin expression during development

    doi: 10.1242/dev.168351

    Figure Lengend Snippet: WNT/β-catenin signaling is crucial for tongue muscle development. (A) β-Galactosidase staining of newborn Myog-Cre:R26R mice. Scale bar: 1 mm. (B) Gross picture of newborn wild-type (WT) control and Ctnnb1 conditional null (cKO)

    Article Snippet: E13.5 tongue tissue extracts were incubated with either active β-catenin antibody (Cell Signaling Technology) or IgG overnight at 4°C, followed by precipitation with magnetic beads.

    Techniques: Staining, Mouse Assay

    Nephrin is a downstream target gene of WNT/β-catenin during muscle fusion. (A) Quantitative RT-PCR for the indicated fusion molecules in the tongues from E13.5 wild-type (WT) (blue bars) and conditional null (cKO) (red bars) embryos. ** P

    Journal: Development (Cambridge, England)

    Article Title: WNT/β-catenin signaling plays a crucial role in myoblast fusion through regulation of nephrin expression during development

    doi: 10.1242/dev.168351

    Figure Lengend Snippet: Nephrin is a downstream target gene of WNT/β-catenin during muscle fusion. (A) Quantitative RT-PCR for the indicated fusion molecules in the tongues from E13.5 wild-type (WT) (blue bars) and conditional null (cKO) (red bars) embryos. ** P

    Article Snippet: E13.5 tongue tissue extracts were incubated with either active β-catenin antibody (Cell Signaling Technology) or IgG overnight at 4°C, followed by precipitation with magnetic beads.

    Techniques: Quantitative RT-PCR

    Loss of β-catenin in the Myog-Cre -positive lineage causes a defect in muscle differentiation. (A) BrdU staining of Myog-Cre;Ctnnb1 F/F ;ZsGreen cKI/cKI (cKO) and Myog-Cre;Ctnnb1 F/+ ;ZsGreen cKI/cKI control (WT) embryos at E13.5 and E14.5. The boxed

    Journal: Development (Cambridge, England)

    Article Title: WNT/β-catenin signaling plays a crucial role in myoblast fusion through regulation of nephrin expression during development

    doi: 10.1242/dev.168351

    Figure Lengend Snippet: Loss of β-catenin in the Myog-Cre -positive lineage causes a defect in muscle differentiation. (A) BrdU staining of Myog-Cre;Ctnnb1 F/F ;ZsGreen cKI/cKI (cKO) and Myog-Cre;Ctnnb1 F/+ ;ZsGreen cKI/cKI control (WT) embryos at E13.5 and E14.5. The boxed

    Article Snippet: E13.5 tongue tissue extracts were incubated with either active β-catenin antibody (Cell Signaling Technology) or IgG overnight at 4°C, followed by precipitation with magnetic beads.

    Techniques: BrdU Staining

    pY654-β-catenin is required for EMT. A , AECs from WT or fl/fl β-catenin mice were treated with AdenoCre or AdenoGFP and seeded on Fn. After 4 days, cells were lysed and immunoblotted with the indicated antibodies. Absence of β-catenin

    Journal: The Journal of Biological Chemistry

    Article Title: Axin Pathway Activity Regulates in Vivo pY654-\u03b2-catenin Accumulation and Pulmonary Fibrosis

    doi: 10.1074/jbc.M111.322123

    Figure Lengend Snippet: pY654-β-catenin is required for EMT. A , AECs from WT or fl/fl β-catenin mice were treated with AdenoCre or AdenoGFP and seeded on Fn. After 4 days, cells were lysed and immunoblotted with the indicated antibodies. Absence of β-catenin

    Article Snippet: 9B11-Myc and Snail monoclonal antibodies, pY416-Src, and total β-catenin polyclonal antibodies were from Cell Signaling.

    Techniques: Mouse Assay

    Src activation mediates TGFβ1-induced pY654-β-catenin generation. A , pY654-β-catenin and p-Smad2 in immunoprecipitates and lysates were measured at various times after TGFβ1 stimulation. B , upper , pY654-β-catenin

    Journal: The Journal of Biological Chemistry

    Article Title: Axin Pathway Activity Regulates in Vivo pY654-\u03b2-catenin Accumulation and Pulmonary Fibrosis

    doi: 10.1074/jbc.M111.322123

    Figure Lengend Snippet: Src activation mediates TGFβ1-induced pY654-β-catenin generation. A , pY654-β-catenin and p-Smad2 in immunoprecipitates and lysates were measured at various times after TGFβ1 stimulation. B , upper , pY654-β-catenin

    Article Snippet: 9B11-Myc and Snail monoclonal antibodies, pY416-Src, and total β-catenin polyclonal antibodies were from Cell Signaling.

    Techniques: Activation Assay

    Raised epithelial cell axin levels abrogate EMT. Primary AECs were plated on Fn and treated with either FT4001 (5 m m ) or vehicle control. After 3 days: A , pY654-β-catenin and its complex with p-Smad2 detected by immunoprecipitation are decreased

    Journal: The Journal of Biological Chemistry

    Article Title: Axin Pathway Activity Regulates in Vivo pY654-\u03b2-catenin Accumulation and Pulmonary Fibrosis

    doi: 10.1074/jbc.M111.322123

    Figure Lengend Snippet: Raised epithelial cell axin levels abrogate EMT. Primary AECs were plated on Fn and treated with either FT4001 (5 m m ) or vehicle control. After 3 days: A , pY654-β-catenin and its complex with p-Smad2 detected by immunoprecipitation are decreased

    Article Snippet: 9B11-Myc and Snail monoclonal antibodies, pY416-Src, and total β-catenin polyclonal antibodies were from Cell Signaling.

    Techniques: Immunoprecipitation

    pY654-β-catenin does not significantly contribute to TGFβ1-induced TCF/β-catenin reporter activity. A , AECTs carrying the TOPflash reporter were seeded on Fn. Cells were lysed, and TOPflash activity was measured after treating

    Journal: The Journal of Biological Chemistry

    Article Title: Axin Pathway Activity Regulates in Vivo pY654-\u03b2-catenin Accumulation and Pulmonary Fibrosis

    doi: 10.1074/jbc.M111.322123

    Figure Lengend Snippet: pY654-β-catenin does not significantly contribute to TGFβ1-induced TCF/β-catenin reporter activity. A , AECTs carrying the TOPflash reporter were seeded on Fn. Cells were lysed, and TOPflash activity was measured after treating

    Article Snippet: 9B11-Myc and Snail monoclonal antibodies, pY416-Src, and total β-catenin polyclonal antibodies were from Cell Signaling.

    Techniques: Activity Assay

    Phosphorylation of LRP6-ICD PPP(S/T)P by GSK3β is not required for LRP6-ICD mediated stabilization of cytoplasmic β-catenin. HEK 293T cells were transfected with LacZ (control), LRP6-ICD-HA (WT-ICD) or LRP6-ICD×5m-HA (M-ICD). Twenty-four

    Journal: Journal of cellular biochemistry

    Article Title: Phosphorylation of PPP(S/T)P Motif of the Free LRP6 Intracellular Domain Is Not Required to Activate the Wnt/?-Catenin Pathway and Attenuate GSK3? Activity

    doi: 10.1002/jcb.22318

    Figure Lengend Snippet: Phosphorylation of LRP6-ICD PPP(S/T)P by GSK3β is not required for LRP6-ICD mediated stabilization of cytoplasmic β-catenin. HEK 293T cells were transfected with LacZ (control), LRP6-ICD-HA (WT-ICD) or LRP6-ICD×5m-HA (M-ICD). Twenty-four

    Article Snippet: The following antibodies were used: polyclonal anti-phospho-β-catenin (Ser 33/37, Thr 41, Cell Signaling), monoclonal anti-β-catenin (BD Biosciences), monoclonal PHF-1 (anti-tau Ser396/404, a gift from Dr. P. Davies [ ]), monoclonal Tau5 (anti-total tau, a gift from Dr. L. Binder [ ]), monoclonal anti-GSK3β (BD Biosciences), polyclonal anti-phospho GSK3β (Ser9, Cell Signaling), monoclonal anti-HA (Covance), monoclonal anti-GFP (Roche Applied Science), monoclonal anti-α-tubulin (Sigma) and polyclonal anti-phospho LRP6 (Ser1490, Cell Signaling).

    Techniques: Transfection

    The expression of fusogenic protein syncytin-1 in SCC-9 and its receptor ASCT-2 were increased by TNF-α. ( a ) Protein level of syncytin-1 was evaluated by western blot in SCC-9, MG-63 and Hela cell line. ( b ) Protein level of ASCT-2 was evaluated by western blot in HUVEC, SCC-9, MG-63 and MCF-7 cell line. ( c , f ) The difference of syncytin-1 and ASCT-2 expression in 6, 12, and 24 h between TNF-α stimulated group and control group. The expression of syncytin-1 in SCC-9 and ASCT-2 in HUVECs were continuous and dramatic highly expressed compared to the negative control group with 10 ng/ml TNF-α treatment. The expression of phosphorylation of β-catenin (P-β-catenin) decreased sharply in TNF-α group, while the amount of β-catenin increased evidently. ( d ) Positive expression of syncytin-1 in squamous cell carcinoma, 200× and 400×, respectively. The expression of syncytin-1 mainly localized in cellular membrane and/or cytoplasm, and the level of protein expression varied in different samples. ( e ) The level of protein expression was evaluated by score 0–7. 22 out of 28 samples (78.6%) showed moderate and high expression of syncytin-1 and scored 4–7; While others (21.4%) scored 0–3.

    Journal: Scientific Reports

    Article Title: Up-regulation of syncytin-1 contributes to TNF-α-enhanced fusion between OSCC and HUVECs partly via Wnt/β-catenin-dependent pathway

    doi: 10.1038/srep40983

    Figure Lengend Snippet: The expression of fusogenic protein syncytin-1 in SCC-9 and its receptor ASCT-2 were increased by TNF-α. ( a ) Protein level of syncytin-1 was evaluated by western blot in SCC-9, MG-63 and Hela cell line. ( b ) Protein level of ASCT-2 was evaluated by western blot in HUVEC, SCC-9, MG-63 and MCF-7 cell line. ( c , f ) The difference of syncytin-1 and ASCT-2 expression in 6, 12, and 24 h between TNF-α stimulated group and control group. The expression of syncytin-1 in SCC-9 and ASCT-2 in HUVECs were continuous and dramatic highly expressed compared to the negative control group with 10 ng/ml TNF-α treatment. The expression of phosphorylation of β-catenin (P-β-catenin) decreased sharply in TNF-α group, while the amount of β-catenin increased evidently. ( d ) Positive expression of syncytin-1 in squamous cell carcinoma, 200× and 400×, respectively. The expression of syncytin-1 mainly localized in cellular membrane and/or cytoplasm, and the level of protein expression varied in different samples. ( e ) The level of protein expression was evaluated by score 0–7. 22 out of 28 samples (78.6%) showed moderate and high expression of syncytin-1 and scored 4–7; While others (21.4%) scored 0–3.

    Article Snippet: The membranes were blocked with 5% non-fat milk in Tris-buffered saline which contained 0.05% Tween 20 (the PVDF membrane should be blocked with TBST with 5% BSA when phosphorylated protein was detected) at room temperature for 1 h. Straight after that the membranes were incubated with anti-GAPDH antibody(1:6000) (Proteintech, Wuhan, China), anti-syncytin-1 antibody (1:750, santa cruz), anti-ASCT-2 antibody (1:4000, sigma), anti-P-β-catenin antibody (1:1000, CST), anti-β-catenin antibody(1:20000, epitomics) overnight at 4 °C.

    Techniques: Expressing, Western Blot, Negative Control

    Wnt/β- catenin signal pathway controlled the expression of syncytin-1 in SCC-9 cells. ( a , b ) The expression of β-catenin, P-β-catenin and syncytin-1 when SCC-9 was treated with DKK-1, the inhibitor of Wnt/β-catenin pathway, led to the decrease of syncytin-1and β-catenin, but the expression of P-β-catenin was increased. ( c , d ) The expression of β-catenin, P-β-catenin and syncytin-1 when SCC-9 was treated with Wnt3α, the activator of Wnt/β-catenin pathway, led to the increased expression of syncytin-1 and β-catenin, while P-β-catenin expression decreased. ( e , f ) The expression of syncytin-1 sharply decreased when β-catenin was silenced. ( g , h , i , j ) salvage experiment with 10 ng/ml TNF-α stimulation could restore syncytin-1 expression in either sh-β-catenin or en-β-catenin-treated SCC-9 cells.

    Journal: Scientific Reports

    Article Title: Up-regulation of syncytin-1 contributes to TNF-α-enhanced fusion between OSCC and HUVECs partly via Wnt/β-catenin-dependent pathway

    doi: 10.1038/srep40983

    Figure Lengend Snippet: Wnt/β- catenin signal pathway controlled the expression of syncytin-1 in SCC-9 cells. ( a , b ) The expression of β-catenin, P-β-catenin and syncytin-1 when SCC-9 was treated with DKK-1, the inhibitor of Wnt/β-catenin pathway, led to the decrease of syncytin-1and β-catenin, but the expression of P-β-catenin was increased. ( c , d ) The expression of β-catenin, P-β-catenin and syncytin-1 when SCC-9 was treated with Wnt3α, the activator of Wnt/β-catenin pathway, led to the increased expression of syncytin-1 and β-catenin, while P-β-catenin expression decreased. ( e , f ) The expression of syncytin-1 sharply decreased when β-catenin was silenced. ( g , h , i , j ) salvage experiment with 10 ng/ml TNF-α stimulation could restore syncytin-1 expression in either sh-β-catenin or en-β-catenin-treated SCC-9 cells.

    Article Snippet: The membranes were blocked with 5% non-fat milk in Tris-buffered saline which contained 0.05% Tween 20 (the PVDF membrane should be blocked with TBST with 5% BSA when phosphorylated protein was detected) at room temperature for 1 h. Straight after that the membranes were incubated with anti-GAPDH antibody(1:6000) (Proteintech, Wuhan, China), anti-syncytin-1 antibody (1:750, santa cruz), anti-ASCT-2 antibody (1:4000, sigma), anti-P-β-catenin antibody (1:1000, CST), anti-β-catenin antibody(1:20000, epitomics) overnight at 4 °C.

    Techniques: Expressing

    Wnt/β-catenin signal pathway was associated with fusion between SCC-9 and HUVEC. ( a) The different numbers of fused cells among control group, TNF-α stimulated group, DKK-1 stimulated group and Wnt3α stimulated group by artificial cell counting. ( b) Different fusion rate among the control group, TNF-α group, DKK-1 group and Wnt3α group by FACS, the fusion rate was 0.748 ± 0.024%, 1.78 ± 0.053%, 0.367 ± 0.016%, 1.76 ± 0.07%, respectively. ( c) The statistics analysis of fusion rate by FACS of the control group, TNF-α group, DKK-1 group and Wnt3α group. ( d) The representative fluorescence images of cell fusion between HUVEC and SCC-9 when β-catenin was silenced or/and over-expressed in SCC-9. (d I) showed the representative images of fluorescence when RFP-HUVECs were co-cultured with GFP-SCC-9; (d II) showed the representative images of fluorescence when RFP-HUVECs were co-cultured with sh-β-catenin-SCC-9; (d III) showed the representative images of fluorescence when RFP-HUVECs were co-cultured with en-β-catenin-SCC-9. ( e ) The number of fused cells was evaluated by artificial counting method. The magnification of images d was 100×. ( f ) Different fusion rate between HUVEC and SCC-9 when β-catenin was silenced or/and overexpressed in SCC-9. The fusion rate was 0.30 ± 0.029% when HUVEC was co-cultured with SCC-9 with β-catenin knocked down. When β-catenin was enhanced, the fusion rate between SCC-9 and HUVEC was raised to 1.80 ± 0.033%. ( g ) The statistics analysis of fusion rate by FACS of fusion between HUVEC and SCC-9 whenβ-catenin was silenced or/and overexpressed in SCC-9. The results revealed that Wnt/β-catenin signal pathway was closely associated with fusion between SCC-9 and HUVECs.

    Journal: Scientific Reports

    Article Title: Up-regulation of syncytin-1 contributes to TNF-α-enhanced fusion between OSCC and HUVECs partly via Wnt/β-catenin-dependent pathway

    doi: 10.1038/srep40983

    Figure Lengend Snippet: Wnt/β-catenin signal pathway was associated with fusion between SCC-9 and HUVEC. ( a) The different numbers of fused cells among control group, TNF-α stimulated group, DKK-1 stimulated group and Wnt3α stimulated group by artificial cell counting. ( b) Different fusion rate among the control group, TNF-α group, DKK-1 group and Wnt3α group by FACS, the fusion rate was 0.748 ± 0.024%, 1.78 ± 0.053%, 0.367 ± 0.016%, 1.76 ± 0.07%, respectively. ( c) The statistics analysis of fusion rate by FACS of the control group, TNF-α group, DKK-1 group and Wnt3α group. ( d) The representative fluorescence images of cell fusion between HUVEC and SCC-9 when β-catenin was silenced or/and over-expressed in SCC-9. (d I) showed the representative images of fluorescence when RFP-HUVECs were co-cultured with GFP-SCC-9; (d II) showed the representative images of fluorescence when RFP-HUVECs were co-cultured with sh-β-catenin-SCC-9; (d III) showed the representative images of fluorescence when RFP-HUVECs were co-cultured with en-β-catenin-SCC-9. ( e ) The number of fused cells was evaluated by artificial counting method. The magnification of images d was 100×. ( f ) Different fusion rate between HUVEC and SCC-9 when β-catenin was silenced or/and overexpressed in SCC-9. The fusion rate was 0.30 ± 0.029% when HUVEC was co-cultured with SCC-9 with β-catenin knocked down. When β-catenin was enhanced, the fusion rate between SCC-9 and HUVEC was raised to 1.80 ± 0.033%. ( g ) The statistics analysis of fusion rate by FACS of fusion between HUVEC and SCC-9 whenβ-catenin was silenced or/and overexpressed in SCC-9. The results revealed that Wnt/β-catenin signal pathway was closely associated with fusion between SCC-9 and HUVECs.

    Article Snippet: The membranes were blocked with 5% non-fat milk in Tris-buffered saline which contained 0.05% Tween 20 (the PVDF membrane should be blocked with TBST with 5% BSA when phosphorylated protein was detected) at room temperature for 1 h. Straight after that the membranes were incubated with anti-GAPDH antibody(1:6000) (Proteintech, Wuhan, China), anti-syncytin-1 antibody (1:750, santa cruz), anti-ASCT-2 antibody (1:4000, sigma), anti-P-β-catenin antibody (1:1000, CST), anti-β-catenin antibody(1:20000, epitomics) overnight at 4 °C.

    Techniques: Cell Counting, FACS, Fluorescence, Cell Culture

    The schema graph and summary of fusion rate between SCC-9 and HUVEC. ( a ) The schema graph of fusion between SCC-9 and HUVEC and the molecular mechanism underlying. When TNF-α was added in SCC-9, the tripolymer of APC, Axin and GSK3β was suppressed, This disrupted phosphorylation/degradation of β-catenin, allowing β-catenin to enrich in the cytoplasm and then enter into nucleus, where it served as a co-activator for TCF/LEF to activate syncytin-1; DKK-1 was the inhibitor of Wnt/β-catenin. DKK-1 could bind to LRP5/6, leading to the activation of the tripolymer (APC, Axin and GSK3β), which on the following would augment the phosphorylation/degradation of β-catenin, and the amount of β-catenin was not sufficient enough to pass into the nucleus, so the expression of syncytin-1 would decreased; Wnt3α could stabilize the status of β-catenin, which would cause the increased expression of β-catenin, afterwards, β-catenin would enter into nucleus and activate TCF/LEF, and then aggrandize the expression of syncytin-1. Our previous studies showed that hypoxia could promote angiogenesis of HUVECs through PI3k/Akt signal pathway. In this study, we preliminary explored and considered that PI3k/Akt signal pathway was closely associated with TNF-α enhanced expression of ASCT-2. There are some reports deemed that some tumor cells could secret TNF-α, we also found that SCC-9 could secret soluble TNF-α, and we thought Wnt/β-catenin could elegantly regulate and control the expression of TNF-α. Further study is needed. When TNF-α was added into HUVEC, it could enlarge the expression of ASCT-2, Next, the ligand, syncytin-1 could bind to the receptor, ASCT-2, and then take part in the control of cell fusion between SCC-9 and HUVEC. ------ steps which had not been confirmed —— steps which had been confirmed. ( b ) The summary of fusion rate with/without stimulus and disposing.

