anti-yy1 Search Results


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  • 99
    Millipore anti yy1 k411me1
    <t>YY1</t> knockdown reduces viral integration into the host genome. (A) Knockdown of YY1 by siRNA was confirmed by Western blotting. (B and C) The amount of total viral cDNA (B) and two-LTR circle (C) of pQEGFP (siRNA no. 1006) and pLNΔAG (siRNA no. 1099) viral vectors is shown. NIH 3T3 cells were transfected twice with YY1 siRNA no. 1006 or no. 1099 and then infected with pQEGFP (MOI of 0.1) or pLNΔAG (MOI of 1). Cellular DNA was extracted at 0, 4, 10, 24, and 48 h and subjected to qPCR analysis. Copy number per cell was calculated by normalizing the value of the viral sequence with that of GAPDH (glyceraldehyde-3-phosphate dehydrogenase). Error bars represent the standard deviations of the results of at least three experiments. Primers used for PCR for GAPDH were 5′-TGTGATGGGTGTGAACCACGAGAA-3′ and 5′-GAGCCCTTCCACAATGCCAAAGTT-3′. Asterisks indicate that values of YY1 siRNA compared to those of scrambled RNA were significantly different (*, P = 0.038, and **, P = 0.031 in panel B; *, P = 0.022, **, P = 0.041, and ***, P = 0.047 in panel C). (D) The integrated form of viral DNA was quantified 14 days postinfection by qPCR and normalized as described for the results presented in panel C. Asterisks indicate significance of values of YY1 siRNA versus those of scrambled RNA (*, P = 0.014, **, P = 0.016, and ***, P
    Anti Yy1 K411me1, supplied by Millipore, used in various techniques. Bioz Stars score: 99/100, based on 2 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    92
    Atlas Antibodies anti yy1
    <t>YY1</t> identify a dominant phenotypic clone in ERα BC A) RIs for the YY1 enhancer within all the individual patients included in the current study YY1 enhancer location with its 3D interactions are shown in the top right inset B) YY1 enhancer ranking analysis of available Epigenome Roadmap H3K27ac datasets. Tissues are displayed from the strongest to the weakest YY1 enhancer activity (based on RI). Representative IHC analysis of normal tissues stained with a YY1 antibody are shown C) eKO cell lines were generated by deleting a 2.4kb containing YY1-A enhancers in MCF7 cells. Actual karyotyping is shown in the bottom panel was performed on 10 individual cells D) YY1 expression in control and eKO cell lines was measured using RT-qPCR. Lines and error bars represent average and 95% CI of five independent experiments. Significance was calculated with a one-way ANOVA followed by Tukey’s test E) Top left: YY1 expression in ERα-positive breast cancer compared to normal breast tissue. Median, lowest and highest values are reported. Top right: YY1 prognostic value in triple negative breast cancers. Bottom left: YY1 prognostic value in luminal breast cancers. Confidence interval (1.19-1.76). Bottom right: multivariate correction for the luminal breast cancer dataset is shown. Analyses included 1476 ERα-positive and 432 ERα-negative patients. Comparison of survival curves was performed using a Log-rank (Mantel-Cox) test. F) IHC analysis of normal breast tissues highlights YY1 functional subclones in normal breast. Similar results were observed in 10 independent clinical specimens from independent individuals G) IHC analysis of ERα positive invasive ductal carcinomas identify YY1 positive clones as the dominant clonal population (Scale bars, 50 μm).
    Anti Yy1, supplied by Atlas Antibodies, used in various techniques. Bioz Stars score: 92/100, based on 6 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    94
    Abcam anti yy1
    Expanded polyQ perturbs the function of <t>YY1</t> to derepress Fuz expression A Knockdown of YY1 expression upregulated Fuz expression in HEK293 cells. Lower panel shows the quantification of Fuz transcript expression relative to controls. Error bars represent s.e.m., n = 3. Statistical analysis was performed using two‐tailed unpaired Student's t ‐test. * P
    Anti Yy1, supplied by Abcam, used in various techniques. Bioz Stars score: 94/100, based on 74 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    94
    Abcam anti yy1 antibody
    CXCR4 regulates let-7a expression via <t>YY1.</t>
    Anti Yy1 Antibody, supplied by Abcam, used in various techniques. Bioz Stars score: 94/100, based on 61 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    94
    Santa Cruz Biotechnology anti yy1
    FABP7 promoter cis -regulatory region identified by promoter deletion analysis . Numbering starts from the FABP7 transcription start site [ 17 ]. Oligonucleotides designated upstream (UP), middle (MP) and downstream (DP) probes are shown along with putative consensus binding sites for OCT1, C/EBP-α, BRN2, OCT6, NFI, C/EBP-β, SP1, and <t>YY1.</t>
    Anti Yy1, supplied by Santa Cruz Biotechnology, used in various techniques. Bioz Stars score: 94/100, based on 537 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    91
    Cell Signaling Technology Inc anti yy1
    G9a silences the expression of HEPH via assembling a co-repressor complex with <t>YY1</t> and HDAC1. a – c MCF-7 cells were transfected with two independent YY1 siRNAs. After 48 h, HEPH protein and mRNA levels and HEPH promoter luciferase activity were examined. Expression of the transfected constructs is shown in the western blotting analysis. d pGL3- HEPH promoter and the indicated constructs were co-transfected into MDA-MB-231 cells. Twenty-four hours after transfection, cell extracts were assayed for luciferase activity. e Silencing and overexpression of HDAC1, but not HDAC2, contributed to the upregulation or downregulation of HEPH mRNA and protein level, respectively. The HDAC1-specific inhibitor MS275 was synergetic with UNC0638 in increasing HEPH expression in a time-dependent manner. f , g The abundance of H3K9-me2 and the binding levels of G9a and HDAC1 in the Pro2 region of the HEPH promoter were determined by ChIP in G9a knockdown or overexpressed cells treated with siYY1. The results are presented as means ± SD from three independent experiments. Two-tailed unpaired Student’s T -test was performed. * P
    Anti Yy1, supplied by Cell Signaling Technology Inc, used in various techniques. Bioz Stars score: 91/100, based on 25 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    91
    Proteintech anti yy1
    <t>YY1</t> depletion represses PCa cell viability and colony forming in a miR-146a-assisted manner. (A–C) Western blot assays were performed to demonstrate the expression effect of siYY1, pCMV6 and sh-YY1 on PCa cell lines. GAPDH was used as endogenous control. (D and E) miR-146a expression by qRT-PCR verified the inhibitory efficiency of si-YY1+antiNC/si-YY1+anti-miR-146a in both cell lines. The cell lines were transfected with si-YY1/si-NC, pCMV6-YY1/pCMV6, and si-YY1+anti-NC/si-YY1+anti-miR-146a. Cell viability and colony forming were measured by MTT (F-K) and colony formation assays (L-N). (O) GSEA showed enriched expression of gene sets involved in cell proliferation in YY1 knocked-down cells. * P
    Anti Yy1, supplied by Proteintech, used in various techniques. Bioz Stars score: 91/100, based on 13 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    93
    Abcam anti yy1 antibody bioworld
    <t>YY1</t> expression for NAFLD at different stages. a The YY1 mRNA levels in the control, steatosis, non-defining NASH, and NASH groups were detected with the quantitative real-time PCR. ** p
    Anti Yy1 Antibody Bioworld, supplied by Abcam, used in various techniques. Bioz Stars score: 93/100, based on 4 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    94
    Abcam anti yy1 antibody epr4651
    <t>YY1</t> expression for NAFLD at different stages. a The YY1 mRNA levels in the control, steatosis, non-defining NASH, and NASH groups were detected with the quantitative real-time PCR. ** p
    Anti Yy1 Antibody Epr4651, supplied by Abcam, used in various techniques. Bioz Stars score: 94/100, based on 2 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    yy1  (Abcam)
    92
    Abcam yy1
    Effect of miR-29a on <t>YY1</t> expression and cell behavior following IL-13 treatment. (A) MTT assay. miR-29a inhibited the cell proliferation promoted by IL-13. (B) miR-29a inhibits cell invasion in A549 cells by Transwell chamber assay. Overexpression of miR-29a or knockdown of miR-29a was carried out by transfection of A549 cells and cells were then treated with IL-13. Cell invasion ability was measured by Transwell assay. (C) Quantification of the number of invasive cells from B. (D) Measurement of the YY1 mRNA level in A549 cells treated with miR-29a overexpression vector or IL-13 stimulation. The mRNA level of YY1 was measured by RT-qPCR and normalized by GAPDH. (E) Determination of the YY1 protein level in A549 cells treated with miR-29a overexpression vector or IL-13 stimulation. The protein level of YY1 was determined by western blot analysis. (F) Quantification of the protein expression level of YY1 from E as analyzed by ImageJ software *P
    Yy1, supplied by Abcam, used in various techniques. Bioz Stars score: 92/100, based on 181 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    85
    Becton Dickinson anti yy1 antibodies
    A purified <t>YY1</t> complex methylates histone H4. Anti-Flag immunoprecipitates obtained either from HeLa cells transfected with plasmids expressing Flag–DRBP76 ( A ) or from immunopurification of Flag–YY1 ( B ) were assayed for methylase activity in the presence of core histones. “Flag competitor” corresponds to the addition of excess Flag peptide immunogen. Negative controls include immunoprecipitates from mock transfected cells and anti-Flag immunopurified materials from cells transduced with adenovirus expressing the GFP. Purified recombinant SUV39H1 was used as a positive control for histone H3 methylation. Each blot was stained with Amido black to ensure proper protein transfer.
    Anti Yy1 Antibodies, supplied by Becton Dickinson, used in various techniques. Bioz Stars score: 85/100, based on 1 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    93
    Santa Cruz Biotechnology anti yy1 h414
    A purified <t>YY1</t> complex methylates histone H4. Anti-Flag immunoprecipitates obtained either from HeLa cells transfected with plasmids expressing Flag–DRBP76 ( A ) or from immunopurification of Flag–YY1 ( B ) were assayed for methylase activity in the presence of core histones. “Flag competitor” corresponds to the addition of excess Flag peptide immunogen. Negative controls include immunoprecipitates from mock transfected cells and anti-Flag immunopurified materials from cells transduced with adenovirus expressing the GFP. Purified recombinant SUV39H1 was used as a positive control for histone H3 methylation. Each blot was stained with Amido black to ensure proper protein transfer.
    Anti Yy1 H414, supplied by Santa Cruz Biotechnology, used in various techniques. Bioz Stars score: 93/100, based on 4 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    77
    Millipore polyclonal anti yy1
    A purified <t>YY1</t> complex methylates histone H4. Anti-Flag immunoprecipitates obtained either from HeLa cells transfected with plasmids expressing Flag–DRBP76 ( A ) or from immunopurification of Flag–YY1 ( B ) were assayed for methylase activity in the presence of core histones. “Flag competitor” corresponds to the addition of excess Flag peptide immunogen. Negative controls include immunoprecipitates from mock transfected cells and anti-Flag immunopurified materials from cells transduced with adenovirus expressing the GFP. Purified recombinant SUV39H1 was used as a positive control for histone H3 methylation. Each blot was stained with Amido black to ensure proper protein transfer.
    Polyclonal Anti Yy1, supplied by Millipore, used in various techniques. Bioz Stars score: 77/100, based on 5 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    85
    Santa Cruz Biotechnology anti yy1 sc7341x
    A purified <t>YY1</t> complex methylates histone H4. Anti-Flag immunoprecipitates obtained either from HeLa cells transfected with plasmids expressing Flag–DRBP76 ( A ) or from immunopurification of Flag–YY1 ( B ) were assayed for methylase activity in the presence of core histones. “Flag competitor” corresponds to the addition of excess Flag peptide immunogen. Negative controls include immunoprecipitates from mock transfected cells and anti-Flag immunopurified materials from cells transduced with adenovirus expressing the GFP. Purified recombinant SUV39H1 was used as a positive control for histone H3 methylation. Each blot was stained with Amido black to ensure proper protein transfer.
    Anti Yy1 Sc7341x, supplied by Santa Cruz Biotechnology, used in various techniques. Bioz Stars score: 85/100, based on 6 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    85
    Santa Cruz Biotechnology anti yy1 sc 1703
    A purified <t>YY1</t> complex methylates histone H4. Anti-Flag immunoprecipitates obtained either from HeLa cells transfected with plasmids expressing Flag–DRBP76 ( A ) or from immunopurification of Flag–YY1 ( B ) were assayed for methylase activity in the presence of core histones. “Flag competitor” corresponds to the addition of excess Flag peptide immunogen. Negative controls include immunoprecipitates from mock transfected cells and anti-Flag immunopurified materials from cells transduced with adenovirus expressing the GFP. Purified recombinant SUV39H1 was used as a positive control for histone H3 methylation. Each blot was stained with Amido black to ensure proper protein transfer.
    Anti Yy1 Sc 1703, supplied by Santa Cruz Biotechnology, used in various techniques. Bioz Stars score: 85/100, based on 9 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    Image Search Results


    YY1 knockdown reduces viral integration into the host genome. (A) Knockdown of YY1 by siRNA was confirmed by Western blotting. (B and C) The amount of total viral cDNA (B) and two-LTR circle (C) of pQEGFP (siRNA no. 1006) and pLNΔAG (siRNA no. 1099) viral vectors is shown. NIH 3T3 cells were transfected twice with YY1 siRNA no. 1006 or no. 1099 and then infected with pQEGFP (MOI of 0.1) or pLNΔAG (MOI of 1). Cellular DNA was extracted at 0, 4, 10, 24, and 48 h and subjected to qPCR analysis. Copy number per cell was calculated by normalizing the value of the viral sequence with that of GAPDH (glyceraldehyde-3-phosphate dehydrogenase). Error bars represent the standard deviations of the results of at least three experiments. Primers used for PCR for GAPDH were 5′-TGTGATGGGTGTGAACCACGAGAA-3′ and 5′-GAGCCCTTCCACAATGCCAAAGTT-3′. Asterisks indicate that values of YY1 siRNA compared to those of scrambled RNA were significantly different (*, P = 0.038, and **, P = 0.031 in panel B; *, P = 0.022, **, P = 0.041, and ***, P = 0.047 in panel C). (D) The integrated form of viral DNA was quantified 14 days postinfection by qPCR and normalized as described for the results presented in panel C. Asterisks indicate significance of values of YY1 siRNA versus those of scrambled RNA (*, P = 0.014, **, P = 0.016, and ***, P