    Journal: Scientific Reports

    Article Title: Up-regulation of syncytin-1 contributes to TNF-α-enhanced fusion between OSCC and HUVECs partly via Wnt/β-catenin-dependent pathway

    doi: 10.1038/srep40983

    Figure Lengend Snippet: The schema graph and summary of fusion rate between SCC-9 and HUVEC. ( a ) The schema graph of fusion between SCC-9 and HUVEC and the molecular mechanism underlying. When TNF-α was added in SCC-9, the tripolymer of APC, Axin and GSK3β was suppressed, This disrupted phosphorylation/degradation of β-catenin, allowing β-catenin to enrich in the cytoplasm and then enter into nucleus, where it served as a co-activator for TCF/LEF to activate syncytin-1; DKK-1 was the inhibitor of Wnt/β-catenin. DKK-1 could bind to LRP5/6, leading to the activation of the tripolymer (APC, Axin and GSK3β), which on the following would augment the phosphorylation/degradation of β-catenin, and the amount of β-catenin was not sufficient enough to pass into the nucleus, so the expression of syncytin-1 would decreased; Wnt3α could stabilize the status of β-catenin, which would cause the increased expression of β-catenin, afterwards, β-catenin would enter into nucleus and activate TCF/LEF, and then aggrandize the expression of syncytin-1. Our previous studies showed that hypoxia could promote angiogenesis of HUVECs through PI3k/Akt signal pathway. In this study, we preliminary explored and considered that PI3k/Akt signal pathway was closely associated with TNF-α enhanced expression of ASCT-2. There are some reports deemed that some tumor cells could secret TNF-α, we also found that SCC-9 could secret soluble TNF-α, and we thought Wnt/β-catenin could elegantly regulate and control the expression of TNF-α. Further study is needed. When TNF-α was added into HUVEC, it could enlarge the expression of ASCT-2, Next, the ligand, syncytin-1 could bind to the receptor, ASCT-2, and then take part in the control of cell fusion between SCC-9 and HUVEC. ------ steps which had not been confirmed —— steps which had been confirmed. ( b ) The summary of fusion rate with/without stimulus and disposing.

    Article Snippet: The membranes were blocked with 5% non-fat milk in Tris-buffered saline which contained 0.05% Tween 20 (the PVDF membrane should be blocked with TBST with 5% BSA when phosphorylated protein was detected) at room temperature for 1 h. Straight after that the membranes were incubated with anti-GAPDH antibody(1:6000) (Proteintech, Wuhan, China), anti-syncytin-1 antibody (1:750, santa cruz), anti-ASCT-2 antibody (1:4000, sigma), anti-P-β-catenin antibody (1:1000, CST), anti-β-catenin antibody(1:20000, epitomics) overnight at 4 °C.

    Techniques: Activation Assay, Expressing

    ELK3 regulates the expression of VE-cadherin and VEGFR-3, and the phosphorylation of β-catenin. (A) VE-cadherin expression in LEC transfected with siELK3 and siNS for 48 h was evaluated using RT-qPCR. (B) VE-cadherin expression was detected by immunocytochemical staining in LEC transfected with siELK3 or siNS for 24 h. Arrows indicates VE-cadherin staining. (C) Accumulation of phosphorylated β-catenin was analyzed using western blot analysis. LEC were transfected with siNS or siELK3 for 24 h and untreated or stimulated with VEGF-C for 30 min. (D) The effect of ELK3 knockdown on the expression of the indicated genes was evaluated using real-time RT-qPCR. LEC were transfected with siELK3 for 24 h and the expression of each gene is presented relative to the levels in control (siNS) cells (siELK3/siNS). (E) The quantity of VEGF-C was determined using ELISA. ELK3, E26 transformation-specific domain-containing protein Elk-3; siNS, small interfering RNA non-specific control; siELK3, small interfering RNA ELK3; VE-cadherin, vascular endothelial-cadherin; LEC, lymphatic endothelial cells; VEGF-C, vascular endothelial growth factor C; RT-qPCR, reverse transcription quantitative polymerase chain reaction.

    Journal: Oncology Letters

    Article Title: Novel function of E26 transformation-specific domain-containing protein ELK3 in lymphatic endothelial cells

    doi: 10.3892/ol.2017.7308

    Figure Lengend Snippet: ELK3 regulates the expression of VE-cadherin and VEGFR-3, and the phosphorylation of β-catenin. (A) VE-cadherin expression in LEC transfected with siELK3 and siNS for 48 h was evaluated using RT-qPCR. (B) VE-cadherin expression was detected by immunocytochemical staining in LEC transfected with siELK3 or siNS for 24 h. Arrows indicates VE-cadherin staining. (C) Accumulation of phosphorylated β-catenin was analyzed using western blot analysis. LEC were transfected with siNS or siELK3 for 24 h and untreated or stimulated with VEGF-C for 30 min. (D) The effect of ELK3 knockdown on the expression of the indicated genes was evaluated using real-time RT-qPCR. LEC were transfected with siELK3 for 24 h and the expression of each gene is presented relative to the levels in control (siNS) cells (siELK3/siNS). (E) The quantity of VEGF-C was determined using ELISA. ELK3, E26 transformation-specific domain-containing protein Elk-3; siNS, small interfering RNA non-specific control; siELK3, small interfering RNA ELK3; VE-cadherin, vascular endothelial-cadherin; LEC, lymphatic endothelial cells; VEGF-C, vascular endothelial growth factor C; RT-qPCR, reverse transcription quantitative polymerase chain reaction.

    Article Snippet: The membrane was blocked by 5% skim milk solution at room temperature for 30 min and then incubated with an anti-ELK3 antibody (#sc-17860, 1:1,000 dilution; Santa Cruz Biotechnology Inc., Dallas, TX, USA), an antibody recognizing active β-catenin (unphosphorylated at Ser33/37/Thr41, #4270S), an anti-phospho-β-catenin antibody (phosphorylated at Ser33/37/Thr41, #9561S) (both from Cell Signaling Technology, Inc.), an anti-lamin B antibody (#sc-6216) or an anti-β-actin antibody (#sc-47778) (both from Santa Cruz Biotechnology, Inc.) overnight at 4°C, followed by incubation with a secondary antibody (#sc-2005, 1:2,000 dilution; Santa Cruz Biotechnology, Inc.) for 1 h at room temperature.

    Techniques: Expressing, Transfection, Quantitative RT-PCR, Staining, Western Blot, Enzyme-linked Immunosorbent Assay, Transformation Assay, Small Interfering RNA, Real-time Polymerase Chain Reaction

    NHE1 regulated EMT markers in ESCC cells The down-regulation of NHE1 affected various EMT markers, particularly Snail and β-catenin. Mean ± SEM; n = 4. * P

    Journal: Oncotarget

    Article Title: Na+/H+ exchanger 1 has tumor suppressive activity and prognostic value in esophageal squamous cell carcinoma

    doi: 10.18632/oncotarget.13645

    Figure Lengend Snippet: NHE1 regulated EMT markers in ESCC cells The down-regulation of NHE1 affected various EMT markers, particularly Snail and β-catenin. Mean ± SEM; n = 4. * P

    Article Snippet: The following antibodies were used in the Western blot analysis; a monoclonal anti-AKT antibody, monoclonal anti-phospho AKT antibody, monoclonal anti-GSK-3β antibody, monoclonal anti-Phospho-GSK-3β antibody, polyclonal anti-β-catenin antibody, and monoclonal anti-p21 antibody were from Cell Signaling Technology, and a monoclonal anti-β-actin antibody and a monoclonal anti-p53 antibody were from Sigma-Aldrich (St. Louis, MO, USA).

    Techniques:

    Chronic alcohol consumption increased a) nuclear expression of β-catenin which corresponded to b) increased cytosolic expression of phosphorylated (Ser 21/9) GSK3β in the TEN+EtOH-treated rats compared to TEN controls. Statistical analysis

    Journal: Advances in experimental medicine and biology

    Article Title: Alcohol consumption, Wnt/β-catenin signaling and hepatocarcinogensis

    doi: 10.1007/978-3-319-09614-8_11

    Figure Lengend Snippet: Chronic alcohol consumption increased a) nuclear expression of β-catenin which corresponded to b) increased cytosolic expression of phosphorylated (Ser 21/9) GSK3β in the TEN+EtOH-treated rats compared to TEN controls. Statistical analysis

    Article Snippet: Blotted cytosolic proteins were incubated with either an anti-active β-catenin antibody used in immunohistochemistry, or with a polyclonal antibody recognizing the phospohorylated form of GSK3β, (Phospho-GSK-3α/β (Ser21/9), Cell Signaling Technology, Beverly, MA).

    Techniques: Expressing

    Quantification of immunohistochemical staining of a) PCNA and b) un-phosphorylated (Ser41/33) β-catenin in paraffin-embedded liver sections from chow+DEN (n=17), PF+DEN (n=18) and EtOH+DEN(n=15) diet groups. Data is expressed as means ±

    Journal: Advances in experimental medicine and biology

    Article Title: Alcohol consumption, Wnt/β-catenin signaling and hepatocarcinogensis

    doi: 10.1007/978-3-319-09614-8_11

    Figure Lengend Snippet: Quantification of immunohistochemical staining of a) PCNA and b) un-phosphorylated (Ser41/33) β-catenin in paraffin-embedded liver sections from chow+DEN (n=17), PF+DEN (n=18) and EtOH+DEN(n=15) diet groups. Data is expressed as means ±

    Article Snippet: Blotted cytosolic proteins were incubated with either an anti-active β-catenin antibody used in immunohistochemistry, or with a polyclonal antibody recognizing the phospohorylated form of GSK3β, (Phospho-GSK-3α/β (Ser21/9), Cell Signaling Technology, Beverly, MA).

    Techniques: Immunohistochemistry, Staining

    Dkk1 targets β-catenin in MDSCs and correlates with CD15 myeloid marker. (A and B) LRP5 and LRP6 mRNA expression in MDSCs isolated from LLC tumor-bearing mice 14 d after tumor injection (A), or in Gr1 + /CD11b + naive cells incubated in vitro in the presence or absence of 10% serum from tumor-bearing mice (B). Mean ± SD ( n = 3 mice/group) *, P

    Journal: The Journal of Experimental Medicine

    Article Title: Dickkopf-related protein 1 (Dkk1) regulates the accumulation and function of myeloid derived suppressor cells in cancer

    doi: 10.1084/jem.20150950

    Figure Lengend Snippet: Dkk1 targets β-catenin in MDSCs and correlates with CD15 myeloid marker. (A and B) LRP5 and LRP6 mRNA expression in MDSCs isolated from LLC tumor-bearing mice 14 d after tumor injection (A), or in Gr1 + /CD11b + naive cells incubated in vitro in the presence or absence of 10% serum from tumor-bearing mice (B). Mean ± SD ( n = 3 mice/group) *, P

    Article Snippet: An antibody against total β-catenin was obtained from Cell Signaling Technology.

    Techniques: Marker, Expressing, Isolation, Mouse Assay, Injection, Incubation, In Vitro

    Meckelin regulation of canonical Wnt signaling. ( A ) IF analysis of β-catenin localization in heterozygous (+/−) and homozygous (−/−), bpck , MEFs. Scale bar = 10 μm. ( B ) Quantitation of nuclear β-catenin levels in MEFs. Data are combined from two representative experiments. ( n = 3336 +/− cells, 2802 bpck cells). Statistics are based on Chi-squared analysis (*** P ≤ 0.0001). ( C ) Western blot displaying levels of β-catenin from subcellular MEF fractionation, showing increased levels in bpck membrane and nuclear fractions. Lamin and N-cadherin were used as loading controls. WCL, whole cell lysate; Memb., membrane fraction; Nuc., nuclear fraction. ( D ) β-Catenin responsive luciferase reporter assay (TOPflash) data from HEK293T cells transiently transfected (48 h) with various cDNA constructs: Wnt signaling reporter (firefly luciferase, TOPflash), transfection control (renilla luciferase, pGL4.73), control plasmid (V5 epitope tag, V5-pcDNA3.1(+)), epitope tagged human TMEM67 cDNA plasmid (V5- TMEM67 [pcDNA3.1(+)]), control plasmid (HA epitope tag, HA-pcDNA3.1(+)), epitope-tagged human DVL2 cDNA (HA- DVL2 [pcDNA3.1(+)]) and epitope-tagged CTNNB1 cDNA (HA-β-cat [pcDNA3.1(+)]). Data were collected from two independent experiments. Each condition tested was prepared in triplicate for each experiment, and luciferase activity measured independently. Luciferase activity is reported as a ratio, TOPflash/renilla luciferase RLUs. Statistics were based on Student's t -test (** P

    Journal: Human Molecular Genetics

    Article Title: The Meckel syndrome protein meckelin (TMEM67) is a key regulator of cilia function but is not required for tissue planar polarity

    doi: 10.1093/hmg/ddt054

    Figure Lengend Snippet: Meckelin regulation of canonical Wnt signaling. ( A ) IF analysis of β-catenin localization in heterozygous (+/−) and homozygous (−/−), bpck , MEFs. Scale bar = 10 μm. ( B ) Quantitation of nuclear β-catenin levels in MEFs. Data are combined from two representative experiments. ( n = 3336 +/− cells, 2802 bpck cells). Statistics are based on Chi-squared analysis (*** P ≤ 0.0001). ( C ) Western blot displaying levels of β-catenin from subcellular MEF fractionation, showing increased levels in bpck membrane and nuclear fractions. Lamin and N-cadherin were used as loading controls. WCL, whole cell lysate; Memb., membrane fraction; Nuc., nuclear fraction. ( D ) β-Catenin responsive luciferase reporter assay (TOPflash) data from HEK293T cells transiently transfected (48 h) with various cDNA constructs: Wnt signaling reporter (firefly luciferase, TOPflash), transfection control (renilla luciferase, pGL4.73), control plasmid (V5 epitope tag, V5-pcDNA3.1(+)), epitope tagged human TMEM67 cDNA plasmid (V5- TMEM67 [pcDNA3.1(+)]), control plasmid (HA epitope tag, HA-pcDNA3.1(+)), epitope-tagged human DVL2 cDNA (HA- DVL2 [pcDNA3.1(+)]) and epitope-tagged CTNNB1 cDNA (HA-β-cat [pcDNA3.1(+)]). Data were collected from two independent experiments. Each condition tested was prepared in triplicate for each experiment, and luciferase activity measured independently. Luciferase activity is reported as a ratio, TOPflash/renilla luciferase RLUs. Statistics were based on Student's t -test (** P

    Article Snippet: Slides were washed twice in PBS and incubated in 5% BSA in PBS for 1 h. Primary anti-β-catenin antibody (Cell Signaling, 9562) was applied overnight in 1% BSA in PBS.

    Techniques: Quantitation Assay, Western Blot, Fractionation, Luciferase, Reporter Assay, Transfection, Construct, Plasmid Preparation, Activity Assay

    (A) Examples of TMA scoring based on staining intensity. Light microscope images of mucinous ovarian tumour tissue (T) and stroma (S) from the TMAs. Tissue samples expressing sFRP4 are stained brown with DAB and counterstained with haematoxylin. Images show examples of ( a ) Negative ( b ) Weak ( c ) Moderate and ( d ) Strong staining. Scale bar = 30 μm. ( B ) Graphical representation of the staining intensity quantified for sFRP4 expression and β-catenin expression respectively, following histological quantification of the tumour cores into benign, weak, or strong. The y axis enumerates the staining intensity within each group of tumours.

    Journal: BMC Cell Biology

    Article Title: Secreted frizzled-related protein 4 expression is positively associated with responsiveness to Cisplatin of ovarian cancer cell lines in vitro and with lower tumour grade in mucinous ovarian cancers

    doi: 10.1186/1471-2121-13-25

    Figure Lengend Snippet: (A) Examples of TMA scoring based on staining intensity. Light microscope images of mucinous ovarian tumour tissue (T) and stroma (S) from the TMAs. Tissue samples expressing sFRP4 are stained brown with DAB and counterstained with haematoxylin. Images show examples of ( a ) Negative ( b ) Weak ( c ) Moderate and ( d ) Strong staining. Scale bar = 30 μm. ( B ) Graphical representation of the staining intensity quantified for sFRP4 expression and β-catenin expression respectively, following histological quantification of the tumour cores into benign, weak, or strong. The y axis enumerates the staining intensity within each group of tumours.

    Article Snippet: The TMAs were probed using anti- β-catenin primary antibody (1:150) (Cell Signaling, Cat. no. 05–601) and anti- sFRP4 (1:100) (Upstate.

    Techniques: Staining, Light Microscopy, Expressing

    (A) β-catenin protein expression across the three cancerous cell lines. ( B ) Graphical representation demonstrating inverse sFRP4 and β-catenin protein expression following siRNA treatment in the chemosensitive cell line A2780. Values represent the mean for each group ± SEM. (** = p

    Journal: BMC Cell Biology

    Article Title: Secreted frizzled-related protein 4 expression is positively associated with responsiveness to Cisplatin of ovarian cancer cell lines in vitro and with lower tumour grade in mucinous ovarian cancers

    doi: 10.1186/1471-2121-13-25

    Figure Lengend Snippet: (A) β-catenin protein expression across the three cancerous cell lines. ( B ) Graphical representation demonstrating inverse sFRP4 and β-catenin protein expression following siRNA treatment in the chemosensitive cell line A2780. Values represent the mean for each group ± SEM. (** = p

    Article Snippet: The TMAs were probed using anti- β-catenin primary antibody (1:150) (Cell Signaling, Cat. no. 05–601) and anti- sFRP4 (1:100) (Upstate.

    Techniques: Expressing

    Graphical representation of the quantified proportion of tumours expressing sFRP4 and β-catenin based on histological classification of mucinous tumours as benign, borderline and adenocarcinomas. Values represent the mean for each group ± SEM. (* p

    Journal: BMC Cell Biology

    Article Title: Secreted frizzled-related protein 4 expression is positively associated with responsiveness to Cisplatin of ovarian cancer cell lines in vitro and with lower tumour grade in mucinous ovarian cancers

    doi: 10.1186/1471-2121-13-25

    Figure Lengend Snippet: Graphical representation of the quantified proportion of tumours expressing sFRP4 and β-catenin based on histological classification of mucinous tumours as benign, borderline and adenocarcinomas. Values represent the mean for each group ± SEM. (* p

    Article Snippet: The TMAs were probed using anti- β-catenin primary antibody (1:150) (Cell Signaling, Cat. no. 05–601) and anti- sFRP4 (1:100) (Upstate.

    Techniques: Expressing

    The functional role of miR-139 in XIST regulating β-catenin and ECM proteins ( A ) miR-139 expression in response to XIST knockdown in MLFCs was determined by using real-time PCR. ( B ) miR-139 mimics or miR-139 inhibitor was transfected into MLFCs to achieve ectopic miR-139 expression or miR-139 inhibition, as verified by real-time PCR assays. ( C ) The expression levels of XIST in response to ectopic miR-139 expression or miR-139 inhibition in MLFCs was determined by using real-time PCR assays. ( D ) and ( E ) After co-processing sh-XIST and miR-139 inhibition in MLFCs, β-catenin protein levels (total, nucleus, cytoplasm) were determined by using Western blot assays. ( F ) After co-processing sh-XIST and miR-139 inhibition in MLFCs, Collagen I and α-SMA protein levels were determined by using Western blot assays. The data are showed as mean± SD of three independent experiments. * P

    Journal: Oncotarget

    Article Title: XIST/miR-139 axis regulates bleomycin (BLM)-induced extracellular matrix (ECM) and pulmonary fibrosis through β-catenin

    doi: 10.18632/oncotarget.18310

    Figure Lengend Snippet: The functional role of miR-139 in XIST regulating β-catenin and ECM proteins ( A ) miR-139 expression in response to XIST knockdown in MLFCs was determined by using real-time PCR. ( B ) miR-139 mimics or miR-139 inhibitor was transfected into MLFCs to achieve ectopic miR-139 expression or miR-139 inhibition, as verified by real-time PCR assays. ( C ) The expression levels of XIST in response to ectopic miR-139 expression or miR-139 inhibition in MLFCs was determined by using real-time PCR assays. ( D ) and ( E ) After co-processing sh-XIST and miR-139 inhibition in MLFCs, β-catenin protein levels (total, nucleus, cytoplasm) were determined by using Western blot assays. ( F ) After co-processing sh-XIST and miR-139 inhibition in MLFCs, Collagen I and α-SMA protein levels were determined by using Western blot assays. The data are showed as mean± SD of three independent experiments. * P

    Article Snippet: The membranes were probed with the following antibodies: β-catenin (Cat# E247, Abcam, USA), Collagen I (ab34710, Abcam), α-SMA (ab5694, Abcam), Histone H3 (ab1791, Abcam), β-actin (Cat# ACTN05 (C4), Abcam) and GAPDH (Cat# 6C5, Abcam).

    Techniques: Functional Assay, Expressing, Real-time Polymerase Chain Reaction, Transfection, Inhibition, Western Blot

    XIST knockdown inhibited human and mouse fibroblast ECM ( A ) After XIST knockdown, the protein levels of β-catenin in nucleus and cytoplasm of IMR-90 and MLFCs were determined by using Western blot assays. ( B ) After XIST knockdown, the protein levels of ECM factor, Collagen I and α-SMA, in IMR-90 and MLFCs were determined by using Western blot assays. The data are showed as mean ± SD of three independent experiments. ** P

    Journal: Oncotarget

    Article Title: XIST/miR-139 axis regulates bleomycin (BLM)-induced extracellular matrix (ECM) and pulmonary fibrosis through β-catenin

    doi: 10.18632/oncotarget.18310

    Figure Lengend Snippet: XIST knockdown inhibited human and mouse fibroblast ECM ( A ) After XIST knockdown, the protein levels of β-catenin in nucleus and cytoplasm of IMR-90 and MLFCs were determined by using Western blot assays. ( B ) After XIST knockdown, the protein levels of ECM factor, Collagen I and α-SMA, in IMR-90 and MLFCs were determined by using Western blot assays. The data are showed as mean ± SD of three independent experiments. ** P

    Article Snippet: The membranes were probed with the following antibodies: β-catenin (Cat# E247, Abcam, USA), Collagen I (ab34710, Abcam), α-SMA (ab5694, Abcam), Histone H3 (ab1791, Abcam), β-actin (Cat# ACTN05 (C4), Abcam) and GAPDH (Cat# 6C5, Abcam).

    Techniques: Western Blot

    XIST competed with β-catenin for miR-139 binding in IMR-90 and MLFCs ( A ) and ( F ) Online tools predicted that XIST and β-catenin shared an almost identical binding site of miR-139. A wt-XIST luciferase reporter gene vector (human or mice), a mut-XIST vector containing a 7 bp (human) or a 6 bp (mice) mutation on putative binding site of miR-139, a wt-β-catenin vector (human or mice), a mut-β-catenin vector containing a 5 bp mutation on putative binding site of miR-139 (human or mice) was constructed. ( B ) The indicated XIST vectors (human) were co-introduced into IMR-90 cells with miR-139 mimics or miR-139 inhibitor; the luciferase activity was determined by using dual luciferase assays. ( C ) The indicated β-catenin vectors (human) were co-introduced into IMR-90 cells with miR-139 inhibitor; the luciferase activity was determined by using dual luciferase assays. ( D ) The indicated β-catenin vectors (human) were co-introduced into IMR-90 cells with sh-XIST; the luciferase activity was determined by using dual luciferase assays. ( E ) The protein levels of β-catenin in miR-139 mimics- or miR-139 inhibitor-transfected IMR-90 cells were evaluated using Western blot assays. ( G ) The indicated XIST vectors (mice) were co-introduced into MLFCs with miR-139 mimics or miR-139 inhibitor; the luciferase activity was determined by using dual luciferase assays. ( H ) The indicated β-catenin vectors (mice) were co-introduced into MLFCs with miR-139 inhibitor; the luciferase activity was determined by using dual luciferase assays. ( I ) The indicated β-catenin vectors (mice) were co-introduced into MLFCs with sh-XIST; the luciferase activity was determined by using dual luciferase assays. ( J ) The protein levels of β-catenin in miR-139 mimics- or miR-139 inhibitor-transfected MLFCs were evaluated using Western blot assays. The data are showed as mean ± SD of three independent experiments. * P

    Journal: Oncotarget

    Article Title: XIST/miR-139 axis regulates bleomycin (BLM)-induced extracellular matrix (ECM) and pulmonary fibrosis through β-catenin

    doi: 10.18632/oncotarget.18310

    Figure Lengend Snippet: XIST competed with β-catenin for miR-139 binding in IMR-90 and MLFCs ( A ) and ( F ) Online tools predicted that XIST and β-catenin shared an almost identical binding site of miR-139. A wt-XIST luciferase reporter gene vector (human or mice), a mut-XIST vector containing a 7 bp (human) or a 6 bp (mice) mutation on putative binding site of miR-139, a wt-β-catenin vector (human or mice), a mut-β-catenin vector containing a 5 bp mutation on putative binding site of miR-139 (human or mice) was constructed. ( B ) The indicated XIST vectors (human) were co-introduced into IMR-90 cells with miR-139 mimics or miR-139 inhibitor; the luciferase activity was determined by using dual luciferase assays. ( C ) The indicated β-catenin vectors (human) were co-introduced into IMR-90 cells with miR-139 inhibitor; the luciferase activity was determined by using dual luciferase assays. ( D ) The indicated β-catenin vectors (human) were co-introduced into IMR-90 cells with sh-XIST; the luciferase activity was determined by using dual luciferase assays. ( E ) The protein levels of β-catenin in miR-139 mimics- or miR-139 inhibitor-transfected IMR-90 cells were evaluated using Western blot assays. ( G ) The indicated XIST vectors (mice) were co-introduced into MLFCs with miR-139 mimics or miR-139 inhibitor; the luciferase activity was determined by using dual luciferase assays. ( H ) The indicated β-catenin vectors (mice) were co-introduced into MLFCs with miR-139 inhibitor; the luciferase activity was determined by using dual luciferase assays. ( I ) The indicated β-catenin vectors (mice) were co-introduced into MLFCs with sh-XIST; the luciferase activity was determined by using dual luciferase assays. ( J ) The protein levels of β-catenin in miR-139 mimics- or miR-139 inhibitor-transfected MLFCs were evaluated using Western blot assays. The data are showed as mean ± SD of three independent experiments. * P

    Article Snippet: The membranes were probed with the following antibodies: β-catenin (Cat# E247, Abcam, USA), Collagen I (ab34710, Abcam), α-SMA (ab5694, Abcam), Histone H3 (ab1791, Abcam), β-actin (Cat# ACTN05 (C4), Abcam) and GAPDH (Cat# 6C5, Abcam).

    Techniques: Binding Assay, Luciferase, Plasmid Preparation, Mouse Assay, Mutagenesis, Construct, Activity Assay, Transfection, Western Blot

    Expression of miR-139 and β-catenin and their correlations with XIST in tissues ( A ) and ( B ) miR-139 and β-catenin expression in PF and normal tissues were determined by using real-time PCR assays. The data are showed as mean ± SD of three independent experiments. ** P

    Journal: Oncotarget

    Article Title: XIST/miR-139 axis regulates bleomycin (BLM)-induced extracellular matrix (ECM) and pulmonary fibrosis through β-catenin

    doi: 10.18632/oncotarget.18310

    Figure Lengend Snippet: Expression of miR-139 and β-catenin and their correlations with XIST in tissues ( A ) and ( B ) miR-139 and β-catenin expression in PF and normal tissues were determined by using real-time PCR assays. The data are showed as mean ± SD of three independent experiments. ** P

    Article Snippet: The membranes were probed with the following antibodies: β-catenin (Cat# E247, Abcam, USA), Collagen I (ab34710, Abcam), α-SMA (ab5694, Abcam), Histone H3 (ab1791, Abcam), β-actin (Cat# ACTN05 (C4), Abcam) and GAPDH (Cat# 6C5, Abcam).

    Techniques: Expressing, Real-time Polymerase Chain Reaction

    Expression and function of XIST in BLM-induced mouse model of PF ( A ) A total of 36 mice were involved in BLM-induced mouse model of PF; a group of 12 mice were injected with saline (control), sacrificed on day 28 after injection, lung tissues obtained for RNA extraction, HE staining or Masson’s trichrome assay; a group of 24 mice were injected with BLM, 12 of them sacrificed on day 28, the rest 12 of them were infected with LV-sh-NC (control) or LV-sh-XIST on day 28, sacrificed 4 more days later for RNA extraction, HE staining or Masson’s trichrome assay. ( B ) Bleomycin-induced mouse model of lung fibrosis was established as verified by H E-staining and Masson’s trichrome assays for collagen deposition. ( C ) XIST expression was determined in PF tissues by using real-time PCR assays. ( D ) LV-sh-XIST was used to achieve XIST knockdown in PF mice, as verified by using real-time PCR assays. ( E ) sh-XIST was used to establish XIST inhibition in BLM-induced mouse PF. The morphologic changes were exhibited by HE staining and Masson’s trichrome assays. ( F ) The protein levels of β-catenin, Collagen I and α-SMA in response to XIST knockdown in PF tissues were determined by using Western blot assays. The data are showed as mean ± SD of three independent experiments. ** P

    Journal: Oncotarget

    Article Title: XIST/miR-139 axis regulates bleomycin (BLM)-induced extracellular matrix (ECM) and pulmonary fibrosis through β-catenin

    doi: 10.18632/oncotarget.18310

    Figure Lengend Snippet: Expression and function of XIST in BLM-induced mouse model of PF ( A ) A total of 36 mice were involved in BLM-induced mouse model of PF; a group of 12 mice were injected with saline (control), sacrificed on day 28 after injection, lung tissues obtained for RNA extraction, HE staining or Masson’s trichrome assay; a group of 24 mice were injected with BLM, 12 of them sacrificed on day 28, the rest 12 of them were infected with LV-sh-NC (control) or LV-sh-XIST on day 28, sacrificed 4 more days later for RNA extraction, HE staining or Masson’s trichrome assay. ( B ) Bleomycin-induced mouse model of lung fibrosis was established as verified by H E-staining and Masson’s trichrome assays for collagen deposition. ( C ) XIST expression was determined in PF tissues by using real-time PCR assays. ( D ) LV-sh-XIST was used to achieve XIST knockdown in PF mice, as verified by using real-time PCR assays. ( E ) sh-XIST was used to establish XIST inhibition in BLM-induced mouse PF. The morphologic changes were exhibited by HE staining and Masson’s trichrome assays. ( F ) The protein levels of β-catenin, Collagen I and α-SMA in response to XIST knockdown in PF tissues were determined by using Western blot assays. The data are showed as mean ± SD of three independent experiments. ** P

    Article Snippet: The membranes were probed with the following antibodies: β-catenin (Cat# E247, Abcam, USA), Collagen I (ab34710, Abcam), α-SMA (ab5694, Abcam), Histone H3 (ab1791, Abcam), β-actin (Cat# ACTN05 (C4), Abcam) and GAPDH (Cat# 6C5, Abcam).

    Techniques: Expressing, Mouse Assay, Injection, RNA Extraction, Staining, Infection, Real-time Polymerase Chain Reaction, Inhibition, Western Blot

    GSK3β phosphorylates Dzip1. (A and B) Dzip1 interacts with GSK3β. Endogenous GSK3β was immunoprecipitated by Dzip1 but not IgG (A), and endogenous Dzip1 was immunoprecipitated with GFP-GSK3β in HEK 293T cells (B). (C and D) Dzip1 is co-localized with GSK3β at the basal body. G0-phase NIH 3T3 cells expressing GFP-GSK3β were immunostained for Dzip1 and AcTub (C), or cells expressing BFP-Centrin2 were immunostained with GSK3β and Dzip1 (D). Scale bar: 5 μm. (E) GSK3β binds Dzip1 in a kinase-substrate interaction manner. Wild-type (WT) GFP-GSK3β and the mutants S9A, K85R, and R96A were each co-expressed with Myc-Dzip1 in G0-phase HEK 293T cells, and treated with the CK1 inhibitor D4476 or the CK2 inhibitor CX4945. Note that treatment with CX4945 but not D4476 led to a significant decrease in the extent of the up-shifted Dzip1 bands, although the binding of Dzip1 to the GFP-GSK3β variants showed no difference. The extent of the up-shifting of the Dzip1 bands was decreased in K85R-expressing cells. (F) Phosphorylation of Dzip1 is coordinated with GSK3β activation. The kinase activity of GSK3β was negatively correlated with serum stimulation in NIH 3T3 cells. Note that the up-shifted bands (arrowheads) of Dzip1 became evident after serum depletion for 24–48 h, and disappeared after serum restimulation. γ-Tubulin was set as a loading control. (G) GSK3β phosphorylates Dzip1 in vivo. In resting mouse embryo fibroblast (MEFs) treated versus not treated with GSK3 and CK2 inhibitors, the Dzip1 bands were up-shifted less in GSK3- and CK2-inhibited cells. The protein levels of total β-Catenin and GSK3β were steady, but the phosphorylated (S33/37/T41) β-Catenin specifically disappeared from GSK3-inhibited cells. α-Tubulin was set as a loading control. (H) GSK3β phosphorylates Dzip1 in vitro. Auto-phosphorylation of GSK3β (55 kD), and the phosphorylated bands of the middle (28 kD), C-terminus (36 kD), and N- terminus (50 kD) of Dzip1 are shown (left panel). Coomassie blue staining of the gel shows the loaded amounts of Dzip1 fragments (right panel). Note that the S520A mutation resulted in decreased phosphorylation of Dzip1 by GSK3β. (I) Inhibition of GSK3 by BIO causes loss of phospho-S520 in Dzip1.

    Journal: PLoS Biology

    Article Title: GSK3β-Dzip1-Rab8 Cascade Regulates Ciliogenesis after Mitosis

    doi: 10.1371/journal.pbio.1002129

    Figure Lengend Snippet: GSK3β phosphorylates Dzip1. (A and B) Dzip1 interacts with GSK3β. Endogenous GSK3β was immunoprecipitated by Dzip1 but not IgG (A), and endogenous Dzip1 was immunoprecipitated with GFP-GSK3β in HEK 293T cells (B). (C and D) Dzip1 is co-localized with GSK3β at the basal body. G0-phase NIH 3T3 cells expressing GFP-GSK3β were immunostained for Dzip1 and AcTub (C), or cells expressing BFP-Centrin2 were immunostained with GSK3β and Dzip1 (D). Scale bar: 5 μm. (E) GSK3β binds Dzip1 in a kinase-substrate interaction manner. Wild-type (WT) GFP-GSK3β and the mutants S9A, K85R, and R96A were each co-expressed with Myc-Dzip1 in G0-phase HEK 293T cells, and treated with the CK1 inhibitor D4476 or the CK2 inhibitor CX4945. Note that treatment with CX4945 but not D4476 led to a significant decrease in the extent of the up-shifted Dzip1 bands, although the binding of Dzip1 to the GFP-GSK3β variants showed no difference. The extent of the up-shifting of the Dzip1 bands was decreased in K85R-expressing cells. (F) Phosphorylation of Dzip1 is coordinated with GSK3β activation. The kinase activity of GSK3β was negatively correlated with serum stimulation in NIH 3T3 cells. Note that the up-shifted bands (arrowheads) of Dzip1 became evident after serum depletion for 24–48 h, and disappeared after serum restimulation. γ-Tubulin was set as a loading control. (G) GSK3β phosphorylates Dzip1 in vivo. In resting mouse embryo fibroblast (MEFs) treated versus not treated with GSK3 and CK2 inhibitors, the Dzip1 bands were up-shifted less in GSK3- and CK2-inhibited cells. The protein levels of total β-Catenin and GSK3β were steady, but the phosphorylated (S33/37/T41) β-Catenin specifically disappeared from GSK3-inhibited cells. α-Tubulin was set as a loading control. (H) GSK3β phosphorylates Dzip1 in vitro. Auto-phosphorylation of GSK3β (55 kD), and the phosphorylated bands of the middle (28 kD), C-terminus (36 kD), and N- terminus (50 kD) of Dzip1 are shown (left panel). Coomassie blue staining of the gel shows the loaded amounts of Dzip1 fragments (right panel). Note that the S520A mutation resulted in decreased phosphorylation of Dzip1 by GSK3β. (I) Inhibition of GSK3 by BIO causes loss of phospho-S520 in Dzip1.

    Article Snippet: Antibodies and Reagents The CK1 inhibitor D4476 (Tocris Bioscience, 2902), rabbit anti-GSK3β (Santa Cruz Biotechnology, sc-9166), β-Catenin (BD, 610154), phosphorylated GSK3β (S9, Cell Signaling Technology, #93365), and phosphorylated β-Catenin (N3, Cell Signaling Technology, #9561) were kind gifts from Dr. Wei Wu (Tsinghua University).

    Techniques: Immunoprecipitation, Expressing, Binding Assay, Activation Assay, Activity Assay, Serum Depletion, In Vivo, In Vitro, Staining, Mutagenesis, Inhibition

    Increased intratumoral proliferation and nuclear β-Catenin in Gpx3 −/− tumors

    Journal: Cancer research

    Article Title: Tumor suppressor function of the plasma glutathione peroxidase Gpx3 in colitis-associated carcinoma

    doi: 10.1158/0008-5472.CAN-12-3150

    Figure Lengend Snippet: Increased intratumoral proliferation and nuclear β-Catenin in Gpx3 −/− tumors

    Article Snippet: After blocking, primary antibody was added (α-Ki67, (NeoMarkers), 1:1000; α-8-hydroxy-2′ deoxyguanosine (Abcam) 1:50; α-β-catenin, (BD Transduction Laboratories), 1:1000; α-arginase I (Arg1), (Santa Cruz), 1:500; α-IL-1β, (R & D Systems), 1:50; FITC-α-F4/80 (eBioscience), 1:1000) and incubated overnight at 4°C.

    Techniques:

    a  qRT-PCR analysis of chromatin modifiers’ fold changes after the inhibition of Wnt signaling. rECA109 and rKyse150 cells were treated with iCRT14 (25 μM for 24 h) and analyzed for expression of chromatin modifiers using qRT-PCR.  b  qRT-PCR analysis of HMGB1 and CHD4. 24 h after treatment of WNT1 protein, qRT-PCR was performed to analyze the expression of HMGB1 and CHD4. Mean ± SD,  N  = 3, * P

    Journal: Cell Death & Disease

    Article Title: Wnt signaling induces radioresistance through upregulating HMGB1 in esophageal squamous cell carcinoma

    doi: 10.1038/s41419-018-0466-4

    Figure Lengend Snippet: a qRT-PCR analysis of chromatin modifiers’ fold changes after the inhibition of Wnt signaling. rECA109 and rKyse150 cells were treated with iCRT14 (25 μM for 24 h) and analyzed for expression of chromatin modifiers using qRT-PCR. b qRT-PCR analysis of HMGB1 and CHD4. 24 h after treatment of WNT1 protein, qRT-PCR was performed to analyze the expression of HMGB1 and CHD4. Mean ± SD, N  = 3, * P

    Article Snippet: The cells were then soaked in normal goat serum for 30 min and incubated with primary antibodies: c-myc (proteintech; 1:50; USA), RAD51 (abcam; 1:1000; UK), KU80 (abcam; 1:1000; UK), β-catenin (proteintech; 1:50; USA), MDC1 (proteintech; 1:50; USA), and phospho-γH2AX (Cell Signaling (20E3); 1:250; USA) overnight at 4 °C.

    Techniques: Quantitative RT-PCR, Inhibition, Expressing

    SLIT/ROBO1 signaling regulates the subcellular localization of β-catenin

    Journal: Developmental cell

    Article Title: SLIT/ROBO1 signaling suppresses mammary branching morphogenesis by limiting basal cell number

    doi: 10.1016/j.devcel.2011.05.012

    Figure Lengend Snippet: SLIT/ROBO1 signaling regulates the subcellular localization of β-catenin

    Article Snippet: To investigate whether β-catenin is downstream of SLIT/ROBO1 in basal cells, we treated HME50 cells with SLIT2 and, using biochemical fractionation, detected a shift in β-catenin from the nuclear to the cytosolic/membrane fractions ( ).

    Techniques:

    TRAIL-TZD regulatesβ-catenin expression via a post-translational mechanism independent of GSK3β. A , Huh-7 cells were treated with DMSO or the TRAIL-TZD combination in the presence of 20 μg/ml cycloheximide ( CHX, lanes 3–12 ). Cells were harvested at 0, 2, 4, 6, and 8 h after treatment followed by Western blot analysis with the indicated antibodies. The Huh-7 cells treated with either DMSO or the TRAIL-TZD combination in the absence of cycloheximide were included in lanes 1 and 2 , respectively, as positive controls to show TRAIL-TZD effects on β-catenin. B , Huh-7 cells were treated with the TRAIL-TZD combination for 24 h in the presence (+) or absence (-) of a pretreatment with the GSK3β inhibitor AR-A014418. At the end of the incubation total cell extracts were prepared and Western blot analysis was performed with the indicated antibodies. C , wild type ( WT +/+ ) or GSK3β knock-out ( KO -/- ) MEFs were incubated with either DMSO or the TRAIL-TZD combination. Total cell extracts were prepared at the indicated time points followed by Western blot analysis with the antibodies shown. D , Huh-7 cells were treated as in B followed by apoptosis assay utilizing the cell death detection ELISA PLUS kit. The data represent the mean ± S.D. of two independent experiments. E , phase-contrast microscopic pictures showing apoptotic morphology of Huh-7 cells following treatment with either DMSO or the TRAIL-TZD combination in the presence (+) or absence (-) of AR-A014418.

    Journal: The Journal of Biological Chemistry

    Article Title: Caspase-mediated Cleavage of ?-Catenin Precedes Drug-induced Apoptosis in Resistant Cancer Cells *Caspase-mediated Cleavage of ?-Catenin Precedes Drug-induced Apoptosis in Resistant Cancer Cells * S⃞

    doi: 10.1074/jbc.M900248200

    Figure Lengend Snippet: TRAIL-TZD regulatesβ-catenin expression via a post-translational mechanism independent of GSK3β. A , Huh-7 cells were treated with DMSO or the TRAIL-TZD combination in the presence of 20 μg/ml cycloheximide ( CHX, lanes 3–12 ). Cells were harvested at 0, 2, 4, 6, and 8 h after treatment followed by Western blot analysis with the indicated antibodies. The Huh-7 cells treated with either DMSO or the TRAIL-TZD combination in the absence of cycloheximide were included in lanes 1 and 2 , respectively, as positive controls to show TRAIL-TZD effects on β-catenin. B , Huh-7 cells were treated with the TRAIL-TZD combination for 24 h in the presence (+) or absence (-) of a pretreatment with the GSK3β inhibitor AR-A014418. At the end of the incubation total cell extracts were prepared and Western blot analysis was performed with the indicated antibodies. C , wild type ( WT +/+ ) or GSK3β knock-out ( KO -/- ) MEFs were incubated with either DMSO or the TRAIL-TZD combination. Total cell extracts were prepared at the indicated time points followed by Western blot analysis with the antibodies shown. D , Huh-7 cells were treated as in B followed by apoptosis assay utilizing the cell death detection ELISA PLUS kit. The data represent the mean ± S.D. of two independent experiments. E , phase-contrast microscopic pictures showing apoptotic morphology of Huh-7 cells following treatment with either DMSO or the TRAIL-TZD combination in the presence (+) or absence (-) of AR-A014418.