    Journal: Journal of Virology

    Article Title: Transcription Factor YY1 Interacts with Retroviral Integrases and Facilitates Integration of Moloney Murine Leukemia Virus cDNA into the Host Chromosomes ▿

    doi: 10.1128/JVI.02681-09

    Figure Lengend Snippet: YY1 knockdown reduces viral integration into the host genome. (A) Knockdown of YY1 by siRNA was confirmed by Western blotting. (B and C) The amount of total viral cDNA (B) and two-LTR circle (C) of pQEGFP (siRNA no. 1006) and pLNΔAG (siRNA no. 1099) viral vectors is shown. NIH 3T3 cells were transfected twice with YY1 siRNA no. 1006 or no. 1099 and then infected with pQEGFP (MOI of 0.1) or pLNΔAG (MOI of 1). Cellular DNA was extracted at 0, 4, 10, 24, and 48 h and subjected to qPCR analysis. Copy number per cell was calculated by normalizing the value of the viral sequence with that of GAPDH (glyceraldehyde-3-phosphate dehydrogenase). Error bars represent the standard deviations of the results of at least three experiments. Primers used for PCR for GAPDH were 5′-TGTGATGGGTGTGAACCACGAGAA-3′ and 5′-GAGCCCTTCCACAATGCCAAAGTT-3′. Asterisks indicate that values of YY1 siRNA compared to those of scrambled RNA were significantly different (*, P = 0.038, and **, P = 0.031 in panel B; *, P = 0.022, **, P = 0.041, and ***, P = 0.047 in panel C). (D) The integrated form of viral DNA was quantified 14 days postinfection by qPCR and normalized as described for the results presented in panel C. Asterisks indicate significance of values of YY1 siRNA versus those of scrambled RNA (*, P = 0.014, **, P = 0.016, and ***, P

    Article Snippet: The partially purified PICs were incubated with anti-MoMLV IN antiserum, anti-YY1 antibody, or control rabbit preimmune serum for 4 h, followed by absorption of the immunocomplex onto salmon sperm DNA-protein A agarose (Millipore) for 1 h. The beads were washed with buffer containing 20 mM HEPES-NaOH (pH 7.5), 5 mM MgCl2 , 150 mM KCl, 6% sucrose, 5 mM DTT, 6 mM EDTA, and 0.1% NP-40.

    Techniques: Western Blot, Transfection, Infection, Real-time Polymerase Chain Reaction, Sequencing, Polymerase Chain Reaction

    Identification of the IN-binding region in YY1 in vitro and in vivo . (A) Schematic drawing of full-length YY1 and its fragments used for pulldown and coimmunoprecipitation experiments. His, histidine-rich domain; GA, glycine-alanine-rich domain; GK, glycine-lysine-rich domain. The results of in vitro pulldown and immunoprecipitation experiments are summarized on the right side of the panel. (B) Results of pulldown assay of MoMLV IN with GST fusion proteins formed with various portions of human YY1. His-MoMLV IN was detected by Western blotting using anti-MoMLV IN antiserum. GST and GST-YY1 fragments were subjected to SDS-PAGE, and the gel was stained with Coomassie brilliant blue. Arrowheads show the intact fragments of GST-YY1 derivatives (lower panel). A result representative of several experiments is presented. (C) Results of coimmunoprecipitation of human YY1 and MoMLV IN expressed in 293FT cells (upper panels). Normal mouse IgG was used in the control, and nonspecific bands are indicated by asterisks. Cells were lysed with buffer A. IN complex was precipitated and washed with buffer A (left and central panels) or buffer B (right panel). In the right panel, controls without IN were included to estimate a nonspecific interaction between YY1 and anti-Flag antibody observed in lane 8, since interaction between IN and some YY1 derivatives seemed to be weak. His-tagged fragments of human YY1 were detected with anti-YY1 or anti-His antibody. These antibodies were also used for detection of input YY1 derivatives (2.5%). Flag-IN was detected with anti-Flag antibody. A result representative of several experiments is presented.

    Journal: Journal of Virology

    Article Title: Transcription Factor YY1 Interacts with Retroviral Integrases and Facilitates Integration of Moloney Murine Leukemia Virus cDNA into the Host Chromosomes ▿

    doi: 10.1128/JVI.02681-09

    Figure Lengend Snippet: Identification of the IN-binding region in YY1 in vitro and in vivo . (A) Schematic drawing of full-length YY1 and its fragments used for pulldown and coimmunoprecipitation experiments. His, histidine-rich domain; GA, glycine-alanine-rich domain; GK, glycine-lysine-rich domain. The results of in vitro pulldown and immunoprecipitation experiments are summarized on the right side of the panel. (B) Results of pulldown assay of MoMLV IN with GST fusion proteins formed with various portions of human YY1. His-MoMLV IN was detected by Western blotting using anti-MoMLV IN antiserum. GST and GST-YY1 fragments were subjected to SDS-PAGE, and the gel was stained with Coomassie brilliant blue. Arrowheads show the intact fragments of GST-YY1 derivatives (lower panel). A result representative of several experiments is presented. (C) Results of coimmunoprecipitation of human YY1 and MoMLV IN expressed in 293FT cells (upper panels). Normal mouse IgG was used in the control, and nonspecific bands are indicated by asterisks. Cells were lysed with buffer A. IN complex was precipitated and washed with buffer A (left and central panels) or buffer B (right panel). In the right panel, controls without IN were included to estimate a nonspecific interaction between YY1 and anti-Flag antibody observed in lane 8, since interaction between IN and some YY1 derivatives seemed to be weak. His-tagged fragments of human YY1 were detected with anti-YY1 or anti-His antibody. These antibodies were also used for detection of input YY1 derivatives (2.5%). Flag-IN was detected with anti-Flag antibody. A result representative of several experiments is presented.

    Article Snippet: The partially purified PICs were incubated with anti-MoMLV IN antiserum, anti-YY1 antibody, or control rabbit preimmune serum for 4 h, followed by absorption of the immunocomplex onto salmon sperm DNA-protein A agarose (Millipore) for 1 h. The beads were washed with buffer containing 20 mM HEPES-NaOH (pH 7.5), 5 mM MgCl2 , 150 mM KCl, 6% sucrose, 5 mM DTT, 6 mM EDTA, and 0.1% NP-40.

    Techniques: Binding Assay, In Vitro, In Vivo, Immunoprecipitation, Western Blot, SDS Page, Staining

    YY1 identify a dominant phenotypic clone in ERα BC A) RIs for the YY1 enhancer within all the individual patients included in the current study YY1 enhancer location with its 3D interactions are shown in the top right inset B) YY1 enhancer ranking analysis of available Epigenome Roadmap H3K27ac datasets. Tissues are displayed from the strongest to the weakest YY1 enhancer activity (based on RI). Representative IHC analysis of normal tissues stained with a YY1 antibody are shown C) eKO cell lines were generated by deleting a 2.4kb containing YY1-A enhancers in MCF7 cells. Actual karyotyping is shown in the bottom panel was performed on 10 individual cells D) YY1 expression in control and eKO cell lines was measured using RT-qPCR. Lines and error bars represent average and 95% CI of five independent experiments. Significance was calculated with a one-way ANOVA followed by Tukey’s test E) Top left: YY1 expression in ERα-positive breast cancer compared to normal breast tissue. Median, lowest and highest values are reported. Top right: YY1 prognostic value in triple negative breast cancers. Bottom left: YY1 prognostic value in luminal breast cancers. Confidence interval (1.19-1.76). Bottom right: multivariate correction for the luminal breast cancer dataset is shown. Analyses included 1476 ERα-positive and 432 ERα-negative patients. Comparison of survival curves was performed using a Log-rank (Mantel-Cox) test. F) IHC analysis of normal breast tissues highlights YY1 functional subclones in normal breast. Similar results were observed in 10 independent clinical specimens from independent individuals G) IHC analysis of ERα positive invasive ductal carcinomas identify YY1 positive clones as the dominant clonal population (Scale bars, 50 μm).

    Journal: Nature medicine

    Article Title: Enhancer mapping uncovers phenotypic heterogeneity and evolution in patients with luminal breast cancer

    doi: 10.1038/s41591-018-0091-x

    Figure Lengend Snippet: YY1 identify a dominant phenotypic clone in ERα BC A) RIs for the YY1 enhancer within all the individual patients included in the current study YY1 enhancer location with its 3D interactions are shown in the top right inset B) YY1 enhancer ranking analysis of available Epigenome Roadmap H3K27ac datasets. Tissues are displayed from the strongest to the weakest YY1 enhancer activity (based on RI). Representative IHC analysis of normal tissues stained with a YY1 antibody are shown C) eKO cell lines were generated by deleting a 2.4kb containing YY1-A enhancers in MCF7 cells. Actual karyotyping is shown in the bottom panel was performed on 10 individual cells D) YY1 expression in control and eKO cell lines was measured using RT-qPCR. Lines and error bars represent average and 95% CI of five independent experiments. Significance was calculated with a one-way ANOVA followed by Tukey’s test E) Top left: YY1 expression in ERα-positive breast cancer compared to normal breast tissue. Median, lowest and highest values are reported. Top right: YY1 prognostic value in triple negative breast cancers. Bottom left: YY1 prognostic value in luminal breast cancers. Confidence interval (1.19-1.76). Bottom right: multivariate correction for the luminal breast cancer dataset is shown. Analyses included 1476 ERα-positive and 432 ERα-negative patients. Comparison of survival curves was performed using a Log-rank (Mantel-Cox) test. F) IHC analysis of normal breast tissues highlights YY1 functional subclones in normal breast. Similar results were observed in 10 independent clinical specimens from independent individuals G) IHC analysis of ERα positive invasive ductal carcinomas identify YY1 positive clones as the dominant clonal population (Scale bars, 50 μm).

    Article Snippet: For YY1 (Protein Atlas HPA001119, Atlas Antibodies Cat#HPA001119, RRID:AB_1858930 ) the flowing conditions were used: tissue sections were incubated with the primary monoclonal. overnight at 4°C, and chromogen development was performed using the Envision system (DAKO Corporation, Glostrup, Denmark).

    Techniques: Activity Assay, Immunohistochemistry, Staining, Generated, Expressing, Quantitative RT-PCR, Functional Assay, Clone Assay

    YY1 marks critical enhancers in breast cancer cells A) ChIP-seq data from ERα-positive MCF7 for YY1 in quiescent or 17ß -estradiol (E2) stimulated cells B) Heatmaps showing global enrichment profiles of several chromatin markers associated with active regulatory regions in MCF7 cells C) Overlap between ERα, YY1 and FOXA1 in MCF7 cells. The right panel shows the potential overlap with in vivo- derived core ERα binding sites D) ERα core binding sites are strongly enriched for YY1 binding in MCF7 cells while patient-specific ERα bindings are generally YY1-free. Proportion were compared using Fisher’s Exact test. E) Genes used to classify luminal breast cancer patients are strongly enriched for ERα-YY1 binding sites. Asterisks represent p

    Journal: Nature medicine

    Article Title: Enhancer mapping uncovers phenotypic heterogeneity and evolution in patients with luminal breast cancer

    doi: 10.1038/s41591-018-0091-x

    Figure Lengend Snippet: YY1 marks critical enhancers in breast cancer cells A) ChIP-seq data from ERα-positive MCF7 for YY1 in quiescent or 17ß -estradiol (E2) stimulated cells B) Heatmaps showing global enrichment profiles of several chromatin markers associated with active regulatory regions in MCF7 cells C) Overlap between ERα, YY1 and FOXA1 in MCF7 cells. The right panel shows the potential overlap with in vivo- derived core ERα binding sites D) ERα core binding sites are strongly enriched for YY1 binding in MCF7 cells while patient-specific ERα bindings are generally YY1-free. Proportion were compared using Fisher’s Exact test. E) Genes used to classify luminal breast cancer patients are strongly enriched for ERα-YY1 binding sites. Asterisks represent p

    Article Snippet: For YY1 (Protein Atlas HPA001119, Atlas Antibodies Cat#HPA001119, RRID:AB_1858930 ) the flowing conditions were used: tissue sections were incubated with the primary monoclonal. overnight at 4°C, and chromogen development was performed using the Envision system (DAKO Corporation, Glostrup, Denmark).

    Techniques: Chromatin Immunoprecipitation, In Vivo, Derivative Assay, Binding Assay

    Endocrine treatment shapes phenotypic evolution. A) Theoretical framework of the analysis. The relative size of phenotypic clones can be tracked using enhancer ranking (RIs). Phenotypic clones can be positively or negatively selected during BC progression in response to endocrine therapies. B) Expanding or contracting phenotypic clones were defined based on the RI-ratio in primary and metastatic samples (RI P /RI M ). Distribution of RI-ratio shows that YY1 enhancers RI does not change significantly during progression compared to other enhancers, while SLC9A3R1 RI ranks among the enhancers with stronger increase in activity during progression. Vertical bars represent 1σ (Standard Deviation) increments from the population median C) Scatterplot of YY1 and SLC9A3R1 enhancer ranking according to patient stage. Bars indicate mean and 95% confidence intervals. Asterisks represent significance at P

    Journal: Nature medicine

    Article Title: Enhancer mapping uncovers phenotypic heterogeneity and evolution in patients with luminal breast cancer

    doi: 10.1038/s41591-018-0091-x

    Figure Lengend Snippet: Endocrine treatment shapes phenotypic evolution. A) Theoretical framework of the analysis. The relative size of phenotypic clones can be tracked using enhancer ranking (RIs). Phenotypic clones can be positively or negatively selected during BC progression in response to endocrine therapies. B) Expanding or contracting phenotypic clones were defined based on the RI-ratio in primary and metastatic samples (RI P /RI M ). Distribution of RI-ratio shows that YY1 enhancers RI does not change significantly during progression compared to other enhancers, while SLC9A3R1 RI ranks among the enhancers with stronger increase in activity during progression. Vertical bars represent 1σ (Standard Deviation) increments from the population median C) Scatterplot of YY1 and SLC9A3R1 enhancer ranking according to patient stage. Bars indicate mean and 95% confidence intervals. Asterisks represent significance at P

    Article Snippet: For YY1 (Protein Atlas HPA001119, Atlas Antibodies Cat#HPA001119, RRID:AB_1858930 ) the flowing conditions were used: tissue sections were incubated with the primary monoclonal. overnight at 4°C, and chromogen development was performed using the Envision system (DAKO Corporation, Glostrup, Denmark).

    Techniques: Clone Assay, Activity Assay, Standard Deviation

    Expanded polyQ perturbs the function of YY1 to derepress Fuz expression A Knockdown of YY1 expression upregulated Fuz expression in HEK293 cells. Lower panel shows the quantification of Fuz transcript expression relative to controls. Error bars represent s.e.m., n = 3. Statistical analysis was performed using two‐tailed unpaired Student's t ‐test. * P

    Journal: EMBO Reports

    Article Title: Planar cell polarity gene Fuz triggers apoptosis in neurodegenerative disease models

    doi: 10.15252/embr.201745409

    Figure Lengend Snippet: Expanded polyQ perturbs the function of YY1 to derepress Fuz expression A Knockdown of YY1 expression upregulated Fuz expression in HEK293 cells. Lower panel shows the quantification of Fuz transcript expression relative to controls. Error bars represent s.e.m., n = 3. Statistical analysis was performed using two‐tailed unpaired Student's t ‐test. * P

    Article Snippet: Primary antibodies used were anti‐cleaved caspase‐3 (9664, 1:500), anti‐p‐JNK (9251, 1:500), anti‐JNK (9252, 1:1,000), anti‐myc (2276, 1:2,000) from Cell Signaling Technology; anti‐p‐MEKK1 (ab138662, 1:1,000), anti‐MEKK1 (ab69533, 1:1,000), anti‐Fuz (ab111842, 1:500), anti‐WT1 (ab89901, 1:1,000), anti‐Flamingo (ab90817, 1:500), anti‐YY1 (ab109237, 1:1,000), anti‐beta‐tubulin (ab6046, 1:2,000) from Abcam; anti‐Tiam1 (sc‐393315, 1:100), anti‐dishevelled (sc‐8027, 1:200), anti‐NR3C1 (sc‐56851, 1:500) from Santa Cruz Biotechnology; anti‐Inturned (LS‐C169884‐50, 1:500) from LifeSpan BioSciences; anti‐Fritz (PA524271, 1:500) from Thermo Fisher Scientific; anti‐ATXN3 (MAB5360, 1:500) from Merck Millipore; anti‐polyglutamine (P1874, 1:1,000), anti‐flag (F3165, 1:500), anti‐HA (H3663, 1:500) and anti‐His (27‐4710‐01, 1:1,000) from Sigma‐Aldrich; anti‐Hsp70 (SPA‐812C, 1:1,000) from Enzo Life Sciences.

    Techniques: Expressing, Two Tailed Test

    YY1 interacts with Fuz promoter in vivo and in vitro A Schematic representation of the human Fuz −1332/+574 promoter region. The Fuz +68/+574 was enlarged to show the detailed nucleotide sequence. A putative CpG island was found within Fuz +68/+574 and was defined as Fuz +117/+347CpG . “+1” is the transcriptional initiation site (arrow). The CpG island is highlighted in blue, and sequence of the YY1 binding site is underlined and highlighted in red. B Chromatin immunoprecipitation assay demonstrated the binding between YY1 protein and Fuz +117/+347CpG DNA fragment. The blue bar indicates the Fuz +117/+347CpG sequence. However, Fuz −802/−612 (orange) represents a region that did not show interaction with YY1 protein. n = 3. C, D (C) In vitro DNA binding assay demonstrated the binding between purified YY1 protein and purified Fuz +117/+347CpG DNA fragment. When a mutation (green) was introduced to the YY1 site (red) within the Fuz +117/+347CpG fragment, YY1 binding was reduced. (D) is the quantification of the intensity of the Fuz +117/+347CpG DNA bands as shown in (C). Error bars represent s.e.m., n = 3. Statistical analysis was performed using two‐tailed unpaired Student's t ‐test. * P

    Journal: EMBO Reports

    Article Title: Planar cell polarity gene Fuz triggers apoptosis in neurodegenerative disease models

    doi: 10.15252/embr.201745409

    Figure Lengend Snippet: YY1 interacts with Fuz promoter in vivo and in vitro A Schematic representation of the human Fuz −1332/+574 promoter region. The Fuz +68/+574 was enlarged to show the detailed nucleotide sequence. A putative CpG island was found within Fuz +68/+574 and was defined as Fuz +117/+347CpG . “+1” is the transcriptional initiation site (arrow). The CpG island is highlighted in blue, and sequence of the YY1 binding site is underlined and highlighted in red. B Chromatin immunoprecipitation assay demonstrated the binding between YY1 protein and Fuz +117/+347CpG DNA fragment. The blue bar indicates the Fuz +117/+347CpG sequence. However, Fuz −802/−612 (orange) represents a region that did not show interaction with YY1 protein. n = 3. C, D (C) In vitro DNA binding assay demonstrated the binding between purified YY1 protein and purified Fuz +117/+347CpG DNA fragment. When a mutation (green) was introduced to the YY1 site (red) within the Fuz +117/+347CpG fragment, YY1 binding was reduced. (D) is the quantification of the intensity of the Fuz +117/+347CpG DNA bands as shown in (C). Error bars represent s.e.m., n = 3. Statistical analysis was performed using two‐tailed unpaired Student's t ‐test. * P

    Article Snippet: Primary antibodies used were anti‐cleaved caspase‐3 (9664, 1:500), anti‐p‐JNK (9251, 1:500), anti‐JNK (9252, 1:1,000), anti‐myc (2276, 1:2,000) from Cell Signaling Technology; anti‐p‐MEKK1 (ab138662, 1:1,000), anti‐MEKK1 (ab69533, 1:1,000), anti‐Fuz (ab111842, 1:500), anti‐WT1 (ab89901, 1:1,000), anti‐Flamingo (ab90817, 1:500), anti‐YY1 (ab109237, 1:1,000), anti‐beta‐tubulin (ab6046, 1:2,000) from Abcam; anti‐Tiam1 (sc‐393315, 1:100), anti‐dishevelled (sc‐8027, 1:200), anti‐NR3C1 (sc‐56851, 1:500) from Santa Cruz Biotechnology; anti‐Inturned (LS‐C169884‐50, 1:500) from LifeSpan BioSciences; anti‐Fritz (PA524271, 1:500) from Thermo Fisher Scientific; anti‐ATXN3 (MAB5360, 1:500) from Merck Millipore; anti‐polyglutamine (P1874, 1:1,000), anti‐flag (F3165, 1:500), anti‐HA (H3663, 1:500) and anti‐His (27‐4710‐01, 1:1,000) from Sigma‐Aldrich; anti‐Hsp70 (SPA‐812C, 1:1,000) from Enzo Life Sciences.

    Techniques: In Vivo, In Vitro, Sequencing, Binding Assay, Chromatin Immunoprecipitation, DNA Binding Assay, Purification, Mutagenesis, Two Tailed Test

    Soluble YY1 protein level is reduced in polyQ diseases A, B . (B) is the quantification of (A). Error bars represent s.e.m., n = 6. Statistical analysis was performed using two‐tailed unpaired Student's t ‐test. ** P

    Journal: EMBO Reports

    Article Title: Planar cell polarity gene Fuz triggers apoptosis in neurodegenerative disease models

    doi: 10.15252/embr.201745409

    Figure Lengend Snippet: Soluble YY1 protein level is reduced in polyQ diseases A, B . (B) is the quantification of (A). Error bars represent s.e.m., n = 6. Statistical analysis was performed using two‐tailed unpaired Student's t ‐test. ** P

    Article Snippet: Primary antibodies used were anti‐cleaved caspase‐3 (9664, 1:500), anti‐p‐JNK (9251, 1:500), anti‐JNK (9252, 1:1,000), anti‐myc (2276, 1:2,000) from Cell Signaling Technology; anti‐p‐MEKK1 (ab138662, 1:1,000), anti‐MEKK1 (ab69533, 1:1,000), anti‐Fuz (ab111842, 1:500), anti‐WT1 (ab89901, 1:1,000), anti‐Flamingo (ab90817, 1:500), anti‐YY1 (ab109237, 1:1,000), anti‐beta‐tubulin (ab6046, 1:2,000) from Abcam; anti‐Tiam1 (sc‐393315, 1:100), anti‐dishevelled (sc‐8027, 1:200), anti‐NR3C1 (sc‐56851, 1:500) from Santa Cruz Biotechnology; anti‐Inturned (LS‐C169884‐50, 1:500) from LifeSpan BioSciences; anti‐Fritz (PA524271, 1:500) from Thermo Fisher Scientific; anti‐ATXN3 (MAB5360, 1:500) from Merck Millipore; anti‐polyglutamine (P1874, 1:1,000), anti‐flag (F3165, 1:500), anti‐HA (H3663, 1:500) and anti‐His (27‐4710‐01, 1:1,000) from Sigma‐Aldrich; anti‐Hsp70 (SPA‐812C, 1:1,000) from Enzo Life Sciences.

    Techniques: Two Tailed Test

    YY1 promotes trophoblast outgrowth in extravillous explant cultures. (A) Extravillous explants were obtained from HCs and RM patients at 6–8 weeks of gestation and cultured on Matrigel. (B) Statistical assay of the migration distance of villous tips (%). Data are presented as means ± SD of three independent experiments. (C) The extravillous explants were cultured on Matrigel for 72 h. Immunofluorescence staining using anti‐YY1 antibodies showed an obvious decrease in the YY1 protein level in the RM group compared with the HC group. Green fluorescence signals indicate bound anti‐YY1 antibodies; CK7 staining is visualized as red; and the DAPI‐stained nuclei are blue. (D) The number of YY1 + cells and CK7 + cells was calculated, respectively, using Leica confocal SP8 software and then the percentage of CK7 + cells normalized to the number of YY1 + invading trophoblasts in RM and HC samples was assessed. (E) Extravillous explants were maintained in culture on Matrigel. Serial pictures of the explants incubated with siYY1 or siCtrl were taken under a light microscope after 24 and 72 h of culture in vitro . (F) Extravillous explants from RM patients were maintained in culture on Matrigel. Serial pictures of the explants incubated with lenti‐ctrl or lenti‐YY1 lentivirus were taken under a light microscope after 24 and 72 h of culture in vitro .

    Journal: The Journal of Pathology

    Article Title: The YY1/ MMP2 axis promotes trophoblast invasion at the maternal–fetal interface

    doi: 10.1002/path.4694

    Figure Lengend Snippet: YY1 promotes trophoblast outgrowth in extravillous explant cultures. (A) Extravillous explants were obtained from HCs and RM patients at 6–8 weeks of gestation and cultured on Matrigel. (B) Statistical assay of the migration distance of villous tips (%). Data are presented as means ± SD of three independent experiments. (C) The extravillous explants were cultured on Matrigel for 72 h. Immunofluorescence staining using anti‐YY1 antibodies showed an obvious decrease in the YY1 protein level in the RM group compared with the HC group. Green fluorescence signals indicate bound anti‐YY1 antibodies; CK7 staining is visualized as red; and the DAPI‐stained nuclei are blue. (D) The number of YY1 + cells and CK7 + cells was calculated, respectively, using Leica confocal SP8 software and then the percentage of CK7 + cells normalized to the number of YY1 + invading trophoblasts in RM and HC samples was assessed. (E) Extravillous explants were maintained in culture on Matrigel. Serial pictures of the explants incubated with siYY1 or siCtrl were taken under a light microscope after 24 and 72 h of culture in vitro . (F) Extravillous explants from RM patients were maintained in culture on Matrigel. Serial pictures of the explants incubated with lenti‐ctrl or lenti‐YY1 lentivirus were taken under a light microscope after 24 and 72 h of culture in vitro .

    Article Snippet: Human villous tissues were labelled with rabbit anti‐YY1 antibodies (ab109237, dilution 1:500; Abcam).

    Techniques: Cell Culture, Migration, Immunofluorescence, Staining, Fluorescence, Software, Incubation, Light Microscopy, In Vitro

    YY1 represses MMP2 expression in trophoblasts. (A) The diagram illustrates the YY1‐binding site in the promoter region of MMP2. (B) Primary trophoblasts (5 × 10 6 ) were isolated from HCs at 6–8 weeks of gestation and a ChIP assay was performed using 2 µg of YY1‐specific antibody to immunoprecipitate the transcriptionally active regions of DNA. The purified DNA was amplified by RT‐qPCR using primer sets specific to UP1–UP4 of MMP2. (C–F) Western blotting and RT‐qPCR analyses of MMP2 expression in HTR‐8 cells transfected as indicated with siCtrl, siYY1, and control vector or YY1 expression vector. (G) Representative immunofluorescence images of YY1 in frozen first‐trimester villous sections obtained at 6–8 weeks of gestation. Fluorescence signals specific to anti‐YY1 antibodies appear green; anti‐MMP2 antibodies appear red; and the DAPI‐stained nuclei appear blue. (H, I) The YY1 and MMP2 mRNA expression levels in the villous tissue of patients with one ( n = 13) or two ( n = 15) pregnancy losses, RM ( n = 31), and HCs ( n = 36) were determined by RT‐qPCR. (J) The MMP2 mRNA expression level measured in the villous tissue of patients ( n = 49) was measured using RT‐qPCR and correlated with the YY1 mRNA expression level in the villous tissue in corresponding patients ( n = 49 ) .