    Article Snippet: The antibodies were obtained from the following sources: poly(ADP-ribose) polymerase (PARP), caspase-3, cleaved caspase-3, caspase-8, GSK3β, phospho-GSK3βSer9 , Akt, and phospho-glycogen synthaseSer641 from Cell Signaling Technology (Danvers, MA); β-catenin from Zymed Laboratories Inc., Invitrogen, and BD Biosciences (San Jose, CA); glycogen synthase from Invitrogen; cyclin-D1 from Neomarkers (Fremont, CA); Bcl-xL from Exalpha Biologicals Inc. (Watertown, MA); GAPDH from Ambion Inc. (Austin, TX).

    Techniques: Expressing, Western Blot, Incubation, Knock-Out, Apoptosis Assay, Enzyme-linked Immunosorbent Assay

    Consecutive sections with foci of (upper left) intense FHL2 expression with concomitant (upper right) reduced E-cadherin expression and (lower) appearance of nuclear expression of  β -catenin (arrows) in areas of EMT. Note the lower FHL2 expression as well as the stronger E-cadherin expression and the absence of nuclear  β -catenin expression in adjacent well-differentiated tumour glands (** magnification × 400).

    Journal: British Journal of Cancer

    Article Title: Epithelial expression of FHL2 is negatively associated with metastasis-free and overall survival in colorectal cancer

    doi: 10.1038/bjc.2013.290

    Figure Lengend Snippet: Consecutive sections with foci of (upper left) intense FHL2 expression with concomitant (upper right) reduced E-cadherin expression and (lower) appearance of nuclear expression of β -catenin (arrows) in areas of EMT. Note the lower FHL2 expression as well as the stronger E-cadherin expression and the absence of nuclear β -catenin expression in adjacent well-differentiated tumour glands (** magnification × 400).

    Article Snippet: For E-cadherin and β -catenin, we used antibodies provided by Dako (Glostrup, Denmark; NCH-38, dilution 1 : 100; β -catenin 1, dilution 1 : 300).

    Techniques: Expressing

    Cad6B CTF2s are stabilized through β-catenin binding. (A) Cad6B CTF2 mutants were generated through alanine substitutions within the β-catenin minimal binding domain and at key adjacent phosphorylation sites. Black bars represent the ratio of coimmunoprecipitated β-catenin to immunoprecipitated (IP) mutant CTF2 levels normalized to the wild-type (WT) ratio (set at 100%); gray bars represent the input levels of mutant CTF2 protein (normalized to β-actin levels) relative to WT CTF2 input levels (set at 100%). Means that share letter superscripts are not significantly different (P

    Journal: The Journal of Cell Biology

    Article Title: Cadherin-6B proteolysis promotes the neural crest cell epithelial-to-mesenchymal transition through transcriptional regulation

    doi: 10.1083/jcb.201604006

    Figure Lengend Snippet: Cad6B CTF2s are stabilized through β-catenin binding. (A) Cad6B CTF2 mutants were generated through alanine substitutions within the β-catenin minimal binding domain and at key adjacent phosphorylation sites. Black bars represent the ratio of coimmunoprecipitated β-catenin to immunoprecipitated (IP) mutant CTF2 levels normalized to the wild-type (WT) ratio (set at 100%); gray bars represent the input levels of mutant CTF2 protein (normalized to β-actin levels) relative to WT CTF2 input levels (set at 100%). Means that share letter superscripts are not significantly different (P

    Article Snippet: To quantify effects on β-catenin distribution, we analyzed electroporated and unelectroporated Snail2-positive migratory neural crest cells in at least six ex vivo neural crest explants per plasmid for the presence of nuclear β-catenin ( JJ).

    Techniques: Binding Assay, Generated, Immunoprecipitation, Mutagenesis

    Transient elevation of Cad6B CTF2 in the cranial neural crest up-regulates multiple EMT effector genes and controls endogenous Snail2 expression via an association with chromatin. (A) Embryos were unilaterally electroporated with GFP (white), MUT9 (gray), or CTF2 (black) expression vectors at 3ss and incubated for an additional 5 h (6–7ss). Excised neural crest cells were pooled and lysed for RNA extraction, followed by generation of cDNA. Gene levels were normalized to 18S ribosomal RNA, and the graph shows differences in gene expression as determined by calculating the fold-change from the control group (CTRL; arbitrarily set to 1; n = 3). (B) Evaluation of the Snail2 -GFP reporter to increased nuclear β-catenin as a result of CTF2 overexpression. Embryos were electroporated as described in A with a GFP reporter driven by a minimal Snail2 promoter, along with either empty vector, CTF2, MUT9, or human β-catenin. Differences in GFP were assessed and depicted graphically, with GFP levels for CTF2 arbitrarily set to 1 ( n = 3). (C) In vivo ChIP-QPCR demonstrates β-catenin–dependent association of CTF2, but not MUT9 or GFP, to chromatin regions within the endogenous Snail2 promoter. Embryos were electroporated as described in A, and fold enrichment of amplicon expression between treatments per primer set was assessed after normalization to GFP group means ( n = 2). The Snail2 promoter schematic (not to scale) depicts the 1.2-kb sequence upstream of the translational start site that was tested. Gray bars represent amplicons not significantly enriched; black bars represent amplicons from primer sets 3 and 6 showing CTF2 association. +, relative position of a validated TCF/LEF site ( Sakai et al., 2005 ) covered by primer set 4; #, putative TCF/LEF site. Data in all graphs show means and SEM, and all asterisks indicate a significant difference in gene expression (A and B) or enrichment (C; P

    Journal: The Journal of Cell Biology

    Article Title: Cadherin-6B proteolysis promotes the neural crest cell epithelial-to-mesenchymal transition through transcriptional regulation

    doi: 10.1083/jcb.201604006

    Figure Lengend Snippet: Transient elevation of Cad6B CTF2 in the cranial neural crest up-regulates multiple EMT effector genes and controls endogenous Snail2 expression via an association with chromatin. (A) Embryos were unilaterally electroporated with GFP (white), MUT9 (gray), or CTF2 (black) expression vectors at 3ss and incubated for an additional 5 h (6–7ss). Excised neural crest cells were pooled and lysed for RNA extraction, followed by generation of cDNA. Gene levels were normalized to 18S ribosomal RNA, and the graph shows differences in gene expression as determined by calculating the fold-change from the control group (CTRL; arbitrarily set to 1; n = 3). (B) Evaluation of the Snail2 -GFP reporter to increased nuclear β-catenin as a result of CTF2 overexpression. Embryos were electroporated as described in A with a GFP reporter driven by a minimal Snail2 promoter, along with either empty vector, CTF2, MUT9, or human β-catenin. Differences in GFP were assessed and depicted graphically, with GFP levels for CTF2 arbitrarily set to 1 ( n = 3). (C) In vivo ChIP-QPCR demonstrates β-catenin–dependent association of CTF2, but not MUT9 or GFP, to chromatin regions within the endogenous Snail2 promoter. Embryos were electroporated as described in A, and fold enrichment of amplicon expression between treatments per primer set was assessed after normalization to GFP group means ( n = 2). The Snail2 promoter schematic (not to scale) depicts the 1.2-kb sequence upstream of the translational start site that was tested. Gray bars represent amplicons not significantly enriched; black bars represent amplicons from primer sets 3 and 6 showing CTF2 association. +, relative position of a validated TCF/LEF site ( Sakai et al., 2005 ) covered by primer set 4; #, putative TCF/LEF site. Data in all graphs show means and SEM, and all asterisks indicate a significant difference in gene expression (A and B) or enrichment (C; P

    Article Snippet: To quantify effects on β-catenin distribution, we analyzed electroporated and unelectroporated Snail2-positive migratory neural crest cells in at least six ex vivo neural crest explants per plasmid for the presence of nuclear β-catenin ( JJ).

    Techniques: Expressing, Incubation, RNA Extraction, Over Expression, Plasmid Preparation, In Vivo, Chromatin Immunoprecipitation, Real-time Polymerase Chain Reaction, Amplification, Sequencing

    Cad6B CTF2 overexpression results in the redistribution and accumulation of β-catenin in cranial neural crest cells in vivo. Representative transverse sections taken through the midbrain region of 8–9ss embryos expressing CTF2-HA (A–C), MUT9-HA (E–G), or GFP (I–K), followed by immunostaining for Snail2 (purple), HA or GFP (green), and β-catenin (red). Boxes in lower-magnification (20×) merge images (A, E, and I) mark respective magnified (63×) areas (B, C, F, G, J, and K). (D, H, and L) Line scans showing subcellular fluorescent intensity of recombinant protein (green), β-catenin (red), and Snail2 (purple) across cell diameters (C, G, and K, dotted lines) from representative electroporated and adjacent unelectroporated cells. (M) Quantification of β-catenin levels in electroporated (E) and unelectroporated (UN) Snail2-positive neural crest cells reveals a statistically significant increase in the number of cells containing elevated nuclear and cytosolic β-catenin levels (black bars) versus cytosolic-only β-catenin (gray bars) in the presence of CTF2 but not MUT9 or GFP (*, P

    Journal: The Journal of Cell Biology

    Article Title: Cadherin-6B proteolysis promotes the neural crest cell epithelial-to-mesenchymal transition through transcriptional regulation

    doi: 10.1083/jcb.201604006

    Figure Lengend Snippet: Cad6B CTF2 overexpression results in the redistribution and accumulation of β-catenin in cranial neural crest cells in vivo. Representative transverse sections taken through the midbrain region of 8–9ss embryos expressing CTF2-HA (A–C), MUT9-HA (E–G), or GFP (I–K), followed by immunostaining for Snail2 (purple), HA or GFP (green), and β-catenin (red). Boxes in lower-magnification (20×) merge images (A, E, and I) mark respective magnified (63×) areas (B, C, F, G, J, and K). (D, H, and L) Line scans showing subcellular fluorescent intensity of recombinant protein (green), β-catenin (red), and Snail2 (purple) across cell diameters (C, G, and K, dotted lines) from representative electroporated and adjacent unelectroporated cells. (M) Quantification of β-catenin levels in electroporated (E) and unelectroporated (UN) Snail2-positive neural crest cells reveals a statistically significant increase in the number of cells containing elevated nuclear and cytosolic β-catenin levels (black bars) versus cytosolic-only β-catenin (gray bars) in the presence of CTF2 but not MUT9 or GFP (*, P

    Article Snippet: To quantify effects on β-catenin distribution, we analyzed electroporated and unelectroporated Snail2-positive migratory neural crest cells in at least six ex vivo neural crest explants per plasmid for the presence of nuclear β-catenin ( JJ).

    Techniques: Over Expression, In Vivo, Expressing, Immunostaining, Recombinant

    Cad6B CTF2 and β-catenin coimport into the nucleus in cranial neural crest cells ex vivo. Representative explants of neural crest cells overexpressing CTF2-HA (pCS2-CTF2-HA, A and F–I; pCAGGs-CTF2-HA, B and L–O), MUT9-HA (pCS2-MUT9-HA, C and R–U; pCAGGs-MUT9-HA, D and X–AA), or GFP (E and DD–GG), followed by immunostaining for Snail2 (purple), HA or GFP (green), and β-catenin (red). Boxes in lower-magnification 20× merge images (A–E) mark respective magnified (63×) areas (F–I, L–O, R–U, X–AA, and DD–GG). Bar: (A–E) 20 µm; (F–GG) 10 µm. (JJ) Quantification of electroporated (E) and unelectroporated (UN) neural crest cells exhibiting β-catenin nuclear localization reveals a statistically significant increase in the presence of CTF2 but not MUT9. Protein subcellular distribution was determined by fluorescence intensity line profiling (J, K, P, Q, V, W, BB, CC, HH, and II) across candidate cell diameters (arrows and dotted lines in F, L, R, X, and DD). Cell counts were performed based on protein distribution pattern (orange, no nuclear β-catenin or CTF2/MUT9/GFP; green, nuclear CTF2/MUT9/GFP only; light blue, both nuclear β-catenin and CTF2/MUT9/GFP; dark blue, nuclear β-catenin only). Samples sizes ( n ) for each group shown in JJ.

    Journal: The Journal of Cell Biology

    Article Title: Cadherin-6B proteolysis promotes the neural crest cell epithelial-to-mesenchymal transition through transcriptional regulation

    doi: 10.1083/jcb.201604006

    Figure Lengend Snippet: Cad6B CTF2 and β-catenin coimport into the nucleus in cranial neural crest cells ex vivo. Representative explants of neural crest cells overexpressing CTF2-HA (pCS2-CTF2-HA, A and F–I; pCAGGs-CTF2-HA, B and L–O), MUT9-HA (pCS2-MUT9-HA, C and R–U; pCAGGs-MUT9-HA, D and X–AA), or GFP (E and DD–GG), followed by immunostaining for Snail2 (purple), HA or GFP (green), and β-catenin (red). Boxes in lower-magnification 20× merge images (A–E) mark respective magnified (63×) areas (F–I, L–O, R–U, X–AA, and DD–GG). Bar: (A–E) 20 µm; (F–GG) 10 µm. (JJ) Quantification of electroporated (E) and unelectroporated (UN) neural crest cells exhibiting β-catenin nuclear localization reveals a statistically significant increase in the presence of CTF2 but not MUT9. Protein subcellular distribution was determined by fluorescence intensity line profiling (J, K, P, Q, V, W, BB, CC, HH, and II) across candidate cell diameters (arrows and dotted lines in F, L, R, X, and DD). Cell counts were performed based on protein distribution pattern (orange, no nuclear β-catenin or CTF2/MUT9/GFP; green, nuclear CTF2/MUT9/GFP only; light blue, both nuclear β-catenin and CTF2/MUT9/GFP; dark blue, nuclear β-catenin only). Samples sizes ( n ) for each group shown in JJ.

    Article Snippet: To quantify effects on β-catenin distribution, we analyzed electroporated and unelectroporated Snail2-positive migratory neural crest cells in at least six ex vivo neural crest explants per plasmid for the presence of nuclear β-catenin ( JJ).

    Techniques: Ex Vivo, Immunostaining, Fluorescence

    Cad6B CTF2s are generated before and throughout cranial neural crest EMT and remain associated with β-catenin. (A) Cad6B is subjected to γ-secretase–mediated proteolysis before, and during, EMT. Premigratory neural crest cells were electroporated at 2–3ss, and cranial neural folds were collected at specific stages for HA immunoblot analysis. (B) CTF2 physically associates with β-catenin, but not p120-catenin, in vivo. Embryo midbrains were electroporated at 4ss and incubated beyond the initiation of EMT. Dorsal neural tubes containing CTF2-3xFLAG– or GFP-3xFLAG–expressing neural crest cells were subjected to coimmunoprecipitation (co-IP) and immunoblotting. (C) HA-tagged CTF2s created via full-length Cad6B-HA proteolysis remain bound to β-catenin in cranial neural crest cells in vivo. Embryos were dorsal/ventrally electroporated at 4ss, and dorsal neural tubes were collected at 8ss. Normal mouse serum (NMS) immunoprecipitation with Cad6B-HA lysate served as a negative control. WT, wild type.

    Journal: The Journal of Cell Biology

    Article Title: Cadherin-6B proteolysis promotes the neural crest cell epithelial-to-mesenchymal transition through transcriptional regulation

    doi: 10.1083/jcb.201604006

    Figure Lengend Snippet: Cad6B CTF2s are generated before and throughout cranial neural crest EMT and remain associated with β-catenin. (A) Cad6B is subjected to γ-secretase–mediated proteolysis before, and during, EMT. Premigratory neural crest cells were electroporated at 2–3ss, and cranial neural folds were collected at specific stages for HA immunoblot analysis. (B) CTF2 physically associates with β-catenin, but not p120-catenin, in vivo. Embryo midbrains were electroporated at 4ss and incubated beyond the initiation of EMT. Dorsal neural tubes containing CTF2-3xFLAG– or GFP-3xFLAG–expressing neural crest cells were subjected to coimmunoprecipitation (co-IP) and immunoblotting. (C) HA-tagged CTF2s created via full-length Cad6B-HA proteolysis remain bound to β-catenin in cranial neural crest cells in vivo. Embryos were dorsal/ventrally electroporated at 4ss, and dorsal neural tubes were collected at 8ss. Normal mouse serum (NMS) immunoprecipitation with Cad6B-HA lysate served as a negative control. WT, wild type.

    Article Snippet: To quantify effects on β-catenin distribution, we analyzed electroporated and unelectroporated Snail2-positive migratory neural crest cells in at least six ex vivo neural crest explants per plasmid for the presence of nuclear β-catenin ( JJ).

    Techniques: Generated, In Vivo, Incubation, Expressing, Co-Immunoprecipitation Assay, Immunoprecipitation, Negative Control

    FoxO1 knockdown promotes migration, invasion potential, and EMT in OSCC cells. (A) FoxO1 expression was detected by qRT-PCR and Western blot after transfected with FoxO1 shRNA in OSCC cells. (B) Images of the wound closure of monolayer Cal-27 and Tca8113 cells with FoxO1 knockdown at the time point of 0 and 24 h are presented on the left. Quantitative results are illustrated on the right. (C) The effect of FoxO1 knockdown on OSCC cells invasion were determined by Transwell assay with Matrigel, and the representative images are on the left. Quantitative results are illustrated on the right. (D) The effects of Cal-27-FoxO1 knockdown on expressions of EMT markers, E-cadherin, N-cadherin, β-catenin, and Vimentin, were measured using qRT-PCR and Western blot. And the effects of Cal-27-sh-2-STAT3/sh-1-FoxO1 on expressions of EMT markers, E-cadherin, N-cadherin, β-catenin, and Vimentin, were measured using qRT-PCR compared with those of sh-2-STAT3/scramble group. β-actin was used as a loading control. (E) Flow cytometry was used to examine the percentage of apoptotic cells in Cal-27 cells with FoxO1 knockdown and scramble control cells. All assays were carried out in triplicate. Results were shown as means ± SD. * P

    Journal: Frontiers in Oncology

    Article Title: STAT3 Promotes Invasion and Aerobic Glycolysis of Human Oral Squamous Cell Carcinoma via Inhibiting FoxO1

    doi: 10.3389/fonc.2019.01175

    Figure Lengend Snippet: FoxO1 knockdown promotes migration, invasion potential, and EMT in OSCC cells. (A) FoxO1 expression was detected by qRT-PCR and Western blot after transfected with FoxO1 shRNA in OSCC cells. (B) Images of the wound closure of monolayer Cal-27 and Tca8113 cells with FoxO1 knockdown at the time point of 0 and 24 h are presented on the left. Quantitative results are illustrated on the right. (C) The effect of FoxO1 knockdown on OSCC cells invasion were determined by Transwell assay with Matrigel, and the representative images are on the left. Quantitative results are illustrated on the right. (D) The effects of Cal-27-FoxO1 knockdown on expressions of EMT markers, E-cadherin, N-cadherin, β-catenin, and Vimentin, were measured using qRT-PCR and Western blot. And the effects of Cal-27-sh-2-STAT3/sh-1-FoxO1 on expressions of EMT markers, E-cadherin, N-cadherin, β-catenin, and Vimentin, were measured using qRT-PCR compared with those of sh-2-STAT3/scramble group. β-actin was used as a loading control. (E) Flow cytometry was used to examine the percentage of apoptotic cells in Cal-27 cells with FoxO1 knockdown and scramble control cells. All assays were carried out in triplicate. Results were shown as means ± SD. * P

    Article Snippet: Reagents Monoclonal mouse anti-human antibodies to STAT3 (sc-293151), FoxO1 (H-128), β-catenin (sc-1496) were purchased from Santa Cruz Biotechnology Inc. (Santa Cruz, CA, USA).

    Techniques: Migration, Expressing, Quantitative RT-PCR, Western Blot, Transfection, shRNA, Transwell Assay, Flow Cytometry, Cytometry

    STAT3 knockdown inhibits migration, invasion potential, and EMT in OSCC cells. (A) STAT3 mRNA levels were evaluated using qRT-PCR in SCC25, Cal-27, HSC3, Tca8113, and UM1 cells. (B) STAT3 expression was detected by qRT-PCR and Western blot after transfected with STAT3 shRNA in OSCC cells. (C) Images of the wound closure of monolayer Cal-27 and Tca8113 cells with STAT3 knockdown at the time point of 0 and 24 h are presented on the left. Quantitative results are illustrated on the right. (D) The effect of STAT3 knockdown on OSCC cells invasion were determined by Transwell assay with Matrigel, and the representative images are on the left. Quantitative results are illustrated on the right. (E) The effects of Cal-27-STAT3 knockdown on expressions of EMT markers, E-cadherin, N-cadherin, β-catenin, and Vimentin, were measured using qRT-PCR and Western blot. β-actin was used as a loading control. (F) Flow cytometry was used to examine the percentage of apoptotic cells in Cal-27 cells with STAT3 knockdown and scramble control cells. All assays were carried out in triplicate. Results were shown as means ± SD. * P

    Journal: Frontiers in Oncology

    Article Title: STAT3 Promotes Invasion and Aerobic Glycolysis of Human Oral Squamous Cell Carcinoma via Inhibiting FoxO1

    doi: 10.3389/fonc.2019.01175

    Figure Lengend Snippet: STAT3 knockdown inhibits migration, invasion potential, and EMT in OSCC cells. (A) STAT3 mRNA levels were evaluated using qRT-PCR in SCC25, Cal-27, HSC3, Tca8113, and UM1 cells. (B) STAT3 expression was detected by qRT-PCR and Western blot after transfected with STAT3 shRNA in OSCC cells. (C) Images of the wound closure of monolayer Cal-27 and Tca8113 cells with STAT3 knockdown at the time point of 0 and 24 h are presented on the left. Quantitative results are illustrated on the right. (D) The effect of STAT3 knockdown on OSCC cells invasion were determined by Transwell assay with Matrigel, and the representative images are on the left. Quantitative results are illustrated on the right. (E) The effects of Cal-27-STAT3 knockdown on expressions of EMT markers, E-cadherin, N-cadherin, β-catenin, and Vimentin, were measured using qRT-PCR and Western blot. β-actin was used as a loading control. (F) Flow cytometry was used to examine the percentage of apoptotic cells in Cal-27 cells with STAT3 knockdown and scramble control cells. All assays were carried out in triplicate. Results were shown as means ± SD. * P

    Article Snippet: Reagents Monoclonal mouse anti-human antibodies to STAT3 (sc-293151), FoxO1 (H-128), β-catenin (sc-1496) were purchased from Santa Cruz Biotechnology Inc. (Santa Cruz, CA, USA).

    Techniques: Migration, Quantitative RT-PCR, Expressing, Western Blot, Transfection, shRNA, Transwell Assay, Flow Cytometry, Cytometry

    PolyP-70-mediated expression of cyclin D1 in the absence and presence of siRNA or specific inhibitors for signaling molecules. (A) The efficiency of gene knockdown of β-catenin ( > 75%) was determined 48h post transfection by Western-blotting using a specific antibody. The (B) PolyP-mediated up-regulation of cyclin D1 was monitored after siRNA knockdown of β-catenin in EA.hy926 endothelial cells. (C–D) PolyP-mediated overexpression of cyclin D1 in the absence and presence of increasing concentrations of two different Wnt signaling inhibitors (iCRT3 and PNU-74654). (E) PolyP-mediated up-regulation of cyclin D1 in the absence and presence of increasing concentrations of PI3K inhibitor (wortmannin). (F) PolyP-mediated overexpression of cyclin D1 in the absence and presence of increasing concentrations of AKT inhibitor (AKT inhibitor VIII). (G) PolyP-mediated overexpression of cyclin D1 in the absence and presence of increasing concentrations of intracellular calcium chelator (BAPTA-AM). (H) PolyP-mediated up-regulation of cyclin D1 in the absence and presence of increasing concentrations of PLC inhibitor (U-73122). (I) PolyP-mediated overexpression of cyclin D1 in the absence and presence of increasing concentrations of PKC inhibitor (BIS). (J) EA.hy926 cells were transiently transfected with control siRNA or siRNA for ERK1/2 and the efficiency of gene knockdown ( > 75%) was determined 48h post transfection by Western-blotting using a specific antibody. (K) The same as J except that polyP-mediated overexpression of cyclin D1 was monitored after siRNA knockdown of ERK1/2. (L) PolyP-mediated overexpression of cyclin D1 in the absence and presence of increasing concentrations of ERK inhibitor (PD-98059). The results are shown as mean ± standard deviation of 3 different experiments. *P

    Journal: Journal of thrombosis and haemostasis : JTH

    Article Title: Inorganic polyphosphate promotes cyclin D1 synthesis through activation of mTOR/Wnt/β-catenin signaling in endothelial cells

    doi: 10.1111/jth.13477

    Figure Lengend Snippet: PolyP-70-mediated expression of cyclin D1 in the absence and presence of siRNA or specific inhibitors for signaling molecules. (A) The efficiency of gene knockdown of β-catenin ( > 75%) was determined 48h post transfection by Western-blotting using a specific antibody. The (B) PolyP-mediated up-regulation of cyclin D1 was monitored after siRNA knockdown of β-catenin in EA.hy926 endothelial cells. (C–D) PolyP-mediated overexpression of cyclin D1 in the absence and presence of increasing concentrations of two different Wnt signaling inhibitors (iCRT3 and PNU-74654). (E) PolyP-mediated up-regulation of cyclin D1 in the absence and presence of increasing concentrations of PI3K inhibitor (wortmannin). (F) PolyP-mediated overexpression of cyclin D1 in the absence and presence of increasing concentrations of AKT inhibitor (AKT inhibitor VIII). (G) PolyP-mediated overexpression of cyclin D1 in the absence and presence of increasing concentrations of intracellular calcium chelator (BAPTA-AM). (H) PolyP-mediated up-regulation of cyclin D1 in the absence and presence of increasing concentrations of PLC inhibitor (U-73122). (I) PolyP-mediated overexpression of cyclin D1 in the absence and presence of increasing concentrations of PKC inhibitor (BIS). (J) EA.hy926 cells were transiently transfected with control siRNA or siRNA for ERK1/2 and the efficiency of gene knockdown ( > 75%) was determined 48h post transfection by Western-blotting using a specific antibody. (K) The same as J except that polyP-mediated overexpression of cyclin D1 was monitored after siRNA knockdown of ERK1/2. (L) PolyP-mediated overexpression of cyclin D1 in the absence and presence of increasing concentrations of ERK inhibitor (PD-98059). The results are shown as mean ± standard deviation of 3 different experiments. *P

    Article Snippet: Vectashield mounting medium was from Vector Laboratories Inc. (Burlingame, CA). siRNA for β-catenin was from Dharmacon (Lafayette, CO).

    Techniques: Expressing, Transfection, Western Blot, Over Expression, Planar Chromatography, Standard Deviation

    Figure 5 Immunoblots for β‐catenin on Triton‐soluble and Triton‐insoluble fractions of HT29 cells cocultured with Helicobacter pylori . A reduction in band intensity is seen for the Triton‐insoluble (membrane‐bound)

    Journal:

    Article Title: Effects of Helicobacter pylori on the cadherin-catenin complex

    doi: 10.1136/jcp.2006.036772

    Figure Lengend Snippet: Figure 5 Immunoblots for β‐catenin on Triton‐soluble and Triton‐insoluble fractions of HT29 cells cocultured with Helicobacter pylori . A reduction in band intensity is seen for the Triton‐insoluble (membrane‐bound)

    Article Snippet: Immunohistochemical analysis was carried out on paraffin‐wax‐embedded sections of antral and corpus biopsy specimens, with staining using specific monoclonal antibodies to E‐cadherin and β‐catenin (Affiniti, Biomol International, Exeter, UK) at dilutions of 1:800 and 1:500, respectively.

    Techniques: Western Blot

    Figure 2 Immunofluorescent images of HT29 cells on confocal microscopy. (A,B) Stained for β‐catenin. (A) Untreated cells, (B) after coculture for 24 h with Helicobacter pylori strain 60190. Decreased membranous staining

    Journal:

    Article Title: Effects of Helicobacter pylori on the cadherin-catenin complex

    doi: 10.1136/jcp.2006.036772

    Figure Lengend Snippet: Figure 2 Immunofluorescent images of HT29 cells on confocal microscopy. (A,B) Stained for β‐catenin. (A) Untreated cells, (B) after coculture for 24 h with Helicobacter pylori strain 60190. Decreased membranous staining

    Article Snippet: Immunohistochemical analysis was carried out on paraffin‐wax‐embedded sections of antral and corpus biopsy specimens, with staining using specific monoclonal antibodies to E‐cadherin and β‐catenin (Affiniti, Biomol International, Exeter, UK) at dilutions of 1:800 and 1:500, respectively.

    Techniques: Confocal Microscopy, Staining

    Figure 10 Gastric corpus biopsy specimens stained for β‐catenin (A) with Helicobacter pylori infection; (B) without Helicobacter pylori infection.

    Journal:

    Article Title: Effects of Helicobacter pylori on the cadherin-catenin complex

    doi: 10.1136/jcp.2006.036772

    Figure Lengend Snippet: Figure 10 Gastric corpus biopsy specimens stained for β‐catenin (A) with Helicobacter pylori infection; (B) without Helicobacter pylori infection.

    Article Snippet: Immunohistochemical analysis was carried out on paraffin‐wax‐embedded sections of antral and corpus biopsy specimens, with staining using specific monoclonal antibodies to E‐cadherin and β‐catenin (Affiniti, Biomol International, Exeter, UK) at dilutions of 1:800 and 1:500, respectively.

    Techniques: Staining, Infection

    Figure 4 (A) Cell lysates or untreated cocultured with Helicobacter pylori strains 60190 or Tx30a, blotted to β‐catenin. Bands are fainter with coculture, particularly for 60190. Duplicate experiments were run in parallel are shown.

    Journal:

    Article Title: Effects of Helicobacter pylori on the cadherin-catenin complex

    doi: 10.1136/jcp.2006.036772

    Figure Lengend Snippet: Figure 4 (A) Cell lysates or untreated cocultured with Helicobacter pylori strains 60190 or Tx30a, blotted to β‐catenin. Bands are fainter with coculture, particularly for 60190. Duplicate experiments were run in parallel are shown.

    Article Snippet: Immunohistochemical analysis was carried out on paraffin‐wax‐embedded sections of antral and corpus biopsy specimens, with staining using specific monoclonal antibodies to E‐cadherin and β‐catenin (Affiniti, Biomol International, Exeter, UK) at dilutions of 1:800 and 1:500, respectively.

    Techniques:

    Figure 6 Immunoblots to β‐catenin for cell lysates treated with Helicobacter pylori wild‐type strain 60190, and its isogenic cagE− and vacA− mutant strains. No differences were seen in β‐catenin

    Journal:

    Article Title: Effects of Helicobacter pylori on the cadherin-catenin complex

    doi: 10.1136/jcp.2006.036772

    Figure Lengend Snippet: Figure 6 Immunoblots to β‐catenin for cell lysates treated with Helicobacter pylori wild‐type strain 60190, and its isogenic cagE− and vacA− mutant strains. No differences were seen in β‐catenin

    Article Snippet: Immunohistochemical analysis was carried out on paraffin‐wax‐embedded sections of antral and corpus biopsy specimens, with staining using specific monoclonal antibodies to E‐cadherin and β‐catenin (Affiniti, Biomol International, Exeter, UK) at dilutions of 1:800 and 1:500, respectively.

    Techniques: Western Blot, Mutagenesis

    Figure 3 Number of cell–cell junctions staining strongly for β‐catenin (out of 200 counted), for HT29 cells untreated and cocultured with Helicobacter pylori strains 60190 (pathogenic) and Tx30a (non‐pathogenic).

    Journal:

    Article Title: Effects of Helicobacter pylori on the cadherin-catenin complex

    doi: 10.1136/jcp.2006.036772

    Figure Lengend Snippet: Figure 3 Number of cell–cell junctions staining strongly for β‐catenin (out of 200 counted), for HT29 cells untreated and cocultured with Helicobacter pylori strains 60190 (pathogenic) and Tx30a (non‐pathogenic).

    Article Snippet: Immunohistochemical analysis was carried out on paraffin‐wax‐embedded sections of antral and corpus biopsy specimens, with staining using specific monoclonal antibodies to E‐cadherin and β‐catenin (Affiniti, Biomol International, Exeter, UK) at dilutions of 1:800 and 1:500, respectively.

    Techniques: Staining

    Immunohistochemistry of human CU-ACC2 tumor and CU-ACC2 PDX. A . and B . The left columns shows CU-ACC2 human tumor sample and the right two column is from CU-ACC2 PDX. A . The immunochemistry stains include H E, SF1, α-inhibin, Melan-A, Ki-67 and B . β-catenin, p53 and MSH2 C . Immunocytochemistry for Ki67 and SF1 (right columns) for CU-ACC2 cells (DAPI left column).

    Journal: Endocrine-related cancer

    Article Title: Development of New Preclinical Models to Advance Adrenocortical Carcinoma Research

    doi: 10.1530/ERC-17-0447

    Figure Lengend Snippet: Immunohistochemistry of human CU-ACC2 tumor and CU-ACC2 PDX. A . and B . The left columns shows CU-ACC2 human tumor sample and the right two column is from CU-ACC2 PDX. A . The immunochemistry stains include H E, SF1, α-inhibin, Melan-A, Ki-67 and B . β-catenin, p53 and MSH2 C . Immunocytochemistry for Ki67 and SF1 (right columns) for CU-ACC2 cells (DAPI left column).

    Article Snippet: Additional sections were deparaffinized, AR performed and immunostained for Ki-67 (Neomarkers/Thermo Scientific, Waltham, MA; clone SP6; 1:300), p53 (Cell Marque, Rocklin, CA; clone D07; 1:300), β-catenin (Cell Marque, Rocklin, CA; clone 14; 1:20), Melan-A, (Agilent, Santa Clara CA; clone A103; 1:10) and SF1 (Perseus Protemics, Inc. Tokyo, Japan; clone N1665; 1:100).

    Techniques: Immunohistochemistry, Immunocytochemistry

    Immunohistochemistry of human CU-ACC1 tumor and CU-ACC1 PDX. A . and B . The left columns show CU-ACC1 human tumor sample and the right two columns are from CU-ACC1 PDX. A . The immunochemistry stains include H E, SF1, α-inhibin, Melan-A, Ki-67 and B . β-catenin, p53 and MSH2 C . Immunocytochemistry for Ki67 and SF1 (right columns) for CU-ACC1 cells (DAPI – left column)

    Journal: Endocrine-related cancer

    Article Title: Development of New Preclinical Models to Advance Adrenocortical Carcinoma Research

    doi: 10.1530/ERC-17-0447

    Figure Lengend Snippet: Immunohistochemistry of human CU-ACC1 tumor and CU-ACC1 PDX. A . and B . The left columns show CU-ACC1 human tumor sample and the right two columns are from CU-ACC1 PDX. A . The immunochemistry stains include H E, SF1, α-inhibin, Melan-A, Ki-67 and B . β-catenin, p53 and MSH2 C . Immunocytochemistry for Ki67 and SF1 (right columns) for CU-ACC1 cells (DAPI – left column)

    Article Snippet: Additional sections were deparaffinized, AR performed and immunostained for Ki-67 (Neomarkers/Thermo Scientific, Waltham, MA; clone SP6; 1:300), p53 (Cell Marque, Rocklin, CA; clone D07; 1:300), β-catenin (Cell Marque, Rocklin, CA; clone 14; 1:20), Melan-A, (Agilent, Santa Clara CA; clone A103; 1:10) and SF1 (Perseus Protemics, Inc. Tokyo, Japan; clone N1665; 1:100).

    Techniques: Immunohistochemistry, Immunocytochemistry

    SJ26 suppressed cell migration in H1299, MCF7 and Hep3B 2.17 cells A. SJ26 suppressed Wnt1/β-catenin-signaling pathway in H1299, MCF7 and Hep3B 2.17 cells. Cells were treated with 10 μM SJ26 and the total cell extracts were prepared. Immunoblotting analysis was conducted using antibodies against Wnt1, β-catenin, or α-tubulin. B. H1299, MCF7 or Hep3B 2.17 cells were incubated with 10 μM of SJ26 and were subjected to trans-well assay analysis. The extent of cell migration across the wells was recorded after 6 hours. The value of migration rate in DMSO-treated cells was defined as 1. Asterisks indicate p

    Journal: Oncotarget

    Article Title: Identification of a new class of WNT1 inhibitor: Cancer cells migration, G-quadruplex stabilization and target validation

    doi: 10.18632/oncotarget.6622

    Figure Lengend Snippet: SJ26 suppressed cell migration in H1299, MCF7 and Hep3B 2.17 cells A. SJ26 suppressed Wnt1/β-catenin-signaling pathway in H1299, MCF7 and Hep3B 2.17 cells. Cells were treated with 10 μM SJ26 and the total cell extracts were prepared. Immunoblotting analysis was conducted using antibodies against Wnt1, β-catenin, or α-tubulin. B. H1299, MCF7 or Hep3B 2.17 cells were incubated with 10 μM of SJ26 and were subjected to trans-well assay analysis. The extent of cell migration across the wells was recorded after 6 hours. The value of migration rate in DMSO-treated cells was defined as 1. Asterisks indicate p

    Article Snippet: Antibodies against Wnt1 (1:1000 dilution, Spring, REF E3960), β-catenin (1:1000 dilution, GeneTex, GTX61089), and α-tubulin (1:5000 dilution, Millipore, MAB374) were used as the primary antibodies.

    Techniques: Migration, Incubation

    EVI1 regulated the Wnt/β-catenin signaling pathway in NPC. ( a ) EVI1 downregulation inhibited β-catenin expression in the total cell lysate and the nuclear fraction. In contrast, forced expression of EVI1 enhanced β-catenin expression in the total cell lysate and the nuclear fraction. ( b ) The Wnt/β-catenin activity was repressed when EVI1 was knocked down as revealed by a TOP/FOP Flash reporter assay (left panel). EVI1 elevated Wnt/β-catenin activity when EVI1 was overexpressed (right panel). ( c ) The expression levels of β-catenin downstream targets AXIN2 and c-myc were downregulated in sh-EVI1 cells and were upregulated in LV-EVI1 cells. ( d ) The β-catenin promoter region contains one putative EVI1-binding site. ( e ) A ChIP assay demonstrated that EVI1 bound to the β-catenin promoter. ( f ) EVI1 positively regulated the β-catenin promoter activity

    Journal: Journal of Experimental & Clinical Cancer Research : CR

    Article Title: EVI1 promotes epithelial-to-mesenchymal transition, cancer stem cell features and chemo−/radioresistance in nasopharyngeal carcinoma

    doi: 10.1186/s13046-019-1077-3

    Figure Lengend Snippet: EVI1 regulated the Wnt/β-catenin signaling pathway in NPC. ( a ) EVI1 downregulation inhibited β-catenin expression in the total cell lysate and the nuclear fraction. In contrast, forced expression of EVI1 enhanced β-catenin expression in the total cell lysate and the nuclear fraction. ( b ) The Wnt/β-catenin activity was repressed when EVI1 was knocked down as revealed by a TOP/FOP Flash reporter assay (left panel). EVI1 elevated Wnt/β-catenin activity when EVI1 was overexpressed (right panel). ( c ) The expression levels of β-catenin downstream targets AXIN2 and c-myc were downregulated in sh-EVI1 cells and were upregulated in LV-EVI1 cells. ( d ) The β-catenin promoter region contains one putative EVI1-binding site. ( e ) A ChIP assay demonstrated that EVI1 bound to the β-catenin promoter. ( f ) EVI1 positively regulated the β-catenin promoter activity

    Article Snippet: Oligonucleotide transfection and lentivirus infection siRNAs for EVI1 and β-catenin were synthesized by GeneChem(Shanghai, China).

    Techniques: Expressing, Activity Assay, Reporter Assay, Binding Assay, Chromatin Immunoprecipitation

    Rhein inhibits β‐catenin expression in tumour xenografts. ( A ) Representative tumour tissue sections (100 × magnification) stained for β‐catenin by immunohistochemistry. ( B ) Tumour lysates were subjected to Western blot analysis of β‐catenin. The blots were subjected to densitometric analysis and relative quantification. ** P

    Journal: Journal of Cellular and Molecular Medicine

    Article Title: The natural agent rhein induces β‐catenin degradation and tumour growth arrest

    doi: 10.1111/jcmm.13346

    Figure Lengend Snippet: Rhein inhibits β‐catenin expression in tumour xenografts. ( A ) Representative tumour tissue sections (100 × magnification) stained for β‐catenin by immunohistochemistry. ( B ) Tumour lysates were subjected to Western blot analysis of β‐catenin. The blots were subjected to densitometric analysis and relative quantification. ** P

    Article Snippet: The plasmid for phosphorylation‐deficient mutant of β‐catenin (S33A, S37A, T41A, S45A) was from Addgene (Cambridge, MA, USA).

    Techniques: Expressing, Staining, Immunohistochemistry, Western Blot

    Rhein decreases the protein level of β‐catenin without changing its mRNA level. ( A ) HepG2 and Hela cells were treated with rhein in increasing dose indicated for 48 hrs followed by Western blot analysis of β‐catenin, c‐myc and cyclin D1. ( B ) HepG2 and Hela cells were treated with 60 μM rhein for 48 hrs, followed by real‐time RT‐PCR analysis of β‐catenin transcription. The relative mRNA levels of β‐catenin were plotted. Values represent means ± S.D. ( n = 3). The mean level of β‐catenin mRNA in rhein‐untreated group was set as 1.