    Journal: The Journal of Pathology

    Article Title: The YY1/ MMP2 axis promotes trophoblast invasion at the maternal–fetal interface

    doi: 10.1002/path.4694

    Figure Lengend Snippet: YY1 represses MMP2 expression in trophoblasts. (A) The diagram illustrates the YY1‐binding site in the promoter region of MMP2. (B) Primary trophoblasts (5 × 10 6 ) were isolated from HCs at 6–8 weeks of gestation and a ChIP assay was performed using 2 µg of YY1‐specific antibody to immunoprecipitate the transcriptionally active regions of DNA. The purified DNA was amplified by RT‐qPCR using primer sets specific to UP1–UP4 of MMP2. (C–F) Western blotting and RT‐qPCR analyses of MMP2 expression in HTR‐8 cells transfected as indicated with siCtrl, siYY1, and control vector or YY1 expression vector. (G) Representative immunofluorescence images of YY1 in frozen first‐trimester villous sections obtained at 6–8 weeks of gestation. Fluorescence signals specific to anti‐YY1 antibodies appear green; anti‐MMP2 antibodies appear red; and the DAPI‐stained nuclei appear blue. (H, I) The YY1 and MMP2 mRNA expression levels in the villous tissue of patients with one ( n = 13) or two ( n = 15) pregnancy losses, RM ( n = 31), and HCs ( n = 36) were determined by RT‐qPCR. (J) The MMP2 mRNA expression level measured in the villous tissue of patients ( n = 49) was measured using RT‐qPCR and correlated with the YY1 mRNA expression level in the villous tissue in corresponding patients ( n = 49 ) .

    Article Snippet: Human villous tissues were labelled with rabbit anti‐YY1 antibodies (ab109237, dilution 1:500; Abcam).

    Techniques: Expressing, Binding Assay, Isolation, Chromatin Immunoprecipitation, Purification, Amplification, Quantitative RT-PCR, Western Blot, Transfection, Plasmid Preparation, Immunofluorescence, Fluorescence, Staining

    YY1 is down‐regulated in cytotrophoblasts and extravillous trophoblasts in RM patients. (A, B) The YY1 levels in first‐trimester villous tissues and third‐trimester placental tissues from the RM and HC groups were determined by RT‐qPCR and western blotting ( n = 6). (C, D) Single staining of maternal villi [cytotrophoblasts (CTBs) and syncytiotrophoblasts (STBs)] using rabbit IgG anti‐human YY1 antibodies and developed with a labelled streptavidin biotin + horseradish peroxidase (HRP) kit. The sections were counterstained with haematoxylin and positive cells were quantified using ImagePro‐plus 6.0. Panel: original magnification × 40; n = 12. (E, F) Representative immunofluorescence images of YY1 in frozen first‐trimester villous sections (6–8 weeks of gestation). Green indicates fluorescence signals specific to anti‐YY1 antibodies and blue indicates nuclei. Fluorescence intensity of the signalling was assessed by Leica confocal SP8 software. Large panel: original magnification × 100; small panel: original magnification × 200; n = 15. (G) Representative immunofluorescence images of YY1 in frozen first‐trimester decidual tissue sections (6–8 weeks of gestation). Fluorescence signals specific to anti‐YY1 antibodies appear green; to the CK7 staining, red; and to the DAPI‐stained nuclei, blue. Panel: original magnification × 100; n = 15. (H) The number of YY1 + cells and CK7 + cells was calculated, respectively, using Leica confocal SP8 software and then the percentage of CK7 + cells normalized to the number of YY1 + cells in the decidual tissue of RM and HC samples was assessed.

    Journal: The Journal of Pathology

    Article Title: The YY1/ MMP2 axis promotes trophoblast invasion at the maternal–fetal interface

    doi: 10.1002/path.4694

    Figure Lengend Snippet: YY1 is down‐regulated in cytotrophoblasts and extravillous trophoblasts in RM patients. (A, B) The YY1 levels in first‐trimester villous tissues and third‐trimester placental tissues from the RM and HC groups were determined by RT‐qPCR and western blotting ( n = 6). (C, D) Single staining of maternal villi [cytotrophoblasts (CTBs) and syncytiotrophoblasts (STBs)] using rabbit IgG anti‐human YY1 antibodies and developed with a labelled streptavidin biotin + horseradish peroxidase (HRP) kit. The sections were counterstained with haematoxylin and positive cells were quantified using ImagePro‐plus 6.0. Panel: original magnification × 40; n = 12. (E, F) Representative immunofluorescence images of YY1 in frozen first‐trimester villous sections (6–8 weeks of gestation). Green indicates fluorescence signals specific to anti‐YY1 antibodies and blue indicates nuclei. Fluorescence intensity of the signalling was assessed by Leica confocal SP8 software. Large panel: original magnification × 100; small panel: original magnification × 200; n = 15. (G) Representative immunofluorescence images of YY1 in frozen first‐trimester decidual tissue sections (6–8 weeks of gestation). Fluorescence signals specific to anti‐YY1 antibodies appear green; to the CK7 staining, red; and to the DAPI‐stained nuclei, blue. Panel: original magnification × 100; n = 15. (H) The number of YY1 + cells and CK7 + cells was calculated, respectively, using Leica confocal SP8 software and then the percentage of CK7 + cells normalized to the number of YY1 + cells in the decidual tissue of RM and HC samples was assessed.

    Article Snippet: Human villous tissues were labelled with rabbit anti‐YY1 antibodies (ab109237, dilution 1:500; Abcam).

    Techniques: Quantitative RT-PCR, Western Blot, Staining, Immunofluorescence, Fluorescence, Software

    Depletion of YY1 Disrupts Gene Expression (A) Model depicting dTAG system used to rapidly deplete YY1 protein. (B) Western blot validation of knockin of FKBP degron tag and ability to inducibly degrade YY1 protein. (C) Change in gene expression (log 2 fold change) upon degradation of YY1 for all genes plotted against the expression in untreated cells. Genes that displayed significant changes in expression (false discovery rate [FDR] adjusted p value

    Journal: Cell

    Article Title: YY1 Is a Structural Regulator of Enhancer-Promoter Loops

    doi: 10.1016/j.cell.2017.11.008

    Figure Lengend Snippet: Depletion of YY1 Disrupts Gene Expression (A) Model depicting dTAG system used to rapidly deplete YY1 protein. (B) Western blot validation of knockin of FKBP degron tag and ability to inducibly degrade YY1 protein. (C) Change in gene expression (log 2 fold change) upon degradation of YY1 for all genes plotted against the expression in untreated cells. Genes that displayed significant changes in expression (false discovery rate [FDR] adjusted p value

    Article Snippet: 7.5 μg of H3K27ac antibody (Abcam, ab4729) or 7.5 μg of YY1 antibody (Abcam, ab109237) was added to the tube and the tube was incubated overnight at 4°C with rotation.

    Techniques: Expressing, Western Blot, Knock-In

    Rescue of Enhancer-Promoter Interactions in Cells (A) Model depicting use of dCas9-YY1 to artificially tether YY1 to a site adjacent to the YY1 binding site mutation in the promoter-proximal region of Etv4 in order to determine whether artificially tethered YY1 can rescue enhancer-promoter interactions. (B) Model depicting dCas9-YY1 rescue experiments. Etv4 promoter-proximal YY1 binding motif mutant cells were transduced with lentivirus to stably express either dCas9 or dCas9-YY1, and two sgRNAs to direct their localization to the sequences adjacent to the deleted YY1 binding motif in the Etv4 promoter-proximal region. The ability to rescue enhancer-promoter looping was assayed by 4C-seq. (C) Western blot results showing that Etv4 promoter-proximal YY1 binding motif mutant cells transduced with lentivirus to stably express either dCas9 or dCas9-YY1 successfully express dCas9 or dCas9-YY1. (D) Artificial tethering of YY1 using dCas9-YY1 was performed at sites adjacent to the YY1 binding site mutation in the promoter-proximal region of Etv4 . The effects of tethering YY1 using dCas9-YY1 on enhancer-promoter looping and expression of the Etv4 gene were measured and compared to dCas9 alone. The genotype of the Etv4 promoter-proximal YY1 binding motif mutant cells and the 4C-seq viewpoint (VP) is shown. The 4C-seq signal is displayed as the smoothed average reads per million per base pair. The mean 4C-seq signal is represented as a line, and the shaded area represents the 95% confidence interval. Three biological replicates were assayed for 4C-seq and CAS9 ChIP-qPCR experiments, and six biological replicates were assayed for RT-qPCR experiments. Error bars represent the SD. All p values were determined using the Student’s t test. (E) Model depicting the loss of looping interactions after the inducible degradation of the structuring factors CTCF and YY1 followed by restoration of looping upon washout of degradation compounds. (F) Change in normalized interaction frequency (log2 fold change) after YY1 and CTCF degradation (treated) and recovery (washout) relative to untreated cells. For YY1 degradation, change in normalized interaction frequency is plotted for YY1-YY1 enhancer-promoter interactions. For CTCF degradation, change in normalized interaction frequency is plotted for CTCF-CTCF interactions. .

    Journal: Cell

    Article Title: YY1 Is a Structural Regulator of Enhancer-Promoter Loops

    doi: 10.1016/j.cell.2017.11.008

    Figure Lengend Snippet: Rescue of Enhancer-Promoter Interactions in Cells (A) Model depicting use of dCas9-YY1 to artificially tether YY1 to a site adjacent to the YY1 binding site mutation in the promoter-proximal region of Etv4 in order to determine whether artificially tethered YY1 can rescue enhancer-promoter interactions. (B) Model depicting dCas9-YY1 rescue experiments. Etv4 promoter-proximal YY1 binding motif mutant cells were transduced with lentivirus to stably express either dCas9 or dCas9-YY1, and two sgRNAs to direct their localization to the sequences adjacent to the deleted YY1 binding motif in the Etv4 promoter-proximal region. The ability to rescue enhancer-promoter looping was assayed by 4C-seq. (C) Western blot results showing that Etv4 promoter-proximal YY1 binding motif mutant cells transduced with lentivirus to stably express either dCas9 or dCas9-YY1 successfully express dCas9 or dCas9-YY1. (D) Artificial tethering of YY1 using dCas9-YY1 was performed at sites adjacent to the YY1 binding site mutation in the promoter-proximal region of Etv4 . The effects of tethering YY1 using dCas9-YY1 on enhancer-promoter looping and expression of the Etv4 gene were measured and compared to dCas9 alone. The genotype of the Etv4 promoter-proximal YY1 binding motif mutant cells and the 4C-seq viewpoint (VP) is shown. The 4C-seq signal is displayed as the smoothed average reads per million per base pair. The mean 4C-seq signal is represented as a line, and the shaded area represents the 95% confidence interval. Three biological replicates were assayed for 4C-seq and CAS9 ChIP-qPCR experiments, and six biological replicates were assayed for RT-qPCR experiments. Error bars represent the SD. All p values were determined using the Student’s t test. (E) Model depicting the loss of looping interactions after the inducible degradation of the structuring factors CTCF and YY1 followed by restoration of looping upon washout of degradation compounds. (F) Change in normalized interaction frequency (log2 fold change) after YY1 and CTCF degradation (treated) and recovery (washout) relative to untreated cells. For YY1 degradation, change in normalized interaction frequency is plotted for YY1-YY1 enhancer-promoter interactions. For CTCF degradation, change in normalized interaction frequency is plotted for CTCF-CTCF interactions. .

    Article Snippet: 7.5 μg of H3K27ac antibody (Abcam, ab4729) or 7.5 μg of YY1 antibody (Abcam, ab109237) was added to the tube and the tube was incubated overnight at 4°C with rotation.

    Techniques: Binding Assay, Mutagenesis, Transduction, Stable Transfection, Western Blot, Expressing, Chromatin Immunoprecipitation, Real-time Polymerase Chain Reaction, Quantitative RT-PCR

    YY1 Generally Occupies Enhancers and Promoters in Mammalian Cells (A and B) Heatmaps displaying the YY1 occupancy at enhancers (A) and active promoters (B) in six human cell types. (C–E) Summaries of the major classes of high-confidence interactions identified with YY1 HiChIP in colorectal cancer cells (C), T cell acute lymphoblastic leukemia cells (D), and chronic myeloid leukemia cells (E). (F–K) Examples of YY1-YY1 enhancer-promoter interactions in three human cell types: colorectal cancer (F and I), T cell acute lymphoblastic leukemia (G and J), and chronic myeloid leukemia (H and K). Displayed examples show YY1-YY1 enhancer-promoter interactions involving typical enhancers (F–H) and involving super-enhancers (I–K). .

    Journal: Cell

    Article Title: YY1 Is a Structural Regulator of Enhancer-Promoter Loops

    doi: 10.1016/j.cell.2017.11.008

    Figure Lengend Snippet: YY1 Generally Occupies Enhancers and Promoters in Mammalian Cells (A and B) Heatmaps displaying the YY1 occupancy at enhancers (A) and active promoters (B) in six human cell types. (C–E) Summaries of the major classes of high-confidence interactions identified with YY1 HiChIP in colorectal cancer cells (C), T cell acute lymphoblastic leukemia cells (D), and chronic myeloid leukemia cells (E). (F–K) Examples of YY1-YY1 enhancer-promoter interactions in three human cell types: colorectal cancer (F and I), T cell acute lymphoblastic leukemia (G and J), and chronic myeloid leukemia (H and K). Displayed examples show YY1-YY1 enhancer-promoter interactions involving typical enhancers (F–H) and involving super-enhancers (I–K). .

    Article Snippet: 7.5 μg of H3K27ac antibody (Abcam, ab4729) or 7.5 μg of YY1 antibody (Abcam, ab109237) was added to the tube and the tube was incubated overnight at 4°C with rotation.

    Techniques: HiChIP

    YY1 Can Enhance DNA Interactions In Vitro (A and D) Models depicting the in vitro DNA circularization assays used to detect the ability of YY1 to enhance DNA looping interactions with no motif control (A) or competitor DNA control (D). (B and E) Results of the in vitro DNA circularization assay visualized by gel electrophoresis with no motif control (B) or competitor DNA control (E). The dominant lower band reflects the starting linear DNA template, while the upper band corresponds to the circularized DNA ligation product. (C and F) Quantifications of DNA template circularization as a function of incubation time with T4 DNA ligase for no motif control (C) or competitor DNA control (F). Values correspond to the percent of DNA template that is circularized and represents the mean and SD of four experiments. .