    Journal: Journal of Cellular and Molecular Medicine

    Article Title: The natural agent rhein induces β‐catenin degradation and tumour growth arrest

    doi: 10.1111/jcmm.13346

    Figure Lengend Snippet: Rhein decreases the protein level of β‐catenin without changing its mRNA level. ( A ) HepG2 and Hela cells were treated with rhein in increasing dose indicated for 48 hrs followed by Western blot analysis of β‐catenin, c‐myc and cyclin D1. ( B ) HepG2 and Hela cells were treated with 60 μM rhein for 48 hrs, followed by real‐time RT‐PCR analysis of β‐catenin transcription. The relative mRNA levels of β‐catenin were plotted. Values represent means ± S.D. ( n = 3). The mean level of β‐catenin mRNA in rhein‐untreated group was set as 1.

    Article Snippet: The plasmid for phosphorylation‐deficient mutant of β‐catenin (S33A, S37A, T41A, S45A) was from Addgene (Cambridge, MA, USA).

    Techniques: Western Blot, Quantitative RT-PCR

    BTRC, Siah1 and Jade1 are not involved in the induction of β‐catenin degradation by rhein. ( A ) HepG2 cells were transduced with negative control siRNA (siCtrl) or BTRC siRNA (siBTRC), followed by treatment with or without rhein (100 μM) for 24 hrs. The levels of β‐catenin and BTRC were detected by Western blotting. ( B ) HepG2 cells were transduced with negative control siRNA (siCtrl) or axin2 siRNA (siAxin2), followed by treatment with or without rhein (100 μM) for 24 hrs. The levels of β‐catenin and axin2 were detected by Western blotting. ( C ) HepG2 cells were transduced with empty vector or HA‐Siah1▵R, followed by treatment with or without rhein (100 μM) for 24 hrs. The levels of β‐catenin and HA‐Siah1▵R were detected by Western blotting. ( D ) HepG2 cells were transduced with empty vector or HA‐Jade1▵2PHD, followed by treatment with or without rhein (100 μM) for 24 hrs. The levels of β‐catenin and HA‐Jade1▵2PHD were detected by Western blotting.

    Journal: Journal of Cellular and Molecular Medicine

    Article Title: The natural agent rhein induces β‐catenin degradation and tumour growth arrest

    doi: 10.1111/jcmm.13346

    Figure Lengend Snippet: BTRC, Siah1 and Jade1 are not involved in the induction of β‐catenin degradation by rhein. ( A ) HepG2 cells were transduced with negative control siRNA (siCtrl) or BTRC siRNA (siBTRC), followed by treatment with or without rhein (100 μM) for 24 hrs. The levels of β‐catenin and BTRC were detected by Western blotting. ( B ) HepG2 cells were transduced with negative control siRNA (siCtrl) or axin2 siRNA (siAxin2), followed by treatment with or without rhein (100 μM) for 24 hrs. The levels of β‐catenin and axin2 were detected by Western blotting. ( C ) HepG2 cells were transduced with empty vector or HA‐Siah1▵R, followed by treatment with or without rhein (100 μM) for 24 hrs. The levels of β‐catenin and HA‐Siah1▵R were detected by Western blotting. ( D ) HepG2 cells were transduced with empty vector or HA‐Jade1▵2PHD, followed by treatment with or without rhein (100 μM) for 24 hrs. The levels of β‐catenin and HA‐Jade1▵2PHD were detected by Western blotting.

    Article Snippet: The plasmid for phosphorylation‐deficient mutant of β‐catenin (S33A, S37A, T41A, S45A) was from Addgene (Cambridge, MA, USA).

    Techniques: Transduction, Negative Control, Western Blot, Plasmid Preparation

    GSK‐3β is involved in rhein‐induced degradation of β‐catenin and inhibition of cell growth. ( A ) HepG2 and Hela cells were treated with or without LiCl (10 mM) and rhein (60 μM) for 48 hrs, followed by Western blot analysis of β‐catenin. ( B ) HepG2 and Hela cells were transfected with GSK‐3β siRNA. Twenty‐four hours after transfection, the cells were treated with or without rhein (60 μM) for 48 hrs, followed by Western blot analysis of β‐catenin and GSK‐3β. ( C ) HepG2 cells were treated with or without rhein (100 μM) for indicated periods, followed by Western blot analysis. ( D ) In vitro kinase assays of immunoprecipitated GSK3β. HepG2 cells were treated with or without 100 μM rhein for 24 hrs, followed by immunoprecipitation with anti‐GSK3β antibody or normal IgG. The immunoprecipitates were subjected to in vitro kinase assays in the presence of a peptide substrate. The relative kinase activity of GSK3β was plotted. ** P

    Journal: Journal of Cellular and Molecular Medicine

    Article Title: The natural agent rhein induces β‐catenin degradation and tumour growth arrest

    doi: 10.1111/jcmm.13346

    Figure Lengend Snippet: GSK‐3β is involved in rhein‐induced degradation of β‐catenin and inhibition of cell growth. ( A ) HepG2 and Hela cells were treated with or without LiCl (10 mM) and rhein (60 μM) for 48 hrs, followed by Western blot analysis of β‐catenin. ( B ) HepG2 and Hela cells were transfected with GSK‐3β siRNA. Twenty‐four hours after transfection, the cells were treated with or without rhein (60 μM) for 48 hrs, followed by Western blot analysis of β‐catenin and GSK‐3β. ( C ) HepG2 cells were treated with or without rhein (100 μM) for indicated periods, followed by Western blot analysis. ( D ) In vitro kinase assays of immunoprecipitated GSK3β. HepG2 cells were treated with or without 100 μM rhein for 24 hrs, followed by immunoprecipitation with anti‐GSK3β antibody or normal IgG. The immunoprecipitates were subjected to in vitro kinase assays in the presence of a peptide substrate. The relative kinase activity of GSK3β was plotted. ** P

    Article Snippet: The plasmid for phosphorylation‐deficient mutant of β‐catenin (S33A, S37A, T41A, S45A) was from Addgene (Cambridge, MA, USA).

    Techniques: Inhibition, Western Blot, Transfection, In Vitro, Immunoprecipitation, Activity Assay

    Rhein promotes proteasomal degradation of β‐catenin. ( A ) HepG2 and Hela cells were treated with a protein translation inhibitor, CHX (100 μg/ml), and with or without rhein (100 μM). At the indicated time‐point, cells were collected followed by Western blot analysis of β‐catenin. ( B ) HepG2 and Hela cells were treated with or without rhein (60 μM) and proteasome inhibitor MG132 (5 μM) for 48 hrs, followed by Western blot analysis of β‐catenin.

    Journal: Journal of Cellular and Molecular Medicine

    Article Title: The natural agent rhein induces β‐catenin degradation and tumour growth arrest

    doi: 10.1111/jcmm.13346

    Figure Lengend Snippet: Rhein promotes proteasomal degradation of β‐catenin. ( A ) HepG2 and Hela cells were treated with a protein translation inhibitor, CHX (100 μg/ml), and with or without rhein (100 μM). At the indicated time‐point, cells were collected followed by Western blot analysis of β‐catenin. ( B ) HepG2 and Hela cells were treated with or without rhein (60 μM) and proteasome inhibitor MG132 (5 μM) for 48 hrs, followed by Western blot analysis of β‐catenin.

    Article Snippet: The plasmid for phosphorylation‐deficient mutant of β‐catenin (S33A, S37A, T41A, S45A) was from Addgene (Cambridge, MA, USA).

    Techniques: Western Blot

    Partial rescue of the cell fate change in Osr2-IresCre;Smad4 fl/fl dental mesenchyme using an exogenous WNT pathway inhibitor in vitro. ( A-C′ ) Immunofluorescence of β-catenin (red) in Smad4 fl/fl dental mesenchymal cells after adenoviral

    Journal: Development (Cambridge, England)

    Article Title: SMAD4-mediated WNT signaling controls the fate of cranial neural crest cells during tooth morphogenesis

    doi: 10.1242/dev.061341

    Figure Lengend Snippet: Partial rescue of the cell fate change in Osr2-IresCre;Smad4 fl/fl dental mesenchyme using an exogenous WNT pathway inhibitor in vitro. ( A-C′ ) Immunofluorescence of β-catenin (red) in Smad4 fl/fl dental mesenchymal cells after adenoviral

    Article Snippet: ELISAs for total β-catenin (Assay Designs) were carried out following the manufacturer's instructions.

    Techniques: In Vitro, Immunofluorescence

    Decreased β-catenin and Vangl2 levels are observed in Atmin Gpg6/Gpg6 E13.5 embryonic kidneys compared with WT littermates. Kidney sections from WT ( A and C ) and Atmin Gpg6/Gpg6 ( B and D ) embryos were immunostained for β-catenin (A and B) and Vangl2 (C and D). β-Catenin immunostaining was significantly reduced in Atmin Gpg6/Gpg6 (B) compared with WT (A). Vangl2 immunostaining was also dramatically decreased and apical enrichment was altered in Atmin Gpg6/Gpg6 (D) compared with WT (C). Overlays of DAPI and Vangl2 localization emphasize the loss of apical enrichment of Vangl2 protein in Atmin Gpg6/Gpg6 (D inset) versus WT (C inset; n = 4). All images are maximum projections of confocal z stacks taken at 0.4 µm intervals. Images were taken in identical conditions and post-acquisition manipulations were identical. Western blotting revealed a 0.4-fold decrease ( I ) in β-catenin ( P

    Journal: Human Molecular Genetics

    Article Title: Atmin mediates kidney morphogenesis by modulating Wnt signaling

    doi: 10.1093/hmg/ddu246

    Figure Lengend Snippet: Decreased β-catenin and Vangl2 levels are observed in Atmin Gpg6/Gpg6 E13.5 embryonic kidneys compared with WT littermates. Kidney sections from WT ( A and C ) and Atmin Gpg6/Gpg6 ( B and D ) embryos were immunostained for β-catenin (A and B) and Vangl2 (C and D). β-Catenin immunostaining was significantly reduced in Atmin Gpg6/Gpg6 (B) compared with WT (A). Vangl2 immunostaining was also dramatically decreased and apical enrichment was altered in Atmin Gpg6/Gpg6 (D) compared with WT (C). Overlays of DAPI and Vangl2 localization emphasize the loss of apical enrichment of Vangl2 protein in Atmin Gpg6/Gpg6 (D inset) versus WT (C inset; n = 4). All images are maximum projections of confocal z stacks taken at 0.4 µm intervals. Images were taken in identical conditions and post-acquisition manipulations were identical. Western blotting revealed a 0.4-fold decrease ( I ) in β-catenin ( P

    Article Snippet: β-Catenin and Vangl2 proteins are altered in AtminGpg6/Gpg6 kidneys Since transcriptional differences relevant to the Wnt signaling pathway were observed between the AtminGpg6/Gpg6 and WT embryonic kidneys, the levels of β-catenin and Vangl2, two key proteins for the canonical and non-canonical/PCP Wnt signaling pathways, respectively, were also examined.

    Techniques: Immunostaining, Western Blot

    SPAG5 regulates SCARA5 expression through the wnt/β-catenin pathway in HCC cells. a , Western blot and qRT-PCR analyses were used to detect SCARA5 expression in Huh7 cells stably transfected with the shNC or the shβ-catenin plasmid. * p

    Journal: Journal of Experimental & Clinical Cancer Research : CR

    Article Title: SPAG5 promotes hepatocellular carcinoma progression by downregulating SCARA5 through modifying β-catenin degradation

    doi: 10.1186/s13046-018-0891-3

    Figure Lengend Snippet: SPAG5 regulates SCARA5 expression through the wnt/β-catenin pathway in HCC cells. a , Western blot and qRT-PCR analyses were used to detect SCARA5 expression in Huh7 cells stably transfected with the shNC or the shβ-catenin plasmid. * p

    Article Snippet: In addition, the plasmid of SPAG5, SCARA5 and β-Catenin were purchased from Genepharma Company.

    Techniques: Expressing, Western Blot, Quantitative RT-PCR, Stable Transfection, Transfection, Plasmid Preparation

    Proposed model by which SAPG5 promotes HCC progression by SCARA5 inhibition through modifying β-catenin ubiquitination

    Journal: Journal of Experimental & Clinical Cancer Research : CR

    Article Title: SPAG5 promotes hepatocellular carcinoma progression by downregulating SCARA5 through modifying β-catenin degradation

    doi: 10.1186/s13046-018-0891-3

    Figure Lengend Snippet: Proposed model by which SAPG5 promotes HCC progression by SCARA5 inhibition through modifying β-catenin ubiquitination

    Article Snippet: In addition, the plasmid of SPAG5, SCARA5 and β-Catenin were purchased from Genepharma Company.

    Techniques: Inhibition

    Nuclear staining associated with mAb 8E7 is observed in β-catenin null cells . A B : Mouse L cell fibroblasts +/- Wnt3a A . Immunofluorescence staining of total (TL-) β-catenin (mAb clone 14, BD Biosciences) and ABC in mouse L cells stably expressing Wnt3a (right) or an empty vector (left). Note prominent nuclear staining in both Wnt3a expressing and non-expressing cells using 8E7 mAb. Exposure times for TL-β-catenin and ABC images are 1313 and 1439 milliseconds, respectively. B . Corresponding immuno-blot. Note that 8E7 detects non-specific band at ~160 kDa (*). C D : WT and β-catenin/plakoglobin null F9 teratocarcinoma cells immunostained with antibodies that recognize TL- and ABC. C . The split panel (upper right) shows the presence of β-catenin/plakoglobin null F9 cells on the coverslip (phase) that are not stained with the TL-β-catenin antibody, but show a diffuse cytoplasmic/nuclear stain with mAb 8E7 (lower right). Inset shows the GAM secondary antibody control. Exposure times for TL-β-catenin and ABC images are 228 and 453 milliseconds, respectively. D . Corresponding immunoblot shows that 8E7 detects a non-specific band at ~160 kDa in F9 cells (*). It is unclear whether this non-specific band contributes to background nuclear staining.

    Journal: Biology Direct

    Article Title: Issues associated with assessing nuclear localization of N-terminally unphosphorylated ?-catenin with monoclonal antibody 8E7

    doi: 10.1186/1745-6150-4-5

    Figure Lengend Snippet: Nuclear staining associated with mAb 8E7 is observed in β-catenin null cells . A B : Mouse L cell fibroblasts +/- Wnt3a A . Immunofluorescence staining of total (TL-) β-catenin (mAb clone 14, BD Biosciences) and ABC in mouse L cells stably expressing Wnt3a (right) or an empty vector (left). Note prominent nuclear staining in both Wnt3a expressing and non-expressing cells using 8E7 mAb. Exposure times for TL-β-catenin and ABC images are 1313 and 1439 milliseconds, respectively. B . Corresponding immuno-blot. Note that 8E7 detects non-specific band at ~160 kDa (*). C D : WT and β-catenin/plakoglobin null F9 teratocarcinoma cells immunostained with antibodies that recognize TL- and ABC. C . The split panel (upper right) shows the presence of β-catenin/plakoglobin null F9 cells on the coverslip (phase) that are not stained with the TL-β-catenin antibody, but show a diffuse cytoplasmic/nuclear stain with mAb 8E7 (lower right). Inset shows the GAM secondary antibody control. Exposure times for TL-β-catenin and ABC images are 228 and 453 milliseconds, respectively. D . Corresponding immunoblot shows that 8E7 detects a non-specific band at ~160 kDa in F9 cells (*). It is unclear whether this non-specific band contributes to background nuclear staining.

    Article Snippet: To address this, we subjected HEK293T cells to Wnt pathway activation using lithium chloride as previously described [ ], followed by immunofluorescence double-labeling with the antibody that specifically recognizes β-catenin that remains unphosphorylated at S37 and T41 (mAb 8E7), and an antibody that presumably recognizes all forms of β-catenin (#06-734; Millipore).

    Techniques: Staining, Immunofluorescence, Stable Transfection, Expressing, Plasmid Preparation

    Subcellular localization of signaling and total forms of β-catenin in HEK 293T cells . A-C : HEK 293T cells were treated with 30 mM NaCl (control, A ) or 30 mM LiCl (B) for 16 hours to induce the post-transcriptional accumulation of cytosolic/nuclear β-catenin. Cells were co-immunostained using a polyclonal antibody raised against the consensus GSK3 phosphorylation sites of human β-catenin (#06-734; Millipore), and a monoclonal antibody that recognizes ABC (8E7, Millipore). DAPI was used to stain cell nuclei. A . NaCl (control)-treated cells display a small amount of junctional and cytoplasmic β-catenin (green), while ABC (red) is found exclusively in the nuclear compartment. B . In the presence of Wnt signaling, cells show increased expression and colocalization of total-β-catenin and ABC in the cytoplasm and nucleus. Note that the punctate, nuclear ABC staining remains largely unchanged after LiCl induction (arrowheads). Rather, a diffuse cyto/nucleoplasmic ABC staining pattern is most evident (arrows in B). No staining is observed with goat-anti-rabbit IgG (GAR) and goat anti-mouse IgG (GAM) secondary antibodies alone (not shown). Exposure times for anti-β-catenin (green) and ABC (red) images are 1215 and 1632 milliseconds, respectively. C . Same experiment as A B except exposure times were matched for the two β-catenin antibodies (1000 ms). D E . Immunoblot analysis of HEK cells treated with LiCl or recombinant mouse Wnt3a protein (R D Systems #1324-WN/CF). Note that 8E7 detects ABC as well as a non-specific band at ~160 kDa (*).

    Journal: Biology Direct

    Article Title: Issues associated with assessing nuclear localization of N-terminally unphosphorylated ?-catenin with monoclonal antibody 8E7

    doi: 10.1186/1745-6150-4-5

    Figure Lengend Snippet: Subcellular localization of signaling and total forms of β-catenin in HEK 293T cells . A-C : HEK 293T cells were treated with 30 mM NaCl (control, A ) or 30 mM LiCl (B) for 16 hours to induce the post-transcriptional accumulation of cytosolic/nuclear β-catenin. Cells were co-immunostained using a polyclonal antibody raised against the consensus GSK3 phosphorylation sites of human β-catenin (#06-734; Millipore), and a monoclonal antibody that recognizes ABC (8E7, Millipore). DAPI was used to stain cell nuclei. A . NaCl (control)-treated cells display a small amount of junctional and cytoplasmic β-catenin (green), while ABC (red) is found exclusively in the nuclear compartment. B . In the presence of Wnt signaling, cells show increased expression and colocalization of total-β-catenin and ABC in the cytoplasm and nucleus. Note that the punctate, nuclear ABC staining remains largely unchanged after LiCl induction (arrowheads). Rather, a diffuse cyto/nucleoplasmic ABC staining pattern is most evident (arrows in B). No staining is observed with goat-anti-rabbit IgG (GAR) and goat anti-mouse IgG (GAM) secondary antibodies alone (not shown). Exposure times for anti-β-catenin (green) and ABC (red) images are 1215 and 1632 milliseconds, respectively. C . Same experiment as A B except exposure times were matched for the two β-catenin antibodies (1000 ms). D E . Immunoblot analysis of HEK cells treated with LiCl or recombinant mouse Wnt3a protein (R D Systems #1324-WN/CF). Note that 8E7 detects ABC as well as a non-specific band at ~160 kDa (*).

    Article Snippet: To address this, we subjected HEK293T cells to Wnt pathway activation using lithium chloride as previously described [ ], followed by immunofluorescence double-labeling with the antibody that specifically recognizes β-catenin that remains unphosphorylated at S37 and T41 (mAb 8E7), and an antibody that presumably recognizes all forms of β-catenin (#06-734; Millipore).

    Techniques: Staining, Expressing, Mass Spectrometry, Recombinant

    Detection of ABC depends on antigen retrieval method . A-D Rat lung paraffin sections. Sections treated with boiling Tris-Urea ( A-C ) or sodium citrate buffer ( D ) before incubation with the following antibodies: ( A ) GAM-HRP secondary alone, ( B ) anti-TL-β-catenin, ( C D ) anti-ABC (8E7). Note that sections treated with Tris-Urea buffer unmask an epitope detected by ABC and present in all nuclei ( C ). Sections treated with sodium citrate buffer show diminished nuclear background, and instead reveal a sub-population of lung airway epithelial cells that contain the ABC epitope (arrows; D ).

    Journal: Biology Direct

    Article Title: Issues associated with assessing nuclear localization of N-terminally unphosphorylated ?-catenin with monoclonal antibody 8E7

    doi: 10.1186/1745-6150-4-5

    Figure Lengend Snippet: Detection of ABC depends on antigen retrieval method . A-D Rat lung paraffin sections. Sections treated with boiling Tris-Urea ( A-C ) or sodium citrate buffer ( D ) before incubation with the following antibodies: ( A ) GAM-HRP secondary alone, ( B ) anti-TL-β-catenin, ( C D ) anti-ABC (8E7). Note that sections treated with Tris-Urea buffer unmask an epitope detected by ABC and present in all nuclei ( C ). Sections treated with sodium citrate buffer show diminished nuclear background, and instead reveal a sub-population of lung airway epithelial cells that contain the ABC epitope (arrows; D ).