    Journal: Cell

    Article Title: YY1 Is a Structural Regulator of Enhancer-Promoter Loops

    doi: 10.1016/j.cell.2017.11.008

    Figure Lengend Snippet: YY1 Can Enhance DNA Interactions In Vitro (A and D) Models depicting the in vitro DNA circularization assays used to detect the ability of YY1 to enhance DNA looping interactions with no motif control (A) or competitor DNA control (D). (B and E) Results of the in vitro DNA circularization assay visualized by gel electrophoresis with no motif control (B) or competitor DNA control (E). The dominant lower band reflects the starting linear DNA template, while the upper band corresponds to the circularized DNA ligation product. (C and F) Quantifications of DNA template circularization as a function of incubation time with T4 DNA ligase for no motif control (C) or competitor DNA control (F). Values correspond to the percent of DNA template that is circularized and represents the mean and SD of four experiments. .

    Article Snippet: 7.5 μg of H3K27ac antibody (Abcam, ab4729) or 7.5 μg of YY1 antibody (Abcam, ab109237) was added to the tube and the tube was incubated overnight at 4°C with rotation.

    Techniques: In Vitro, Nucleic Acid Electrophoresis, DNA Ligation, Incubation

    Deletion of YY1 Binding Sites Causes Loss of Enhancer-Promoter Interactions (A) Model depicting CRISPR/Cas9-mediated deletion of a YY1 binding motif in the regulatory region of a gene. (B and C) CRISPR/Cas9-mediated deletion of YY1 binding motifs in the regulatory regions of two genes, Raf1 (B) and Etv4 ), and the shaded area represents the 95% confidence interval. Three biological replicates were assayed for 4C-seq and ChIP-qPCR experiments, and six biological replicates were assayed for RT-qPCR experiments. Error bars represent the SD. All p values were determined using the Student’s t test. .

    Journal: Cell

    Article Title: YY1 Is a Structural Regulator of Enhancer-Promoter Loops

    doi: 10.1016/j.cell.2017.11.008

    Figure Lengend Snippet: Deletion of YY1 Binding Sites Causes Loss of Enhancer-Promoter Interactions (A) Model depicting CRISPR/Cas9-mediated deletion of a YY1 binding motif in the regulatory region of a gene. (B and C) CRISPR/Cas9-mediated deletion of YY1 binding motifs in the regulatory regions of two genes, Raf1 (B) and Etv4 ), and the shaded area represents the 95% confidence interval. Three biological replicates were assayed for 4C-seq and ChIP-qPCR experiments, and six biological replicates were assayed for RT-qPCR experiments. Error bars represent the SD. All p values were determined using the Student’s t test. .

    Article Snippet: 7.5 μg of H3K27ac antibody (Abcam, ab4729) or 7.5 μg of YY1 antibody (Abcam, ab109237) was added to the tube and the tube was incubated overnight at 4°C with rotation.

    Techniques: Binding Assay, CRISPR, Chromatin Immunoprecipitation, Real-time Polymerase Chain Reaction, Quantitative RT-PCR

    Depletion of YY1 Disrupts Enhancer-Promoter Looping (A) Scatterplot displaying for all YY1-YY1 enhancer-promoter interactions the change in normalized interaction frequency (log 2 fold change) upon degradation of YY1, as measured by H3K27ac HiChIP, and plotted against the normalized interaction frequency in untreated cells. (B) Change in normalized interaction frequency (log 2 fold change) upon degradation of YY1 for three different classes of interactions: all interactions, interactions not associated with YY1 ChIP-seq peaks, and YY1-YY1 enhancer-promoter interactions. (C) Scatterplot displaying for each gene associated with a YY1-YY1 enhancer-promoter interaction the change in gene expression (log 2 fold change) upon degradation of YY1 plotted against the expression in untreated cells. Genes that showed significant changes in expression (FDR adjusted p value

    Journal: Cell

    Article Title: YY1 Is a Structural Regulator of Enhancer-Promoter Loops

    doi: 10.1016/j.cell.2017.11.008

    Figure Lengend Snippet: Depletion of YY1 Disrupts Enhancer-Promoter Looping (A) Scatterplot displaying for all YY1-YY1 enhancer-promoter interactions the change in normalized interaction frequency (log 2 fold change) upon degradation of YY1, as measured by H3K27ac HiChIP, and plotted against the normalized interaction frequency in untreated cells. (B) Change in normalized interaction frequency (log 2 fold change) upon degradation of YY1 for three different classes of interactions: all interactions, interactions not associated with YY1 ChIP-seq peaks, and YY1-YY1 enhancer-promoter interactions. (C) Scatterplot displaying for each gene associated with a YY1-YY1 enhancer-promoter interaction the change in gene expression (log 2 fold change) upon degradation of YY1 plotted against the expression in untreated cells. Genes that showed significant changes in expression (FDR adjusted p value

    Article Snippet: 7.5 μg of H3K27ac antibody (Abcam, ab4729) or 7.5 μg of YY1 antibody (Abcam, ab109237) was added to the tube and the tube was incubated overnight at 4°C with rotation.

    Techniques: HiChIP, Chromatin Immunoprecipitation, Expressing

    YY1 Is a Candidate Enhancer-Promoter Structuring Factor . Candidate enhancer-promoter structuring factors identified by ChIP-MS are indicated as dots, and those identified as cell essential (CS

    Journal: Cell

    Article Title: YY1 Is a Structural Regulator of Enhancer-Promoter Loops

    doi: 10.1016/j.cell.2017.11.008

    Figure Lengend Snippet: YY1 Is a Candidate Enhancer-Promoter Structuring Factor . Candidate enhancer-promoter structuring factors identified by ChIP-MS are indicated as dots, and those identified as cell essential (CS

    Article Snippet: 7.5 μg of H3K27ac antibody (Abcam, ab4729) or 7.5 μg of YY1 antibody (Abcam, ab109237) was added to the tube and the tube was incubated overnight at 4°C with rotation.

    Techniques: Chromatin Immunoprecipitation, Mass Spectrometry

    CXCR4 regulates let-7a expression via YY1.

    Journal: The Journal of Clinical Investigation

    Article Title: CXCR4 downregulation of let-7a drives chemoresistance in acute myeloid leukemia

    doi: 10.1172/JCI66553

    Figure Lengend Snippet: CXCR4 regulates let-7a expression via YY1.

    Article Snippet: ChIP assays were performed with anti-YY1 antibody (ab12132; Abcam) and anti-H3K27 antibody (07-449; Millipore) as recommended by the manufacturer (EZ-Chip; Millipore).

    Techniques: Expressing

    Summary of temporal expression profiles of key transcription factors in the SC lineage. Sox9 and Sox10 are expressed throughout SC genesis, beginning at E9.0 in migrating NCCs and persisting until P65 in myelinating and non-myelinating SCs in the sciatic nerve. AP2α, Pax3 and Etv5 are also expressed in NCCs, persisting until E12.5 in SCPs with Nfatc4 expression being restricted to NCCs. Egr1 and Sox2 expression is initiated in SCPs at E12.5, persisting until E14.5 in iSCs. At E18.5, AP2α is expressed in the DRG but is undetectable in the nerves while the expression of Jun, Oct6, and Yy1 is initiated and persists till P65. Rare Sox2 + Sox10 + positive cells are also detected in the P65 nerve. Egr2 expression is not detected until P7 in myelinating SCs in the sciatic nerve. Post-injury, high upregulation in expression of Sox2, Oct6, and Jun is observed, while distinct nuclear Egr1 expression is also detected. The straight lines represent continued expression of the markers through the different stages, while the dotted lines represent declining or low expression. The asterisk indicates where expression is restricted to the DRG and is undetectable in the nerves at E18.5. Markers expressed in the nerve after an acute injury have been denoted with ‘√’. The green cells represent the developing axon, while the beige cells represent the NCCs at E9.0-E10.5 and the developing SCs at E12.5 to post-natal stages.

    Journal: PLoS ONE

    Article Title: Temporal Analysis of Gene Expression in the Murine Schwann Cell Lineage and the Acutely Injured Postnatal Nerve

    doi: 10.1371/journal.pone.0153256

    Figure Lengend Snippet: Summary of temporal expression profiles of key transcription factors in the SC lineage. Sox9 and Sox10 are expressed throughout SC genesis, beginning at E9.0 in migrating NCCs and persisting until P65 in myelinating and non-myelinating SCs in the sciatic nerve. AP2α, Pax3 and Etv5 are also expressed in NCCs, persisting until E12.5 in SCPs with Nfatc4 expression being restricted to NCCs. Egr1 and Sox2 expression is initiated in SCPs at E12.5, persisting until E14.5 in iSCs. At E18.5, AP2α is expressed in the DRG but is undetectable in the nerves while the expression of Jun, Oct6, and Yy1 is initiated and persists till P65. Rare Sox2 + Sox10 + positive cells are also detected in the P65 nerve. Egr2 expression is not detected until P7 in myelinating SCs in the sciatic nerve. Post-injury, high upregulation in expression of Sox2, Oct6, and Jun is observed, while distinct nuclear Egr1 expression is also detected. The straight lines represent continued expression of the markers through the different stages, while the dotted lines represent declining or low expression. The asterisk indicates where expression is restricted to the DRG and is undetectable in the nerves at E18.5. Markers expressed in the nerve after an acute injury have been denoted with ‘√’. The green cells represent the developing axon, while the beige cells represent the NCCs at E9.0-E10.5 and the developing SCs at E12.5 to post-natal stages.

    Article Snippet: Primary antibodies included: rabbit anti-AP2α (Abcam ab52222; 1:200), rabbit anti-Egr1 (Aviva Systems Biology ARP32241_P050; 1:200), rabbit anti-Egr2 (Bioss Antibodies bs-8368R; 1:200, Santa Cruz Biotechnology sc-20690; 1:200), mouse anti-Egr2 (Abcam ab168771; 1:50), rabbit anti-Etv5 (Abcam ab102010; 1:300), rabbit anti-Jun (Abcam ab31419; 1:300), rabbit anti-Ki67 (Vector Laboratories #VP-K451), rabbit anti-Nfatc4 (Abcam ab3447; 1:200), mouse anti-NeuN (Millipore MAB377; 1:200), goat anti-Oct6 C-20 (Santa Cruz Biotechnology sc-11661; 1:50), mouse anti-Pax3 (Developmental Studies Hybridoma Bank; 1:5), rabbit anti-Sox2 (Cell Signaling #3728; 1:200), rabbit anti-Sox9 (Millipore AB5535; 1:500), goat anti-Sox10 (Santa Cruz Biotechnology sc-17343; 1:400), rabbit anti-Sox10 (Millipore AB5727; 1:200), and rabbit anti-Yy1 (Abcam ab12132; 1:200).

    Techniques: Expressing

    Expression of SC lineage markers in E18.5 late immature/pro-myelinating SCs. (A-K) Co-labeling of Sox10 with Sox9 (A-B'), AP2α (C-D'), Jun (E-F') and Yy1 (I-J'), and co-labeling of Sox9 and Oct6 (G-H') in transverse sections through the E18.5 trunk. Low magnification merged images of protein of interest (red) and Sox10 (green) (A,C,E,I) or Sox9 (G). High magnification images of the DRG, showing merged images of the protein of interest (red) and Sox10 (or Sox9) in green (B,D,F,H,J), and single protein of interest images in white (B',D',F',H',J'). Blue is DAPI counterstain in A-I. Arrows indicate co-expression of proteins of interest with Sox10 (or Sox9) in late immature and pro-myelinating SCs. Quantification of percentage of Sox10 + cells that co-express each of the proteins of interest (K). Error bars = S.E.M. drg, dorsal root ganglion; sc, spinal cord; sn, spinal nerve; vr, ventral root. Scale bars, 60μm.

    Journal: PLoS ONE

    Article Title: Temporal Analysis of Gene Expression in the Murine Schwann Cell Lineage and the Acutely Injured Postnatal Nerve

    doi: 10.1371/journal.pone.0153256

    Figure Lengend Snippet: Expression of SC lineage markers in E18.5 late immature/pro-myelinating SCs. (A-K) Co-labeling of Sox10 with Sox9 (A-B'), AP2α (C-D'), Jun (E-F') and Yy1 (I-J'), and co-labeling of Sox9 and Oct6 (G-H') in transverse sections through the E18.5 trunk. Low magnification merged images of protein of interest (red) and Sox10 (green) (A,C,E,I) or Sox9 (G). High magnification images of the DRG, showing merged images of the protein of interest (red) and Sox10 (or Sox9) in green (B,D,F,H,J), and single protein of interest images in white (B',D',F',H',J'). Blue is DAPI counterstain in A-I. Arrows indicate co-expression of proteins of interest with Sox10 (or Sox9) in late immature and pro-myelinating SCs. Quantification of percentage of Sox10 + cells that co-express each of the proteins of interest (K). Error bars = S.E.M. drg, dorsal root ganglion; sc, spinal cord; sn, spinal nerve; vr, ventral root. Scale bars, 60μm.

    Article Snippet: Primary antibodies included: rabbit anti-AP2α (Abcam ab52222; 1:200), rabbit anti-Egr1 (Aviva Systems Biology ARP32241_P050; 1:200), rabbit anti-Egr2 (Bioss Antibodies bs-8368R; 1:200, Santa Cruz Biotechnology sc-20690; 1:200), mouse anti-Egr2 (Abcam ab168771; 1:50), rabbit anti-Etv5 (Abcam ab102010; 1:300), rabbit anti-Jun (Abcam ab31419; 1:300), rabbit anti-Ki67 (Vector Laboratories #VP-K451), rabbit anti-Nfatc4 (Abcam ab3447; 1:200), mouse anti-NeuN (Millipore MAB377; 1:200), goat anti-Oct6 C-20 (Santa Cruz Biotechnology sc-11661; 1:50), mouse anti-Pax3 (Developmental Studies Hybridoma Bank; 1:5), rabbit anti-Sox2 (Cell Signaling #3728; 1:200), rabbit anti-Sox9 (Millipore AB5535; 1:500), goat anti-Sox10 (Santa Cruz Biotechnology sc-17343; 1:400), rabbit anti-Sox10 (Millipore AB5727; 1:200), and rabbit anti-Yy1 (Abcam ab12132; 1:200).

    Techniques: Expressing, Labeling

    Expression of SC lineage markers in P7 mature SCs. (A-F'') Co-labeling of Sox10 with Sox9 (A-A''), Nfatc4 (B-B''), Jun (C-C''), Oct6 (D-D''), Yy1 (E-E''), and Egr2 (F-F'') in longitudinal sections of the P7 sciatic nerve. Merged images of the protein of interest in red and Sox10 in green (A-F). Blue is DAPI counterstain. Expression profiles of the protein of interest (A'-F') and Sox10 (A''-F''). Arrows indicate co-expression of proteins of interest with Sox10 in P7 mature SCs Quantification of percentage of Sox10 + cells that co-express each of the proteins of interest (G). Error bars = S.E.M. Scale bars, 40μm.

    Journal: PLoS ONE

    Article Title: Temporal Analysis of Gene Expression in the Murine Schwann Cell Lineage and the Acutely Injured Postnatal Nerve

    doi: 10.1371/journal.pone.0153256

    Figure Lengend Snippet: Expression of SC lineage markers in P7 mature SCs. (A-F'') Co-labeling of Sox10 with Sox9 (A-A''), Nfatc4 (B-B''), Jun (C-C''), Oct6 (D-D''), Yy1 (E-E''), and Egr2 (F-F'') in longitudinal sections of the P7 sciatic nerve. Merged images of the protein of interest in red and Sox10 in green (A-F). Blue is DAPI counterstain. Expression profiles of the protein of interest (A'-F') and Sox10 (A''-F''). Arrows indicate co-expression of proteins of interest with Sox10 in P7 mature SCs Quantification of percentage of Sox10 + cells that co-express each of the proteins of interest (G). Error bars = S.E.M. Scale bars, 40μm.

    Article Snippet: Primary antibodies included: rabbit anti-AP2α (Abcam ab52222; 1:200), rabbit anti-Egr1 (Aviva Systems Biology ARP32241_P050; 1:200), rabbit anti-Egr2 (Bioss Antibodies bs-8368R; 1:200, Santa Cruz Biotechnology sc-20690; 1:200), mouse anti-Egr2 (Abcam ab168771; 1:50), rabbit anti-Etv5 (Abcam ab102010; 1:300), rabbit anti-Jun (Abcam ab31419; 1:300), rabbit anti-Ki67 (Vector Laboratories #VP-K451), rabbit anti-Nfatc4 (Abcam ab3447; 1:200), mouse anti-NeuN (Millipore MAB377; 1:200), goat anti-Oct6 C-20 (Santa Cruz Biotechnology sc-11661; 1:50), mouse anti-Pax3 (Developmental Studies Hybridoma Bank; 1:5), rabbit anti-Sox2 (Cell Signaling #3728; 1:200), rabbit anti-Sox9 (Millipore AB5535; 1:500), goat anti-Sox10 (Santa Cruz Biotechnology sc-17343; 1:400), rabbit anti-Sox10 (Millipore AB5727; 1:200), and rabbit anti-Yy1 (Abcam ab12132; 1:200).

    Techniques: Expressing, Labeling

    Expression of SC lineage markers in P65 sciatic nerve after acute nerve injury. (A-E) Co-labeling of Sox10 (red) with Sox9 (A-A''), Nfatc4 (B-B''), Egr1 (C-C''), Yy1 (D-D''), and Egr2 (E-E'') in longitudinal sections of the injured P65 sciatic nerve. Merged images of protein of interest (green), Sox10 (red) and Hoechst (blue) (A-E). Yellow arrowheads indicate co-labelled cells. Scale bars, 20μm.

    Journal: PLoS ONE

    Article Title: Temporal Analysis of Gene Expression in the Murine Schwann Cell Lineage and the Acutely Injured Postnatal Nerve

    doi: 10.1371/journal.pone.0153256

    Figure Lengend Snippet: Expression of SC lineage markers in P65 sciatic nerve after acute nerve injury. (A-E) Co-labeling of Sox10 (red) with Sox9 (A-A''), Nfatc4 (B-B''), Egr1 (C-C''), Yy1 (D-D''), and Egr2 (E-E'') in longitudinal sections of the injured P65 sciatic nerve. Merged images of protein of interest (green), Sox10 (red) and Hoechst (blue) (A-E). Yellow arrowheads indicate co-labelled cells. Scale bars, 20μm.

    Article Snippet: Primary antibodies included: rabbit anti-AP2α (Abcam ab52222; 1:200), rabbit anti-Egr1 (Aviva Systems Biology ARP32241_P050; 1:200), rabbit anti-Egr2 (Bioss Antibodies bs-8368R; 1:200, Santa Cruz Biotechnology sc-20690; 1:200), mouse anti-Egr2 (Abcam ab168771; 1:50), rabbit anti-Etv5 (Abcam ab102010; 1:300), rabbit anti-Jun (Abcam ab31419; 1:300), rabbit anti-Ki67 (Vector Laboratories #VP-K451), rabbit anti-Nfatc4 (Abcam ab3447; 1:200), mouse anti-NeuN (Millipore MAB377; 1:200), goat anti-Oct6 C-20 (Santa Cruz Biotechnology sc-11661; 1:50), mouse anti-Pax3 (Developmental Studies Hybridoma Bank; 1:5), rabbit anti-Sox2 (Cell Signaling #3728; 1:200), rabbit anti-Sox9 (Millipore AB5535; 1:500), goat anti-Sox10 (Santa Cruz Biotechnology sc-17343; 1:400), rabbit anti-Sox10 (Millipore AB5727; 1:200), and rabbit anti-Yy1 (Abcam ab12132; 1:200).

    Techniques: Expressing, Labeling

    Expression of SC lineage markers in adult P65 uninjured sciatic nerve. (A-I'') Co-labeling of Sox10 with fluoromyelin (A-A‴), Sox9 (B-B''), Nfatc4 (C-C''), Sox2 (D-D''), Egr1 (E-E''), and Oct6 (F-F''), Jun (G-G''), Yy1 (H-H''), Egr2 (I-I'') in longitudinal sections of the uninjured P65 sciatic nerve. Merged images of protein of interest (green), Sox10 (red) and Hoechst (blue) (B-I). Yellow arrowheads indicate co-labelled cells. Scale bars, 20μm.

    Journal: PLoS ONE

    Article Title: Temporal Analysis of Gene Expression in the Murine Schwann Cell Lineage and the Acutely Injured Postnatal Nerve

    doi: 10.1371/journal.pone.0153256

    Figure Lengend Snippet: Expression of SC lineage markers in adult P65 uninjured sciatic nerve. (A-I'') Co-labeling of Sox10 with fluoromyelin (A-A‴), Sox9 (B-B''), Nfatc4 (C-C''), Sox2 (D-D''), Egr1 (E-E''), and Oct6 (F-F''), Jun (G-G''), Yy1 (H-H''), Egr2 (I-I'') in longitudinal sections of the uninjured P65 sciatic nerve. Merged images of protein of interest (green), Sox10 (red) and Hoechst (blue) (B-I). Yellow arrowheads indicate co-labelled cells. Scale bars, 20μm.

    Article Snippet: Primary antibodies included: rabbit anti-AP2α (Abcam ab52222; 1:200), rabbit anti-Egr1 (Aviva Systems Biology ARP32241_P050; 1:200), rabbit anti-Egr2 (Bioss Antibodies bs-8368R; 1:200, Santa Cruz Biotechnology sc-20690; 1:200), mouse anti-Egr2 (Abcam ab168771; 1:50), rabbit anti-Etv5 (Abcam ab102010; 1:300), rabbit anti-Jun (Abcam ab31419; 1:300), rabbit anti-Ki67 (Vector Laboratories #VP-K451), rabbit anti-Nfatc4 (Abcam ab3447; 1:200), mouse anti-NeuN (Millipore MAB377; 1:200), goat anti-Oct6 C-20 (Santa Cruz Biotechnology sc-11661; 1:50), mouse anti-Pax3 (Developmental Studies Hybridoma Bank; 1:5), rabbit anti-Sox2 (Cell Signaling #3728; 1:200), rabbit anti-Sox9 (Millipore AB5535; 1:500), goat anti-Sox10 (Santa Cruz Biotechnology sc-17343; 1:400), rabbit anti-Sox10 (Millipore AB5727; 1:200), and rabbit anti-Yy1 (Abcam ab12132; 1:200).

    Techniques: Expressing, Labeling

    FABP7 promoter cis -regulatory region identified by promoter deletion analysis . Numbering starts from the FABP7 transcription start site [ 17 ]. Oligonucleotides designated upstream (UP), middle (MP) and downstream (DP) probes are shown along with putative consensus binding sites for OCT1, C/EBP-α, BRN2, OCT6, NFI, C/EBP-β, SP1, and YY1.

    Journal: BMC Molecular Biology

    Article Title: Analysis of the regulation of fatty acid binding protein 7 expression in human renal carcinoma cell lines

    doi: 10.1186/1471-2199-12-31

    Figure Lengend Snippet: FABP7 promoter cis -regulatory region identified by promoter deletion analysis . Numbering starts from the FABP7 transcription start site [ 17 ]. Oligonucleotides designated upstream (UP), middle (MP) and downstream (DP) probes are shown along with putative consensus binding sites for OCT1, C/EBP-α, BRN2, OCT6, NFI, C/EBP-β, SP1, and YY1.

    Article Snippet: Goat anti-mouse IgG-HRP (sc-2005), anti-Brn-2 (C-20: sc-6029 X), anti-Lamin B (M-20: sc-6217), anti-NF-1 (H-300: sc-5567 X), anti-Oct-1 (C-21: sc-232 X), anti-Oct-6 (H-13: sc-11660 X) and anti-YY1 (H-414: sc-1703 X) were purchased from Santa Cruz Biotechnology, Inc. (Santa Cruz, CA).

    Techniques: Binding Assay

    YY1 knockdown reduces viral integration into the host genome. (A) Knockdown of YY1 by siRNA was confirmed by Western blotting. (B and C) The amount of total viral cDNA (B) and two-LTR circle (C) of pQEGFP (siRNA no. 1006) and pLNΔAG (siRNA no. 1099) viral vectors is shown. NIH 3T3 cells were transfected twice with YY1 siRNA no. 1006 or no. 1099 and then infected with pQEGFP (MOI of 0.1) or pLNΔAG (MOI of 1). Cellular DNA was extracted at 0, 4, 10, 24, and 48 h and subjected to qPCR analysis. Copy number per cell was calculated by normalizing the value of the viral sequence with that of GAPDH (glyceraldehyde-3-phosphate dehydrogenase). Error bars represent the standard deviations of the results of at least three experiments. Primers used for PCR for GAPDH were 5′-TGTGATGGGTGTGAACCACGAGAA-3′ and 5′-GAGCCCTTCCACAATGCCAAAGTT-3′. Asterisks indicate that values of YY1 siRNA compared to those of scrambled RNA were significantly different (*, P = 0.038, and **, P = 0.031 in panel B; *, P = 0.022, **, P = 0.041, and ***, P = 0.047 in panel C). (D) The integrated form of viral DNA was quantified 14 days postinfection by qPCR and normalized as described for the results presented in panel C. Asterisks indicate significance of values of YY1 siRNA versus those of scrambled RNA (*, P = 0.014, **, P = 0.016, and ***, P

    Journal: Journal of Virology

    Article Title: Transcription Factor YY1 Interacts with Retroviral Integrases and Facilitates Integration of Moloney Murine Leukemia Virus cDNA into the Host Chromosomes ▿

    doi: 10.1128/JVI.02681-09

    Figure Lengend Snippet: YY1 knockdown reduces viral integration into the host genome. (A) Knockdown of YY1 by siRNA was confirmed by Western blotting. (B and C) The amount of total viral cDNA (B) and two-LTR circle (C) of pQEGFP (siRNA no. 1006) and pLNΔAG (siRNA no. 1099) viral vectors is shown. NIH 3T3 cells were transfected twice with YY1 siRNA no. 1006 or no. 1099 and then infected with pQEGFP (MOI of 0.1) or pLNΔAG (MOI of 1). Cellular DNA was extracted at 0, 4, 10, 24, and 48 h and subjected to qPCR analysis. Copy number per cell was calculated by normalizing the value of the viral sequence with that of GAPDH (glyceraldehyde-3-phosphate dehydrogenase). Error bars represent the standard deviations of the results of at least three experiments. Primers used for PCR for GAPDH were 5′-TGTGATGGGTGTGAACCACGAGAA-3′ and 5′-GAGCCCTTCCACAATGCCAAAGTT-3′. Asterisks indicate that values of YY1 siRNA compared to those of scrambled RNA were significantly different (*, P = 0.038, and **, P = 0.031 in panel B; *, P = 0.022, **, P = 0.041, and ***, P = 0.047 in panel C). (D) The integrated form of viral DNA was quantified 14 days postinfection by qPCR and normalized as described for the results presented in panel C. Asterisks indicate significance of values of YY1 siRNA versus those of scrambled RNA (*, P = 0.014, **, P = 0.016, and ***, P

    Article Snippet: Anti-YY1 antibody (C-20) was obtained from Santa Cruz.

    Techniques: Western Blot, Transfection, Infection, Real-time Polymerase Chain Reaction, Sequencing, Polymerase Chain Reaction

    Identification of the IN-binding region in YY1 in vitro and in vivo . (A) Schematic drawing of full-length YY1 and its fragments used for pulldown and coimmunoprecipitation experiments. His, histidine-rich domain; GA, glycine-alanine-rich domain; GK, glycine-lysine-rich domain. The results of in vitro pulldown and immunoprecipitation experiments are summarized on the right side of the panel. (B) Results of pulldown assay of MoMLV IN with GST fusion proteins formed with various portions of human YY1. His-MoMLV IN was detected by Western blotting using anti-MoMLV IN antiserum. GST and GST-YY1 fragments were subjected to SDS-PAGE, and the gel was stained with Coomassie brilliant blue. Arrowheads show the intact fragments of GST-YY1 derivatives (lower panel). A result representative of several experiments is presented. (C) Results of coimmunoprecipitation of human YY1 and MoMLV IN expressed in 293FT cells (upper panels). Normal mouse IgG was used in the control, and nonspecific bands are indicated by asterisks. Cells were lysed with buffer A. IN complex was precipitated and washed with buffer A (left and central panels) or buffer B (right panel). In the right panel, controls without IN were included to estimate a nonspecific interaction between YY1 and anti-Flag antibody observed in lane 8, since interaction between IN and some YY1 derivatives seemed to be weak. His-tagged fragments of human YY1 were detected with anti-YY1 or anti-His antibody. These antibodies were also used for detection of input YY1 derivatives (2.5%). Flag-IN was detected with anti-Flag antibody. A result representative of several experiments is presented.