    Article Snippet: To address this, we subjected HEK293T cells to Wnt pathway activation using lithium chloride as previously described [ ], followed by immunofluorescence double-labeling with the antibody that specifically recognizes β-catenin that remains unphosphorylated at S37 and T41 (mAb 8E7), and an antibody that presumably recognizes all forms of β-catenin (#06-734; Millipore).

    Techniques: Incubation

    Analysis of intracellular components of Wnt pathway displayed downregulation of Wnt canonical pathway regulators in sFRP4 OE. There was a decrease in β-catenin as shown by immunocytochemical staining of β-catenin (red) in sFRP4 OE cells while there was an increase in nuclear β-catenin in sFRP4 SI cells (scale bar = 10 μm) ( A ). Western blot analysis showed a decrease in β-catenin and increase of GSK3β and APAF1 in sFRP4 OE cells ( B ), FURA-2AM analysis by flow cytometry showed an increase in intracellular calcium level in sFRP4 OE compared to sFRP4 SI cells ( C ).

    Journal: Cancers

    Article Title: Stemness, Pluripotentiality, and Wnt Antagonism: sFRP4, a Wnt antagonist Mediates Pluripotency and Stemness in Glioblastoma

    doi: 10.3390/cancers11010025

    Figure Lengend Snippet: Analysis of intracellular components of Wnt pathway displayed downregulation of Wnt canonical pathway regulators in sFRP4 OE. There was a decrease in β-catenin as shown by immunocytochemical staining of β-catenin (red) in sFRP4 OE cells while there was an increase in nuclear β-catenin in sFRP4 SI cells (scale bar = 10 μm) ( A ). Western blot analysis showed a decrease in β-catenin and increase of GSK3β and APAF1 in sFRP4 OE cells ( B ), FURA-2AM analysis by flow cytometry showed an increase in intracellular calcium level in sFRP4 OE compared to sFRP4 SI cells ( C ).

    Article Snippet: Western Blotting Protein expression levels of sFRP4 (Invitrogen), β-catenin (BioLegend), GSK-3β (Cell Signaling Technology), and APAF1 (R & D Systems, Minneapolis, MN, USA) were analyzed by Western blotting as previously described by Warrier et al. [ ].

    Techniques: Staining, Western Blot, Flow Cytometry, Cytometry

    The interaction of Merlin with both β-catenin and Par3 is required for the establishment of functional junctions (A) IVTT-produced Merlin or Merlin 18-595 was mixed with either GST-β-catenin (top) or Myc-Par3 (middle). Complexes were isolated using either GST beads (top panel) or an anti-Myc antibody (middle panel) and immunoblotted with anti-GST, -Myc, or -Nf2 antibodies. Merlin 18-595 can readily bind Myc-Par3 but not GST-β-catenin in vitro . 10% of the input is shown in the bottom panel. (B) PAM212 keratinocytes were transfected with Myc-Par3, Myc-Par3 511-1266 or Myc-Par3 1-373 and cultured in calcium-containing media for the indicated times. Immunoprecipitation of Par3 followed by immunoblotting with Myc-, AJ- or TJ-specific antibodies revealed that full-length Myc-Par3 and Myc-Par3 511-1266 can associate with Merlin and with AJ and TJ proteins. In contrast, Myc-Par3 1-373 does not associate with Merlin or with AJ and TJ proteins. Note that Myc-Par3 511-1266 and Myc-Par3 1-373 are produced from a Par3 splice variant that does not contain the aPKC-binding site. NRS, normal rabbit serum. (C) TER was measured across calcium-stimulated PAM212 keratinocyte monolayers that stably express Merlin, Merlin 18-595 , or Myc-Par3 1-373 . Note that both Merlin 18-595 and Myc-Par3 1-373 dominantly interfered with the establishment of TER. Values = mean +/− SD. (D–G) Primary wild-type (D and F) or K14-Cre;Nf2 lox/lox (E and G) keratinocytes were stimulated with calcium-containing media for 8 hours, and either processed for indirect immunofluorescence (Fix 1, D and E) or subject to a more stringent in situ extraction (Fix 2, F and G). Immunostaining revealed normal localization of Par3 (green) at cell-cell boundaries in K14-Cre;Nf2 lox/lox .

    Journal: Developmental cell

    Article Title: The Nf2 Tumor Suppressor, Merlin, Regulates Epidermal Development Through the Establishment of a Junctional Polarity Complex

    doi: 10.1016/j.devcel.2010.10.008

    Figure Lengend Snippet: The interaction of Merlin with both β-catenin and Par3 is required for the establishment of functional junctions (A) IVTT-produced Merlin or Merlin 18-595 was mixed with either GST-β-catenin (top) or Myc-Par3 (middle). Complexes were isolated using either GST beads (top panel) or an anti-Myc antibody (middle panel) and immunoblotted with anti-GST, -Myc, or -Nf2 antibodies. Merlin 18-595 can readily bind Myc-Par3 but not GST-β-catenin in vitro . 10% of the input is shown in the bottom panel. (B) PAM212 keratinocytes were transfected with Myc-Par3, Myc-Par3 511-1266 or Myc-Par3 1-373 and cultured in calcium-containing media for the indicated times. Immunoprecipitation of Par3 followed by immunoblotting with Myc-, AJ- or TJ-specific antibodies revealed that full-length Myc-Par3 and Myc-Par3 511-1266 can associate with Merlin and with AJ and TJ proteins. In contrast, Myc-Par3 1-373 does not associate with Merlin or with AJ and TJ proteins. Note that Myc-Par3 511-1266 and Myc-Par3 1-373 are produced from a Par3 splice variant that does not contain the aPKC-binding site. NRS, normal rabbit serum. (C) TER was measured across calcium-stimulated PAM212 keratinocyte monolayers that stably express Merlin, Merlin 18-595 , or Myc-Par3 1-373 . Note that both Merlin 18-595 and Myc-Par3 1-373 dominantly interfered with the establishment of TER. Values = mean +/− SD. (D–G) Primary wild-type (D and F) or K14-Cre;Nf2 lox/lox (E and G) keratinocytes were stimulated with calcium-containing media for 8 hours, and either processed for indirect immunofluorescence (Fix 1, D and E) or subject to a more stringent in situ extraction (Fix 2, F and G). Immunostaining revealed normal localization of Par3 (green) at cell-cell boundaries in K14-Cre;Nf2 lox/lox .

    Article Snippet: For IP and western blot the following antibodies were used: NF2 (1:1000, A-19 and C-18, Santa Cruz), Keratin 14 (1:5000, AF64, Covance), Actin (1:1000, AC40, Sigma), β-tubulin (1:1000, SAP4G5, Sigma), Par3 (1:500, 07-330, Upstate), aPKC (1:500, C-20, Santa Cruz), HA (1:1000, clone 12CA5), Myc (1:1000, 9E10, Santa Cruz), ZO-1 (1:500, Z-R1, Zymed), E-cadherin (1:2000, #36, Pharmingen), β-catenin (1:2000, #14, Pharmingen), and α-catenin (1:1000, #5, Pharmingen).

    Techniques: Functional Assay, Produced, Isolation, In Vitro, Transfection, Cell Culture, Immunoprecipitation, Variant Assay, Binding Assay, Stable Transfection, Immunofluorescence, In Situ, Immunostaining

    Merlin binds α-catenin and is required for the formation of mature junctional belts in keratinocytes (A,B) Primary wild-type (A) or K14-Cre;Nf2 lox/lox (B) keratinocytes were cultured in media containing high calcium for the indicated times. Immunostaining revealed that Merlin (green) colocalizes with E-cadherin (red) and actin (white) in both immature PAs (arrows) and in the mature junctional belt (arrowheads) in wild-type cells. K14-Cre;Nf2 lox/lox keratinocytes form PAs but after 24 hours actin remains disorganized and a mature junctional belt does not form. Bars, 5 μm. (C) Immunoprecipitation of endogenous E-cadherin from immortalized wild-type PAM212 keratinocytes revealed that endogenous Merlin associates with the E-cadherin complex prior to calcium stimulation when only PAs are present (day 0). Although Merlin localizes to the mature junctional belt (A), the levels of Merlin that physically associate with the core cadherin complex diminish after calcium stimulation and junctional maturation. NRS, normal rabbit serum. (D) Flag-Merlin produced in vitro in Sf9 cells associates with bacterially produced GST-α-catenin but not with GST-E-cad-cyto or GST-β-catenin. Input equals 10% of the Flag-Merlin used in the binding assay. (E) Full-length (FL)-Merlin-HA, N-terminal (NT)-Merlin-HA and C-terminal (CT)-Merlin-HA were produced by in vitro transcription and translation (IVTT) and incubated with GST-α-catenin. Isolation of GST complexes and immunoblotting with anti-HA antibodies revealed that FL- and NT-Merlin-HA but not CT-Merlin-HA associated with GST-α-catenin in vitro . Input equals 10% of the IVTT products. (F) Immunoprecipitation of endogenous α-catenin from PAM212 keratinocytes cultured in calcium-depleted media (day 0) revealed the association of endogenous Merlin with endogenous α-catenin. .

    Journal: Developmental cell

    Article Title: The Nf2 Tumor Suppressor, Merlin, Regulates Epidermal Development Through the Establishment of a Junctional Polarity Complex

    doi: 10.1016/j.devcel.2010.10.008

    Figure Lengend Snippet: Merlin binds α-catenin and is required for the formation of mature junctional belts in keratinocytes (A,B) Primary wild-type (A) or K14-Cre;Nf2 lox/lox (B) keratinocytes were cultured in media containing high calcium for the indicated times. Immunostaining revealed that Merlin (green) colocalizes with E-cadherin (red) and actin (white) in both immature PAs (arrows) and in the mature junctional belt (arrowheads) in wild-type cells. K14-Cre;Nf2 lox/lox keratinocytes form PAs but after 24 hours actin remains disorganized and a mature junctional belt does not form. Bars, 5 μm. (C) Immunoprecipitation of endogenous E-cadherin from immortalized wild-type PAM212 keratinocytes revealed that endogenous Merlin associates with the E-cadherin complex prior to calcium stimulation when only PAs are present (day 0). Although Merlin localizes to the mature junctional belt (A), the levels of Merlin that physically associate with the core cadherin complex diminish after calcium stimulation and junctional maturation. NRS, normal rabbit serum. (D) Flag-Merlin produced in vitro in Sf9 cells associates with bacterially produced GST-α-catenin but not with GST-E-cad-cyto or GST-β-catenin. Input equals 10% of the Flag-Merlin used in the binding assay. (E) Full-length (FL)-Merlin-HA, N-terminal (NT)-Merlin-HA and C-terminal (CT)-Merlin-HA were produced by in vitro transcription and translation (IVTT) and incubated with GST-α-catenin. Isolation of GST complexes and immunoblotting with anti-HA antibodies revealed that FL- and NT-Merlin-HA but not CT-Merlin-HA associated with GST-α-catenin in vitro . Input equals 10% of the IVTT products. (F) Immunoprecipitation of endogenous α-catenin from PAM212 keratinocytes cultured in calcium-depleted media (day 0) revealed the association of endogenous Merlin with endogenous α-catenin. .

    Article Snippet: For IP and western blot the following antibodies were used: NF2 (1:1000, A-19 and C-18, Santa Cruz), Keratin 14 (1:5000, AF64, Covance), Actin (1:1000, AC40, Sigma), β-tubulin (1:1000, SAP4G5, Sigma), Par3 (1:500, 07-330, Upstate), aPKC (1:500, C-20, Santa Cruz), HA (1:1000, clone 12CA5), Myc (1:1000, 9E10, Santa Cruz), ZO-1 (1:500, Z-R1, Zymed), E-cadherin (1:2000, #36, Pharmingen), β-catenin (1:2000, #14, Pharmingen), and α-catenin (1:1000, #5, Pharmingen).

    Techniques: Cell Culture, Immunostaining, Immunoprecipitation, Produced, In Vitro, Binding Assay, Incubation, Isolation

    Expressions of signaling molecules of Wnt pathway, Wnt5b (A and B) and β-catenin (C and D). The expression of Wnt5b is observed in both tumor cells and CAFs, at the tumor surface (A) and the invasive margin (B). The increased expression and nuclear

    Journal: International Journal of Clinical and Experimental Pathology

    Article Title: Microenvironmental interactions and expression of molecular markers associated with epithelial-to-mesenchymal transition in colorectal carcinoma

    doi:

    Figure Lengend Snippet: Expressions of signaling molecules of Wnt pathway, Wnt5b (A and B) and β-catenin (C and D). The expression of Wnt5b is observed in both tumor cells and CAFs, at the tumor surface (A) and the invasive margin (B). The increased expression and nuclear

    Article Snippet: Simple immunohistochemical stains of β-catenin, Wnt5b, TGF-β1, p-Smad2, Smad4, ZEB1 and anti-SNAIL were done using a Bond polymer refine detection kit (DAB, 3,3’-diaminobenzidine tetrahydrochloride, Leica Biosystems).

    Techniques: Expressing

    Differentiation. HE staining of mammary tumors (A–B) and pulmonary metastases (C) from Apc +/1572T mice shows typical mammary glandular architecture and squamous differentiation. (D–F) Luminal epithelial differentiation as shown by cytokeratin 8 (Ck8) IHC staining. (G–I) Myoepithelial differentiation revealed by IHC staining with the Sma antibody. (J–L) IHC analysis with antibodies directed against cytokeratin 14 (Ck14) confirm the presence of squamous differentiation (hair follicle and skin cellular types). (M–O) β-catenin IHC analysis shows heterogeneous subcellular localization and intracellular accumulation with fewer cells characterized by positive nuclear staining. The results shown in this figure were confirmed in 12 independent primary tumors.

    Journal: PLoS Genetics

    Article Title: A Targeted Constitutive Mutation in the Apc Tumor Suppressor Gene Underlies Mammary But Not Intestinal Tumorigenesis

    doi: 10.1371/journal.pgen.1000547

    Figure Lengend Snippet: Differentiation. HE staining of mammary tumors (A–B) and pulmonary metastases (C) from Apc +/1572T mice shows typical mammary glandular architecture and squamous differentiation. (D–F) Luminal epithelial differentiation as shown by cytokeratin 8 (Ck8) IHC staining. (G–I) Myoepithelial differentiation revealed by IHC staining with the Sma antibody. (J–L) IHC analysis with antibodies directed against cytokeratin 14 (Ck14) confirm the presence of squamous differentiation (hair follicle and skin cellular types). (M–O) β-catenin IHC analysis shows heterogeneous subcellular localization and intracellular accumulation with fewer cells characterized by positive nuclear staining. The results shown in this figure were confirmed in 12 independent primary tumors.

    Article Snippet: Antibodies employed for IHC analysis include: β-catenin (1∶2000, 1247-1, Epitomics), Troma1 which recognizes a Ck8 epitope (1∶400, Hybridoma Bank), Sma (1∶200, M0851, DakoCytmomation), Ck14 (1∶10000, PRB-155P, Covance), Ck6 (1∶5000, PRB-169P, Covance).

    Techniques: Staining, Mouse Assay, Immunohistochemistry

    Biochemical characterization of the targeted Apc 1572T allele. (A) Schematic representation of the APC tumor suppressor protein, its functional domains, and the truncated proteins resulting from the Apc 1572T, Apc 1638N, and Apc 1638T targeted alleles. Only residual amounts (2%) of the truncated Apc1638N protein are encoded by the targeted allele, as shown by immuno-precipitation analysis of Apc 1638N/1638N ES lines [5] . (B) β-catenin/TCF reporter assay (TOP-FLASH) analysis of Apc +/+ (1) and Apc -mutant ES cell lines: Apc 1638T/1638T (2); Apc 1572T/1572T (3); Apc 1638N/1638N (4). Each bar represents the average measurement of the luciferase units from triplicate assays. For each cell line, 3 independent experiments were performed with the TOP (filled bars) and FOP (empty bars) reporter constructs. The bold figures represent the average TOP/FOP ratio of all independent experiments. Depicted error bars correspond to standard deviation. In brief, ES cells were plated on dishes coated with MEFs and subsequently transfected by lipofection with either the TOP-FLASH or FOP-FLASH reporter constructs [10] together with the Renilla luciferase vector for normalization purposes. (C) Immuno-precipitation (IP) analysis of Apc-bound β-catenin in Ap c-mutant ES cell lines. For comparative purposes, immuno-precipitates obtained from equal amounts of total cellular lysates were loaded.

    Journal: PLoS Genetics

    Article Title: A Targeted Constitutive Mutation in the Apc Tumor Suppressor Gene Underlies Mammary But Not Intestinal Tumorigenesis

    doi: 10.1371/journal.pgen.1000547

    Figure Lengend Snippet: Biochemical characterization of the targeted Apc 1572T allele. (A) Schematic representation of the APC tumor suppressor protein, its functional domains, and the truncated proteins resulting from the Apc 1572T, Apc 1638N, and Apc 1638T targeted alleles. Only residual amounts (2%) of the truncated Apc1638N protein are encoded by the targeted allele, as shown by immuno-precipitation analysis of Apc 1638N/1638N ES lines [5] . (B) β-catenin/TCF reporter assay (TOP-FLASH) analysis of Apc +/+ (1) and Apc -mutant ES cell lines: Apc 1638T/1638T (2); Apc 1572T/1572T (3); Apc 1638N/1638N (4). Each bar represents the average measurement of the luciferase units from triplicate assays. For each cell line, 3 independent experiments were performed with the TOP (filled bars) and FOP (empty bars) reporter constructs. The bold figures represent the average TOP/FOP ratio of all independent experiments. Depicted error bars correspond to standard deviation. In brief, ES cells were plated on dishes coated with MEFs and subsequently transfected by lipofection with either the TOP-FLASH or FOP-FLASH reporter constructs [10] together with the Renilla luciferase vector for normalization purposes. (C) Immuno-precipitation (IP) analysis of Apc-bound β-catenin in Ap c-mutant ES cell lines. For comparative purposes, immuno-precipitates obtained from equal amounts of total cellular lysates were loaded.

    Article Snippet: Antibodies employed for IHC analysis include: β-catenin (1∶2000, 1247-1, Epitomics), Troma1 which recognizes a Ck8 epitope (1∶400, Hybridoma Bank), Sma (1∶200, M0851, DakoCytmomation), Ck14 (1∶10000, PRB-155P, Covance), Ck6 (1∶5000, PRB-169P, Covance).

    Techniques: Functional Assay, Immunoprecipitation, Reporter Assay, Mutagenesis, Luciferase, Construct, Standard Deviation, Transfection, Plasmid Preparation

    Phenotypic characterization of Apc +/1572T mice: mammary adenocarcinomas are composed by mixed differentiation lineages with heterogeneous patterns of β-catenin intracellular accumulation and subcellular localization. Survival curves of (A) female and (B) male Apc +/1572T mice, respectively. The black, green and red lines are representative of mice in the 129Ola, F1 B6x129Ola, and B6 respectively. Please note that in these graphs, age of death represents the moment at which, due to the presence of signs of discomfort or because the tumor size exceeded 2 cm 3 , mice had to be euthanized according to institutional and national regulations. (C) Macroscopic image of the appearance of the mammary adenocarcinomas characteristic of the Apc 1572T model. (D) Examples of global digital microscopy scans of two mammary adenocarcinomas from Apc +/1572T mice illustrative of the multi-lineage nature of these lesions.

    Journal: PLoS Genetics

    Article Title: A Targeted Constitutive Mutation in the Apc Tumor Suppressor Gene Underlies Mammary But Not Intestinal Tumorigenesis

    doi: 10.1371/journal.pgen.1000547

    Figure Lengend Snippet: Phenotypic characterization of Apc +/1572T mice: mammary adenocarcinomas are composed by mixed differentiation lineages with heterogeneous patterns of β-catenin intracellular accumulation and subcellular localization. Survival curves of (A) female and (B) male Apc +/1572T mice, respectively. The black, green and red lines are representative of mice in the 129Ola, F1 B6x129Ola, and B6 respectively. Please note that in these graphs, age of death represents the moment at which, due to the presence of signs of discomfort or because the tumor size exceeded 2 cm 3 , mice had to be euthanized according to institutional and national regulations. (C) Macroscopic image of the appearance of the mammary adenocarcinomas characteristic of the Apc 1572T model. (D) Examples of global digital microscopy scans of two mammary adenocarcinomas from Apc +/1572T mice illustrative of the multi-lineage nature of these lesions.

    Article Snippet: Antibodies employed for IHC analysis include: β-catenin (1∶2000, 1247-1, Epitomics), Troma1 which recognizes a Ck8 epitope (1∶400, Hybridoma Bank), Sma (1∶200, M0851, DakoCytmomation), Ck14 (1∶10000, PRB-155P, Covance), Ck6 (1∶5000, PRB-169P, Covance).