    Journal: Journal of Virology

    Article Title: Transcription Factor YY1 Interacts with Retroviral Integrases and Facilitates Integration of Moloney Murine Leukemia Virus cDNA into the Host Chromosomes ▿

    doi: 10.1128/JVI.02681-09

    Figure Lengend Snippet: Identification of the IN-binding region in YY1 in vitro and in vivo . (A) Schematic drawing of full-length YY1 and its fragments used for pulldown and coimmunoprecipitation experiments. His, histidine-rich domain; GA, glycine-alanine-rich domain; GK, glycine-lysine-rich domain. The results of in vitro pulldown and immunoprecipitation experiments are summarized on the right side of the panel. (B) Results of pulldown assay of MoMLV IN with GST fusion proteins formed with various portions of human YY1. His-MoMLV IN was detected by Western blotting using anti-MoMLV IN antiserum. GST and GST-YY1 fragments were subjected to SDS-PAGE, and the gel was stained with Coomassie brilliant blue. Arrowheads show the intact fragments of GST-YY1 derivatives (lower panel). A result representative of several experiments is presented. (C) Results of coimmunoprecipitation of human YY1 and MoMLV IN expressed in 293FT cells (upper panels). Normal mouse IgG was used in the control, and nonspecific bands are indicated by asterisks. Cells were lysed with buffer A. IN complex was precipitated and washed with buffer A (left and central panels) or buffer B (right panel). In the right panel, controls without IN were included to estimate a nonspecific interaction between YY1 and anti-Flag antibody observed in lane 8, since interaction between IN and some YY1 derivatives seemed to be weak. His-tagged fragments of human YY1 were detected with anti-YY1 or anti-His antibody. These antibodies were also used for detection of input YY1 derivatives (2.5%). Flag-IN was detected with anti-Flag antibody. A result representative of several experiments is presented.

    Article Snippet: Anti-YY1 antibody (C-20) was obtained from Santa Cruz.

    Techniques: Binding Assay, In Vitro, In Vivo, Immunoprecipitation, Western Blot, SDS Page, Staining

    G9a silences the expression of HEPH via assembling a co-repressor complex with YY1 and HDAC1. a – c MCF-7 cells were transfected with two independent YY1 siRNAs. After 48 h, HEPH protein and mRNA levels and HEPH promoter luciferase activity were examined. Expression of the transfected constructs is shown in the western blotting analysis. d pGL3- HEPH promoter and the indicated constructs were co-transfected into MDA-MB-231 cells. Twenty-four hours after transfection, cell extracts were assayed for luciferase activity. e Silencing and overexpression of HDAC1, but not HDAC2, contributed to the upregulation or downregulation of HEPH mRNA and protein level, respectively. The HDAC1-specific inhibitor MS275 was synergetic with UNC0638 in increasing HEPH expression in a time-dependent manner. f , g The abundance of H3K9-me2 and the binding levels of G9a and HDAC1 in the Pro2 region of the HEPH promoter were determined by ChIP in G9a knockdown or overexpressed cells treated with siYY1. The results are presented as means ± SD from three independent experiments. Two-tailed unpaired Student’s T -test was performed. * P

    Journal: Nature Communications

    Article Title: G9a regulates breast cancer growth by modulating iron homeostasis through the repression of ferroxidase hephaestin

    doi: 10.1038/s41467-017-00350-9

    Figure Lengend Snippet: G9a silences the expression of HEPH via assembling a co-repressor complex with YY1 and HDAC1. a – c MCF-7 cells were transfected with two independent YY1 siRNAs. After 48 h, HEPH protein and mRNA levels and HEPH promoter luciferase activity were examined. Expression of the transfected constructs is shown in the western blotting analysis. d pGL3- HEPH promoter and the indicated constructs were co-transfected into MDA-MB-231 cells. Twenty-four hours after transfection, cell extracts were assayed for luciferase activity. e Silencing and overexpression of HDAC1, but not HDAC2, contributed to the upregulation or downregulation of HEPH mRNA and protein level, respectively. The HDAC1-specific inhibitor MS275 was synergetic with UNC0638 in increasing HEPH expression in a time-dependent manner. f , g The abundance of H3K9-me2 and the binding levels of G9a and HDAC1 in the Pro2 region of the HEPH promoter were determined by ChIP in G9a knockdown or overexpressed cells treated with siYY1. The results are presented as means ± SD from three independent experiments. Two-tailed unpaired Student’s T -test was performed. * P

    Article Snippet: The soluble fraction was collected and the chromatins were incubated with 5 μl of anti-K9 dimethylated histone H3, anti-histone H3, anti-G9a or anti-YY1 (Cell Signaling) at 4 °C overnight.

    Techniques: Expressing, Transfection, Luciferase, Activity Assay, Construct, Western Blot, Multiple Displacement Amplification, Over Expression, Binding Assay, Chromatin Immunoprecipitation, Two Tailed Test

    High levels of G9a and low levels of HEPH correlate with poor survival in breast cancer. a G9a and HEPH prognostic interactions. Associations between OS and high or low G9a and HEPH expression levels (based on mean partitioning) in a combined multi-institutional population-based cohort consisting of 75 breast cancer cases. Kaplan–Meier plots and log-rank P -values are shown for (I) HEPH expression, (II) G9a expression, (III) high G9a dichotomized by low HEPH. b Public breast cancer database (KM-Plotter) was queried to examine the association between patients with breast cancer RFS and HEPH expression, the log-rank test P -value was indicated. c Schematic diagram depicting the regulation of HEPH in breast tumor cells. G9a as an HMTase activity-dependent repressor collaborates in the complex with YY1 and HDAC1, and works coordinately to contribute to the reduction of HEPH expression. Silencing of G9a upregulates HEPH, inhibits breast cancer cell proliferation and cell survival via upregulation of HEPH transcription and induces HEPH-mediated iron homeostasis disruption upon greater iron export. Green particles represent ferrous iron; red particles represent ferric iron

    Journal: Nature Communications

    Article Title: G9a regulates breast cancer growth by modulating iron homeostasis through the repression of ferroxidase hephaestin

    doi: 10.1038/s41467-017-00350-9

    Figure Lengend Snippet: High levels of G9a and low levels of HEPH correlate with poor survival in breast cancer. a G9a and HEPH prognostic interactions. Associations between OS and high or low G9a and HEPH expression levels (based on mean partitioning) in a combined multi-institutional population-based cohort consisting of 75 breast cancer cases. Kaplan–Meier plots and log-rank P -values are shown for (I) HEPH expression, (II) G9a expression, (III) high G9a dichotomized by low HEPH. b Public breast cancer database (KM-Plotter) was queried to examine the association between patients with breast cancer RFS and HEPH expression, the log-rank test P -value was indicated. c Schematic diagram depicting the regulation of HEPH in breast tumor cells. G9a as an HMTase activity-dependent repressor collaborates in the complex with YY1 and HDAC1, and works coordinately to contribute to the reduction of HEPH expression. Silencing of G9a upregulates HEPH, inhibits breast cancer cell proliferation and cell survival via upregulation of HEPH transcription and induces HEPH-mediated iron homeostasis disruption upon greater iron export. Green particles represent ferrous iron; red particles represent ferric iron

    Article Snippet: The soluble fraction was collected and the chromatins were incubated with 5 μl of anti-K9 dimethylated histone H3, anti-histone H3, anti-G9a or anti-YY1 (Cell Signaling) at 4 °C overnight.

    Techniques: Expressing, Activity Assay

    SP1 and YY1 silencing increases HAS2 mRNA. HaCaT cell cultures transfected with control, SP1 , and YY1 siRNAs were analyzed for HAS2 mRNA normalized to the control gene RPLP0 as described under “Experimental Procedures.” The values are related to non-transfected cultures and represent means ± S.E. of four independent experiments (each with two replicates). Statistical significances between cultures treated with control siRNA versus SP1 and YY1 siRNA are as follows: *, p

    Journal: The Journal of Biological Chemistry

    Article Title: Cellular Content of UDP-N-acetylhexosamines Controls Hyaluronan Synthase 2 Expression and Correlates with O-Linked N-Acetylglucosamine Modification of Transcription Factors YY1 and SP1 *

    doi: 10.1074/jbc.M111.265637

    Figure Lengend Snippet: SP1 and YY1 silencing increases HAS2 mRNA. HaCaT cell cultures transfected with control, SP1 , and YY1 siRNAs were analyzed for HAS2 mRNA normalized to the control gene RPLP0 as described under “Experimental Procedures.” The values are related to non-transfected cultures and represent means ± S.E. of four independent experiments (each with two replicates). Statistical significances between cultures treated with control siRNA versus SP1 and YY1 siRNA are as follows: *, p

    Article Snippet: Total protein (500–900 μg) was incubated with 1.5 μg of anti-SP1 antibody (Santa Cruz Biotechnology or Abcam, Cambridge, United Kingdom) or anti-YY1 antibody (Cell Signaling Technology, Danvers, MA) linked to 20 μl of Protein AG magnetic beads (Ademtech, Pessac, France) or protein A/G-agarose (Santa Cruz Biotechnology) overnight at 4 °C.

    Techniques: Transfection

    YY1 depletion represses PCa cell viability and colony forming in a miR-146a-assisted manner. (A–C) Western blot assays were performed to demonstrate the expression effect of siYY1, pCMV6 and sh-YY1 on PCa cell lines. GAPDH was used as endogenous control. (D and E) miR-146a expression by qRT-PCR verified the inhibitory efficiency of si-YY1+antiNC/si-YY1+anti-miR-146a in both cell lines. The cell lines were transfected with si-YY1/si-NC, pCMV6-YY1/pCMV6, and si-YY1+anti-NC/si-YY1+anti-miR-146a. Cell viability and colony forming were measured by MTT (F-K) and colony formation assays (L-N). (O) GSEA showed enriched expression of gene sets involved in cell proliferation in YY1 knocked-down cells. * P

    Journal: International Journal of Oncology

    Article Title: Upregulation of miR-146a by YY1 depletion correlates with delayed progression of prostate cancer

    doi: 10.3892/ijo.2017.3840

    Figure Lengend Snippet: YY1 depletion represses PCa cell viability and colony forming in a miR-146a-assisted manner. (A–C) Western blot assays were performed to demonstrate the expression effect of siYY1, pCMV6 and sh-YY1 on PCa cell lines. GAPDH was used as endogenous control. (D and E) miR-146a expression by qRT-PCR verified the inhibitory efficiency of si-YY1+antiNC/si-YY1+anti-miR-146a in both cell lines. The cell lines were transfected with si-YY1/si-NC, pCMV6-YY1/pCMV6, and si-YY1+anti-NC/si-YY1+anti-miR-146a. Cell viability and colony forming were measured by MTT (F-K) and colony formation assays (L-N). (O) GSEA showed enriched expression of gene sets involved in cell proliferation in YY1 knocked-down cells. * P

    Article Snippet: Total cell extracts were pre-cleared with Protein A+G beads (Beyotime) at 4°C for 2 h. The supernatant was incubated with the anti-EZH2 or anti-YY1 (EZH2, 1:200; YY1, 1:200; Proteintech, Chicago, IL, USA) with gentle shacking for 10 h at 4°C followed by addition of 30 µ l of Protein A+G beads for another 2 h. The beads were re-suspended in 100 µ l of 2X loading buffer and boiled for 10 min. Then western blot assay was performed according to the instructions previously described to detect the YY1 and EZH2.

    Techniques: Western Blot, Expressing, Quantitative RT-PCR, Transfection, MTT Assay

    miR-146a is negatively regulated by YY1 at the transcriptional level. (A–D) miR-146a expression by qRT-PCR detected the effect of si-YY1/si-NC, pCMV6-YY1/pCMV6 in both PCa cell lines. (E) Schematic of putative YY1 binding motifs in the miR-146a promoter region (−1,262 bp upstream of the miR-146a promoter). Mutant binding motif for YY1 was chemically synthesized to generate the pGL3-basic-mutant reporter. (F) Luciferase reporter assay were performed to detect the junction of YY1 at the miR-146a promoter region (−912 to −923 bp before the transcription start site). pGL3-miR-146a reporter vector (1 µ g), together with 2 µ g of pCMV6-YY1 or pCMV6 were co-transfected into PC3 cells. Luciferase activities were determined and normalized to Renilla activity after transfection for 24 h. * P

    Journal: International Journal of Oncology

    Article Title: Upregulation of miR-146a by YY1 depletion correlates with delayed progression of prostate cancer

    doi: 10.3892/ijo.2017.3840

    Figure Lengend Snippet: miR-146a is negatively regulated by YY1 at the transcriptional level. (A–D) miR-146a expression by qRT-PCR detected the effect of si-YY1/si-NC, pCMV6-YY1/pCMV6 in both PCa cell lines. (E) Schematic of putative YY1 binding motifs in the miR-146a promoter region (−1,262 bp upstream of the miR-146a promoter). Mutant binding motif for YY1 was chemically synthesized to generate the pGL3-basic-mutant reporter. (F) Luciferase reporter assay were performed to detect the junction of YY1 at the miR-146a promoter region (−912 to −923 bp before the transcription start site). pGL3-miR-146a reporter vector (1 µ g), together with 2 µ g of pCMV6-YY1 or pCMV6 were co-transfected into PC3 cells. Luciferase activities were determined and normalized to Renilla activity after transfection for 24 h. * P

    Article Snippet: Total cell extracts were pre-cleared with Protein A+G beads (Beyotime) at 4°C for 2 h. The supernatant was incubated with the anti-EZH2 or anti-YY1 (EZH2, 1:200; YY1, 1:200; Proteintech, Chicago, IL, USA) with gentle shacking for 10 h at 4°C followed by addition of 30 µ l of Protein A+G beads for another 2 h. The beads were re-suspended in 100 µ l of 2X loading buffer and boiled for 10 min. Then western blot assay was performed according to the instructions previously described to detect the YY1 and EZH2.