    Techniques: Mouse Assay, Microscopy

    GTP-Rac1 and pAKT did not participate in TCF/β-catenin signaling pathway after SLP-2 inhibition in NSCLC cells. a Western blot analyses for the expressions of pAkt in A549, H460, H838, and H157 cells treated with either AdSLP2i or AdCtrl at m.o.i. of 100. b Downregulation of SLP-2 increased the GTP-Rac1 proteins in A549 and H157 cells, but decreased the GTP-Rac1 proteins in H838 and H460 cells as determined by pull-down assay. Total cell lysates were subjected to the pull-down assay for GTP-Rac1 activities. Total cell lysates were analyzed for Rac1 expression as a loading control. The ratio of GTP-Rac1/total Rac1 in cells was analyzed by densitometry of the blot

    Journal: Cell Death & Disease

    Article Title: Stomatin-like protein 2 regulates survivin expression in non-small cell lung cancer cells through β-catenin signaling pathway

    doi: 10.1038/s41419-018-0461-9

    Figure Lengend Snippet: GTP-Rac1 and pAKT did not participate in TCF/β-catenin signaling pathway after SLP-2 inhibition in NSCLC cells. a Western blot analyses for the expressions of pAkt in A549, H460, H838, and H157 cells treated with either AdSLP2i or AdCtrl at m.o.i. of 100. b Downregulation of SLP-2 increased the GTP-Rac1 proteins in A549 and H157 cells, but decreased the GTP-Rac1 proteins in H838 and H460 cells as determined by pull-down assay. Total cell lysates were subjected to the pull-down assay for GTP-Rac1 activities. Total cell lysates were analyzed for Rac1 expression as a loading control. The ratio of GTP-Rac1/total Rac1 in cells was analyzed by densitometry of the blot

    Article Snippet: We further utilized the antibody recognizing active form of β-catenin with dephosphorylation on Ser37 or Thr41 (05-665, Anti-ABC, Merck) and immunofluorescence to confirm the change of β-catenin localization after AdSLP2i or AdCtrl - transfection (Fig. ).

    Techniques: Inhibition, Western Blot, Pull Down Assay, Expressing

    Western blotting and immunofluorescence analysis for the nuclear localization of β-catenin by SLP-2 inhibition in NSCLC cells. a Western blot analyses for the expressions of nuclear active β-catenin in A549, H460, H838, and H157 cells treated with either AdSLP2i or AdCtrl at m.o.i. of 100. b Immunofluorescence analyses for the nuclear localization of β-catenin in A549, H460, H838, and H157 cells treated with either AdSLP2i or AdCtrl at m.o.i. of 100. On day 3 post infection, the nuclear localization of β-catenin was clearly reduced in all four AdSLP2i-treated cell lines

    Journal: Cell Death & Disease

    Article Title: Stomatin-like protein 2 regulates survivin expression in non-small cell lung cancer cells through β-catenin signaling pathway

    doi: 10.1038/s41419-018-0461-9

    Figure Lengend Snippet: Western blotting and immunofluorescence analysis for the nuclear localization of β-catenin by SLP-2 inhibition in NSCLC cells. a Western blot analyses for the expressions of nuclear active β-catenin in A549, H460, H838, and H157 cells treated with either AdSLP2i or AdCtrl at m.o.i. of 100. b Immunofluorescence analyses for the nuclear localization of β-catenin in A549, H460, H838, and H157 cells treated with either AdSLP2i or AdCtrl at m.o.i. of 100. On day 3 post infection, the nuclear localization of β-catenin was clearly reduced in all four AdSLP2i-treated cell lines

    Article Snippet: We further utilized the antibody recognizing active form of β-catenin with dephosphorylation on Ser37 or Thr41 (05-665, Anti-ABC, Merck) and immunofluorescence to confirm the change of β-catenin localization after AdSLP2i or AdCtrl - transfection (Fig. ).

    Techniques: Western Blot, Immunofluorescence, Inhibition, Infection

    Model of Egfr activity requirements during intestinal tumorigenesis. Red dots represent nuclear β-catenin-positive cells. Solid green lines indicate evidence for Egfr activity during establishment and adenocarcinoma expansion. Dashed green lines indicate that a requirement for Egfr activity during adenoma expansion, progression, and invasion has yet to be demonstrated conclusively.

    Journal: Proceedings of the National Academy of Sciences of the United States of America

    Article Title: Importance of epidermal growth factor receptor signaling in establishment of adenomas and maintenance of carcinomas during intestinal tumorigenesis

    doi: 10.1073/pnas.032678499

    Figure Lengend Snippet: Model of Egfr activity requirements during intestinal tumorigenesis. Red dots represent nuclear β-catenin-positive cells. Solid green lines indicate evidence for Egfr activity during establishment and adenocarcinoma expansion. Dashed green lines indicate that a requirement for Egfr activity during adenoma expansion, progression, and invasion has yet to be demonstrated conclusively.

    Article Snippet: Microwave antigen retrieval with citrate buffer and the Mouse-on-Mouse kit (Vector Laboratories) were used in conjunction with primary Ab for β-catenin (Transduction Laboratories, Lexington, KY, clone 14, 1:500 dilution) or Ki67 (NovoCastra, Newcastle, U.K., 1:100 dilution).

    Techniques: Activity Assay

    Early ileal lesions in Apc Min mice. ( A and B ) Adjacent sections of an Egfr + cystic microadenoma. ( C ) Egfr + crypt, containing a cluster of three nuclear β-catenin-positive cells, flanked by normal crypts. ( D ) Close up of boxed crypt from C . ( E ) Egfr wa2 crypt with several β-catenin-positive nuclei. ( A ) H E staining. ( B – E ) β-catenin immunohistochemistry. Arrowheads in D mark the β-catenin-positive nuclei.

    Journal: Proceedings of the National Academy of Sciences of the United States of America

    Article Title: Importance of epidermal growth factor receptor signaling in establishment of adenomas and maintenance of carcinomas during intestinal tumorigenesis

    doi: 10.1073/pnas.032678499

    Figure Lengend Snippet: Early ileal lesions in Apc Min mice. ( A and B ) Adjacent sections of an Egfr + cystic microadenoma. ( C ) Egfr + crypt, containing a cluster of three nuclear β-catenin-positive cells, flanked by normal crypts. ( D ) Close up of boxed crypt from C . ( E ) Egfr wa2 crypt with several β-catenin-positive nuclei. ( A ) H E staining. ( B – E ) β-catenin immunohistochemistry. Arrowheads in D mark the β-catenin-positive nuclei.

    Article Snippet: Microwave antigen retrieval with citrate buffer and the Mouse-on-Mouse kit (Vector Laboratories) were used in conjunction with primary Ab for β-catenin (Transduction Laboratories, Lexington, KY, clone 14, 1:500 dilution) or Ki67 (NovoCastra, Newcastle, U.K., 1:100 dilution).

    Techniques: Mouse Assay, Staining, Immunohistochemistry

    Analysis of Akt and Wnt/β-catenin signaling in Tcl1- deficient and -overexpressing ES cells. (A) Western blot analysis of GSK, Akt, and β-catenin in wild-type (WT), Tcl1 −/− (KO) #2 and #4, Tcl1 −/− (CAG- Tcl1 ) #1 and #3, and Tcl1 −/− (CAG-EGFP) #5 ES cells. (B) Western blot analysis of active β-catenin, Oct3/4, and HSP90 in the cytoplasmic (C) and nuclear (N) fractions of wild-type (WT), Tcl1 −/− (KO) #2 and #4, and Tcl1 −/− (CAG- Tcl1 ) #10 and #1 ES cells. Tcl1 −/− (CAG- Tcl1 ) #10 and #1 were derived from Tcl1 −/− (KO) #2 and #4, respectively. Proper fractionation was confirmed by western blotting of Oct3/4 and HSP90, which localize to the nucleus and cytoplasm, respectively. Be8cause active β-catenin levels in the nuclear fractions were much lower than those in the cytoplasmic fractions, active β-catenin in the nuclear fractions was detected by approximately two-fold longer exposure compared with that in the cytoplasmic fractions. (C) TOPflash assay. P values of wild-type ES cells (WT) compared with Tcl1 −/− (CAG- Tcl1 ) #1 and #3 ES cells were less than 0.01. P values of Tcl1 −/− (KO) #4 and #5 ES cells compared with Tcl1 −/− (CAG- Tcl1 ) #1 and #3 ES cells were less than 0.02.

    Journal: PLoS ONE

    Article Title: Functional Analysis of Tcl1 Using Tcl1-Deficient Mouse Embryonic Stem Cells

    doi: 10.1371/journal.pone.0071645

    Figure Lengend Snippet: Analysis of Akt and Wnt/β-catenin signaling in Tcl1- deficient and -overexpressing ES cells. (A) Western blot analysis of GSK, Akt, and β-catenin in wild-type (WT), Tcl1 −/− (KO) #2 and #4, Tcl1 −/− (CAG- Tcl1 ) #1 and #3, and Tcl1 −/− (CAG-EGFP) #5 ES cells. (B) Western blot analysis of active β-catenin, Oct3/4, and HSP90 in the cytoplasmic (C) and nuclear (N) fractions of wild-type (WT), Tcl1 −/− (KO) #2 and #4, and Tcl1 −/− (CAG- Tcl1 ) #10 and #1 ES cells. Tcl1 −/− (CAG- Tcl1 ) #10 and #1 were derived from Tcl1 −/− (KO) #2 and #4, respectively. Proper fractionation was confirmed by western blotting of Oct3/4 and HSP90, which localize to the nucleus and cytoplasm, respectively. Be8cause active β-catenin levels in the nuclear fractions were much lower than those in the cytoplasmic fractions, active β-catenin in the nuclear fractions was detected by approximately two-fold longer exposure compared with that in the cytoplasmic fractions. (C) TOPflash assay. P values of wild-type ES cells (WT) compared with Tcl1 −/− (CAG- Tcl1 ) #1 and #3 ES cells were less than 0.01. P values of Tcl1 −/− (KO) #4 and #5 ES cells compared with Tcl1 −/− (CAG- Tcl1 ) #1 and #3 ES cells were less than 0.02.

    Article Snippet: The primary antibodies used were: rabbit anti-Akt (Cat#CST9272; Cell Signaling, Beverly, MA), rabbit anti-phospho-Akt (Ser473) (Cat#CST9271; Cell Signaling), rabbit anti-phospho-GSK3α/β (Ser21/9) (Cat#CST9331; Cell Signaling), rabbit anti-phospho-β-catenin (Thr41/Ser45) (Cat#CST9565; Cell Signaling), rabbit anti-phospho-β-catenin (Ser33/37/Thr41) (Cat#CST9561; Cell Signaling), rabbit anti-Tcl1 (Cat#CST4042; Cell Signaling), mouse monoclonal anti-β-catenin (Cat#610153; BD Biosciences, Franklin Lakes, NJ), mouse monoclonal antibody recognizing active β-catenin that is unphosphorylated at Ser37 and Thr41 (mAb 8E7) (Cat#05-665; Merck Millipore), mouse monoclonal anti-HSP90 (Cat#ADI-SPA-830; Enzo Life Sciences, Farmingdale, NY), mouse monoclonal anti-Oct3/4 (Cat#sc-5279; Santa Cruz Biotechnol., Santa Cruz, CA), and horseradish peroxidise (HRP)-conjugated mouse monoclonal anti-β-actin (Cat#ab20272; Abcam, Cambridge, MA).

    Techniques: Western Blot, Derivative Assay, Fractionation, TOPFlash assay

    Effects of PA on the expression of E-cadherin, β-catenin and COX-2 in HO-8910 cells. After treatment with or without different concentrations of PA, corresponding proteins were determined at 48 h.

    Journal: International Journal of Clinical and Experimental Pathology

    Article Title: Inhibition of ovarian cancer proliferation and invasion by pachymic acid

    doi:

    Figure Lengend Snippet: Effects of PA on the expression of E-cadherin, β-catenin and COX-2 in HO-8910 cells. After treatment with or without different concentrations of PA, corresponding proteins were determined at 48 h.

    Article Snippet: Total protein (20 μg) was loaded to sodium dodecyl sulfate-polyacrylamide electrophoresis gels (SDS-PAGE) and then transferred to nitrocellulose filter membrane, which was incubated with anti-E cadherin, β-catenin, COX-2 and GAPDH antibodies at 4°C for 12 h. After that, the membranes were incubated with corresponding horseradish-peroxidase-conjugated secondary antibodies and the immunoreactive band was visualized with ECL-detecting reagents (Beyotime, Jiangsu, China).

    Techniques: Expressing

    Up-regulation of β-catenin mediates 5-HT-induced PPAR γ reduction

    Journal: Oncotarget

    Article Title: 5-HT induces PPAR γ reduction and proliferation of pulmonary artery smooth muscle cells via modulating GSK-3β/β-catenin pathway

    doi: 10.18632/oncotarget.20582

    Figure Lengend Snippet: Up-regulation of β-catenin mediates 5-HT-induced PPAR γ reduction

    Article Snippet: The primers for β-catenin (Sangon Biotech, Shanghai, China) were: forward 5′-CCACGACTAGTTCAGCTGCTTGTAC-3′ and reverse 5′-ACTGCACAAACAGTGGAATGGTATT-3′.

    Techniques:

    5-HT inactivates GSK-3β, up-regulates β-catenin and reduces PPAR γ by PI3K/Akt pathway

    Journal: Oncotarget

    Article Title: 5-HT induces PPAR γ reduction and proliferation of pulmonary artery smooth muscle cells via modulating GSK-3β/β-catenin pathway

    doi: 10.18632/oncotarget.20582

    Figure Lengend Snippet: 5-HT inactivates GSK-3β, up-regulates β-catenin and reduces PPAR γ by PI3K/Akt pathway

    Article Snippet: The primers for β-catenin (Sangon Biotech, Shanghai, China) were: forward 5′-CCACGACTAGTTCAGCTGCTTGTAC-3′ and reverse 5′-ACTGCACAAACAGTGGAATGGTATT-3′.

    Techniques:

    GSK-3β insufficiency or inhibition of proteasome activity up-regulates β-catenin and reduces PPAR γ

    Journal: Oncotarget

    Article Title: 5-HT induces PPAR γ reduction and proliferation of pulmonary artery smooth muscle cells via modulating GSK-3β/β-catenin pathway

    doi: 10.18632/oncotarget.20582

    Figure Lengend Snippet: GSK-3β insufficiency or inhibition of proteasome activity up-regulates β-catenin and reduces PPAR γ

    Article Snippet: The primers for β-catenin (Sangon Biotech, Shanghai, China) were: forward 5′-CCACGACTAGTTCAGCTGCTTGTAC-3′ and reverse 5′-ACTGCACAAACAGTGGAATGGTATT-3′.

    Techniques: Inhibition, Activity Assay

    Up-regulation of β-catenin and subsequent PPAR γ reduction mediates 5-HT-stimulated PASMCs proliferation

    Journal: Oncotarget

    Article Title: 5-HT induces PPAR γ reduction and proliferation of pulmonary artery smooth muscle cells via modulating GSK-3β/β-catenin pathway

    doi: 10.18632/oncotarget.20582

    Figure Lengend Snippet: Up-regulation of β-catenin and subsequent PPAR γ reduction mediates 5-HT-stimulated PASMCs proliferation

    Article Snippet: The primers for β-catenin (Sangon Biotech, Shanghai, China) were: forward 5′-CCACGACTAGTTCAGCTGCTTGTAC-3′ and reverse 5′-ACTGCACAAACAGTGGAATGGTATT-3′.

    Techniques:

    Nodal LAM’s immunohistochemical expression of HMB45 is typically focal with a granular or gritty quality that fills just a portion of the cytoplasm (A, 400X). β-catenin consistently strongly and diffusely labels the cytoplasm of nodal

    Journal: The American journal of surgical pathology

    Article Title: Incidental Nodal Lymphangioleiomyomatosis Is Not a Harbinger of Pulmonary Lymphangioleiomyomatosis: A Study of 19 Cases with Evaluation of Diagnostic Immunohistochemistry

    doi: 10.1097/PAS.0000000000000470

    Figure Lengend Snippet: Nodal LAM’s immunohistochemical expression of HMB45 is typically focal with a granular or gritty quality that fills just a portion of the cytoplasm (A, 400X). β-catenin consistently strongly and diffusely labels the cytoplasm of nodal

    Article Snippet: IHC was performed using a panel of three antibodies: HMB45 (clone HMB45, no dilution, Ventana Medical Systems, Tucson, AZ), A103 (clone A103, no dilution; Ventana Medical Systems) and β-catenin (clone 14, no dilution, Ventana).

    Techniques: Laser Capture Microdissection, Immunohistochemistry, Expressing

    Effect of galantamine and/or vildagliptin on Wnt/β-catenin pathway. Effect of different doses of galantamine (Galan 2.5, 5 10 mg/kg), vildagliptin (Vilda 3, 10 30 mg/kg) and their combination (Galan5 + Vilda 30) on the hepatic and muscular contents of p-GSK-3β (A, B) and β-catenin (C, D) in n-STZ diabetic rats. Drugs were gavaged orally for four weeks. Values are means of 10 rats ± S.E.M as compared with normal control (*), diabetic control (#), Galan5 (η), Galan10 (ɸ) and Vilda30 (Ψ)-treated groups (one-way ANOVA followed by Tukey post hoc test) at P

    Journal: PLoS ONE

    Article Title: Antidiabetic Effect of Galantamine: Novel Effect for a Known Centrally Acting Drug

    doi: 10.1371/journal.pone.0134648

    Figure Lengend Snippet: Effect of galantamine and/or vildagliptin on Wnt/β-catenin pathway. Effect of different doses of galantamine (Galan 2.5, 5 10 mg/kg), vildagliptin (Vilda 3, 10 30 mg/kg) and their combination (Galan5 + Vilda 30) on the hepatic and muscular contents of p-GSK-3β (A, B) and β-catenin (C, D) in n-STZ diabetic rats. Drugs were gavaged orally for four weeks. Values are means of 10 rats ± S.E.M as compared with normal control (*), diabetic control (#), Galan5 (η), Galan10 (ɸ) and Vilda30 (Ψ)-treated groups (one-way ANOVA followed by Tukey post hoc test) at P

    Article Snippet: Phosphorylated glycogen synthase kinase-3β (p-GSK-3β) at serine 9 and β-catenin were determined using the corresponding ELISA kit (Enzo Life Sciences, Plymouth Meeting, PA, USA).

    Techniques:

    D-DT-mediated β-catenin stabilization contributes to, but is not wholly responsible for, D-DT-dependent COX-2 expression A, Cells were plated in 6-well plates and were transfected the following day with pCDNA3.1/D-DT or pCDNA3.1/β-catenin

    Journal: Molecular cancer research : MCR

    Article Title: The MIF homolog, D-dopachrome tautomerase (D-DT), promotes COX-2 expression through ?-catenin-dependent and independent mechanisms

    doi: 10.1158/1541-7786.MCR-10-0101

    Figure Lengend Snippet: D-DT-mediated β-catenin stabilization contributes to, but is not wholly responsible for, D-DT-dependent COX-2 expression A, Cells were plated in 6-well plates and were transfected the following day with pCDNA3.1/D-DT or pCDNA3.1/β-catenin

    Article Snippet: Adenovirus for D-DT was prepared as previously described ( ). β-catenin and constitutively active MKK7 recombinant adenoviruses were purchased from Vector Biolabs (San Diego, CA) and were used to infect cells at ~ 5 × 107 virus particles/ml.

    Techniques: Expressing, Transfection

    Reduced β-catenin expression and activity in D-DT-deficient cells A, HT-29 cells were plated in 6-well plates for 24 hours followed by oligotransfection with 50 nM scrambled shRNA oligos or 50 nM D-DT-specific shRNA oligos for an additional 48

    Journal: Molecular cancer research : MCR

    Article Title: The MIF homolog, D-dopachrome tautomerase (D-DT), promotes COX-2 expression through ?-catenin-dependent and independent mechanisms

    doi: 10.1158/1541-7786.MCR-10-0101

    Figure Lengend Snippet: Reduced β-catenin expression and activity in D-DT-deficient cells A, HT-29 cells were plated in 6-well plates for 24 hours followed by oligotransfection with 50 nM scrambled shRNA oligos or 50 nM D-DT-specific shRNA oligos for an additional 48

    Article Snippet: Adenovirus for D-DT was prepared as previously described ( ). β-catenin and constitutively active MKK7 recombinant adenoviruses were purchased from Vector Biolabs (San Diego, CA) and were used to infect cells at ~ 5 × 107 virus particles/ml.

    Techniques: Expressing, Activity Assay, shRNA

    CSN5 participates in the regulation of COX-2 and β-catenin. A, Nonspecific control (Scr) and CSN5-specific shRNA-transfected cells were harvested, lysed and immunoblotted for COX-2, CSN5 and α-tubulin. B, 24 hours after transfection with

    Journal: Molecular cancer research : MCR

    Article Title: The MIF homolog, D-dopachrome tautomerase (D-DT), promotes COX-2 expression through ?-catenin-dependent and independent mechanisms

    doi: 10.1158/1541-7786.MCR-10-0101

    Figure Lengend Snippet: CSN5 participates in the regulation of COX-2 and β-catenin. A, Nonspecific control (Scr) and CSN5-specific shRNA-transfected cells were harvested, lysed and immunoblotted for COX-2, CSN5 and α-tubulin. B, 24 hours after transfection with

    Article Snippet: Adenovirus for D-DT was prepared as previously described ( ). β-catenin and constitutively active MKK7 recombinant adenoviruses were purchased from Vector Biolabs (San Diego, CA) and were used to infect cells at ~ 5 × 107 virus particles/ml.

    Techniques: shRNA, Transfection