    Techniques: Expressing, Quantitative RT-PCR, Binding Assay, Mutagenesis, Synthesized, Luciferase, Reporter Assay, Plasmid Preparation, Transfection, Activity Assay

    Depletion of YY1 induces cell apoptosis in a miR-146a-assisted manner. Both cell lines were transfected with si-YY1/si-NC, pCMV6-YY1/pCMV6, and si-YY1+anti-NC/si-YY1+anti-miR-146a. (A–C) Quantification of apoptosis with a fluorescence-activated cell sorter. Significance was determined by Student's t-test. * P

    Journal: International Journal of Oncology

    Article Title: Upregulation of miR-146a by YY1 depletion correlates with delayed progression of prostate cancer

    doi: 10.3892/ijo.2017.3840

    Figure Lengend Snippet: Depletion of YY1 induces cell apoptosis in a miR-146a-assisted manner. Both cell lines were transfected with si-YY1/si-NC, pCMV6-YY1/pCMV6, and si-YY1+anti-NC/si-YY1+anti-miR-146a. (A–C) Quantification of apoptosis with a fluorescence-activated cell sorter. Significance was determined by Student's t-test. * P

    Article Snippet: Total cell extracts were pre-cleared with Protein A+G beads (Beyotime) at 4°C for 2 h. The supernatant was incubated with the anti-EZH2 or anti-YY1 (EZH2, 1:200; YY1, 1:200; Proteintech, Chicago, IL, USA) with gentle shacking for 10 h at 4°C followed by addition of 30 µ l of Protein A+G beads for another 2 h. The beads were re-suspended in 100 µ l of 2X loading buffer and boiled for 10 min. Then western blot assay was performed according to the instructions previously described to detect the YY1 and EZH2.

    Techniques: Transfection, Fluorescence

    ONCOMINE database reanalysis and clinical significance of YY1 in PCa specimens. (A) Co-relationship between YY1 expression and biochemical recurrence by reanalysis of the Nakagawa prostate. (B) Co-relationship between YY1 expression and Gleason scores by reanalysis of the Nakagawa prostate. (C) Co-relationship between YY1 expression (IHC) and Gleason scores in 34 PCa specimens. * P

    Journal: International Journal of Oncology

    Article Title: Upregulation of miR-146a by YY1 depletion correlates with delayed progression of prostate cancer

    doi: 10.3892/ijo.2017.3840

    Figure Lengend Snippet: ONCOMINE database reanalysis and clinical significance of YY1 in PCa specimens. (A) Co-relationship between YY1 expression and biochemical recurrence by reanalysis of the Nakagawa prostate. (B) Co-relationship between YY1 expression and Gleason scores by reanalysis of the Nakagawa prostate. (C) Co-relationship between YY1 expression (IHC) and Gleason scores in 34 PCa specimens. * P

    Article Snippet: Total cell extracts were pre-cleared with Protein A+G beads (Beyotime) at 4°C for 2 h. The supernatant was incubated with the anti-EZH2 or anti-YY1 (EZH2, 1:200; YY1, 1:200; Proteintech, Chicago, IL, USA) with gentle shacking for 10 h at 4°C followed by addition of 30 µ l of Protein A+G beads for another 2 h. The beads were re-suspended in 100 µ l of 2X loading buffer and boiled for 10 min. Then western blot assay was performed according to the instructions previously described to detect the YY1 and EZH2.

    Techniques: Expressing, Immunohistochemistry

    Biological insight into the co-relationship between YY1 and miR-146a. (A) GSEA revealed enriched expression of miR-146a target gene sets after YY1 depletion in PC3. (B) Spearman correlation analysis was performed to assess the correlation between miR-146a expression (ISH) and YY1 expression (IHC) in 34 PCa specimens. (C) Representative images of miR-146a expression (ISH) and YY1 expression (IHC) in PCa tissues. The positive brown signals indicate the relative expression levels of YY1 and miR-146a. Original magnification, ×200.

    Journal: International Journal of Oncology

    Article Title: Upregulation of miR-146a by YY1 depletion correlates with delayed progression of prostate cancer

    doi: 10.3892/ijo.2017.3840

    Figure Lengend Snippet: Biological insight into the co-relationship between YY1 and miR-146a. (A) GSEA revealed enriched expression of miR-146a target gene sets after YY1 depletion in PC3. (B) Spearman correlation analysis was performed to assess the correlation between miR-146a expression (ISH) and YY1 expression (IHC) in 34 PCa specimens. (C) Representative images of miR-146a expression (ISH) and YY1 expression (IHC) in PCa tissues. The positive brown signals indicate the relative expression levels of YY1 and miR-146a. Original magnification, ×200.

    Article Snippet: Total cell extracts were pre-cleared with Protein A+G beads (Beyotime) at 4°C for 2 h. The supernatant was incubated with the anti-EZH2 or anti-YY1 (EZH2, 1:200; YY1, 1:200; Proteintech, Chicago, IL, USA) with gentle shacking for 10 h at 4°C followed by addition of 30 µ l of Protein A+G beads for another 2 h. The beads were re-suspended in 100 µ l of 2X loading buffer and boiled for 10 min. Then western blot assay was performed according to the instructions previously described to detect the YY1 and EZH2.

    Techniques: Expressing, In Situ Hybridization, Immunohistochemistry

    YY1 depletion suppresses xenografts growth accompanied by miR-146 upregulation. (A and B) After establishment of subcutaneous tumors, tumor volumes were measured every week. The tumor volume of the sh-YY1 group was significantly decreased compared with the sh-NC group. (C) qRT-PCR detected the increase of miR-146a expression in sh-YY1 xenografts. * P

    Journal: International Journal of Oncology

    Article Title: Upregulation of miR-146a by YY1 depletion correlates with delayed progression of prostate cancer

    doi: 10.3892/ijo.2017.3840

    Figure Lengend Snippet: YY1 depletion suppresses xenografts growth accompanied by miR-146 upregulation. (A and B) After establishment of subcutaneous tumors, tumor volumes were measured every week. The tumor volume of the sh-YY1 group was significantly decreased compared with the sh-NC group. (C) qRT-PCR detected the increase of miR-146a expression in sh-YY1 xenografts. * P

    Article Snippet: Total cell extracts were pre-cleared with Protein A+G beads (Beyotime) at 4°C for 2 h. The supernatant was incubated with the anti-EZH2 or anti-YY1 (EZH2, 1:200; YY1, 1:200; Proteintech, Chicago, IL, USA) with gentle shacking for 10 h at 4°C followed by addition of 30 µ l of Protein A+G beads for another 2 h. The beads were re-suspended in 100 µ l of 2X loading buffer and boiled for 10 min. Then western blot assay was performed according to the instructions previously described to detect the YY1 and EZH2.

    Techniques: Quantitative RT-PCR, Expressing

    EZH2 is recruited at the promoter region of miR-146a by YY1, thereby repressing the expression of miR-146a. (A) Positively enriched expression of gene sets repressed by EZH2 are shown in YY1 knocked down PC3 cell line by GSEA. (B) Expression of miR-146a in sh-YY1 PC3 cells with EZH2 depletion by qRT-PCR showed further upregulation of miR-146a after EZH2 knockdown, suggested the potential role of EZH2 participating in the YY1-mediated regulatory process. * P

    Journal: International Journal of Oncology

    Article Title: Upregulation of miR-146a by YY1 depletion correlates with delayed progression of prostate cancer

    doi: 10.3892/ijo.2017.3840

    Figure Lengend Snippet: EZH2 is recruited at the promoter region of miR-146a by YY1, thereby repressing the expression of miR-146a. (A) Positively enriched expression of gene sets repressed by EZH2 are shown in YY1 knocked down PC3 cell line by GSEA. (B) Expression of miR-146a in sh-YY1 PC3 cells with EZH2 depletion by qRT-PCR showed further upregulation of miR-146a after EZH2 knockdown, suggested the potential role of EZH2 participating in the YY1-mediated regulatory process. * P

    Article Snippet: Total cell extracts were pre-cleared with Protein A+G beads (Beyotime) at 4°C for 2 h. The supernatant was incubated with the anti-EZH2 or anti-YY1 (EZH2, 1:200; YY1, 1:200; Proteintech, Chicago, IL, USA) with gentle shacking for 10 h at 4°C followed by addition of 30 µ l of Protein A+G beads for another 2 h. The beads were re-suspended in 100 µ l of 2X loading buffer and boiled for 10 min. Then western blot assay was performed according to the instructions previously described to detect the YY1 and EZH2.

    Techniques: Expressing, Quantitative RT-PCR

    YY1 expression for NAFLD at different stages. a The YY1 mRNA levels in the control, steatosis, non-defining NASH, and NASH groups were detected with the quantitative real-time PCR. ** p

    Journal: BMC Gastroenterology

    Article Title: Hepatic expression of Yin Yang 1 (YY1) is associated with the non-alcoholic fatty liver disease (NAFLD) progression in patients undergoing bariatric surgery

    doi: 10.1186/s12876-018-0871-2

    Figure Lengend Snippet: YY1 expression for NAFLD at different stages. a The YY1 mRNA levels in the control, steatosis, non-defining NASH, and NASH groups were detected with the quantitative real-time PCR. ** p

    Article Snippet: Sections were incubated with the rabbit anti-YY1 primary antibody (1:250dilution; Abcam, Cambridge, MA, USA) at 4 °C overnight.

    Techniques: Expressing, Real-time Polymerase Chain Reaction

    Immunohistochemistry detection of YY1 for NAFLD at different stages ( a ) Immunohistochemistry analysis was performed to detect the expression of YY1 in the control, steatosis, non-defining NASH, and NASH groups.Scale bar, 100 μm. b The cases positive or negative for YY1 expression were analyzed. * p

    Journal: BMC Gastroenterology

    Article Title: Hepatic expression of Yin Yang 1 (YY1) is associated with the non-alcoholic fatty liver disease (NAFLD) progression in patients undergoing bariatric surgery

    doi: 10.1186/s12876-018-0871-2

    Figure Lengend Snippet: Immunohistochemistry detection of YY1 for NAFLD at different stages ( a ) Immunohistochemistry analysis was performed to detect the expression of YY1 in the control, steatosis, non-defining NASH, and NASH groups.Scale bar, 100 μm. b The cases positive or negative for YY1 expression were analyzed. * p

    Article Snippet: Sections were incubated with the rabbit anti-YY1 primary antibody (1:250dilution; Abcam, Cambridge, MA, USA) at 4 °C overnight.

    Techniques: Immunohistochemistry, Expressing

    Effect of miR-29a on YY1 expression and cell behavior following IL-13 treatment. (A) MTT assay. miR-29a inhibited the cell proliferation promoted by IL-13. (B) miR-29a inhibits cell invasion in A549 cells by Transwell chamber assay. Overexpression of miR-29a or knockdown of miR-29a was carried out by transfection of A549 cells and cells were then treated with IL-13. Cell invasion ability was measured by Transwell assay. (C) Quantification of the number of invasive cells from B. (D) Measurement of the YY1 mRNA level in A549 cells treated with miR-29a overexpression vector or IL-13 stimulation. The mRNA level of YY1 was measured by RT-qPCR and normalized by GAPDH. (E) Determination of the YY1 protein level in A549 cells treated with miR-29a overexpression vector or IL-13 stimulation. The protein level of YY1 was determined by western blot analysis. (F) Quantification of the protein expression level of YY1 from E as analyzed by ImageJ software *P

    Journal: Oncology Reports

    Article Title: miR-29a suppresses IL-13-induced cell invasion by inhibiting YY1 in the AKT pathway in lung adenocarcinoma A549 cells

    doi: 10.3892/or.2018.6352

    Figure Lengend Snippet: Effect of miR-29a on YY1 expression and cell behavior following IL-13 treatment. (A) MTT assay. miR-29a inhibited the cell proliferation promoted by IL-13. (B) miR-29a inhibits cell invasion in A549 cells by Transwell chamber assay. Overexpression of miR-29a or knockdown of miR-29a was carried out by transfection of A549 cells and cells were then treated with IL-13. Cell invasion ability was measured by Transwell assay. (C) Quantification of the number of invasive cells from B. (D) Measurement of the YY1 mRNA level in A549 cells treated with miR-29a overexpression vector or IL-13 stimulation. The mRNA level of YY1 was measured by RT-qPCR and normalized by GAPDH. (E) Determination of the YY1 protein level in A549 cells treated with miR-29a overexpression vector or IL-13 stimulation. The protein level of YY1 was determined by western blot analysis. (F) Quantification of the protein expression level of YY1 from E as analyzed by ImageJ software *P

    Article Snippet: Membranes were incubated with primary antibodies: YY1 (cat. no. ab10923; Abcam, Cambridge, MA, USA), AKT (cat. no. C67E7; Cell Signaling Technology, Danvers, MA, USA), pAKT Ser473 (cat. no. 9271; Cell Signaling Technology) and N-cadherin (cat. no. D4R1H; Cell Signaling Technology) (diluted by 1:2,000) and β-actin antibody (cat. no. ZB2305; ZEGB-Bio Ltd., Beijing, China), respectively, at 4°C overnight.

    Techniques: Expressing, MTT Assay, Transwell Chamber Assay, Over Expression, Transfection, Transwell Assay, Plasmid Preparation, Quantitative RT-PCR, Western Blot, Software

    Regulation of YY1 and N-cadherin by miR-29a. (A) miR-29a inhibits the mRNA level of YY1 in A549 cells. miR-29a was overexpressed and miR-29a was knocked down by transfection of the A549 cells before measuring the mRNA level by RT-qPCR assay. (B) miR-29a reduced the protein level of YY1 in A549 cells. The protein level of YY1 was measured by western blot analysis. (C) Quantification of the protein expression level of YY1 as analyzed by ImageJ software. (D) miR-29a reduced the protein level of N-cadherin in the A549 cells. The protein level of N-cadherin was measured by western blot analysis. (E) Quantification of the protein expression level of N-cadherin as analyzed by ImageJ software. *P

    Journal: Oncology Reports

    Article Title: miR-29a suppresses IL-13-induced cell invasion by inhibiting YY1 in the AKT pathway in lung adenocarcinoma A549 cells

    doi: 10.3892/or.2018.6352

    Figure Lengend Snippet: Regulation of YY1 and N-cadherin by miR-29a. (A) miR-29a inhibits the mRNA level of YY1 in A549 cells. miR-29a was overexpressed and miR-29a was knocked down by transfection of the A549 cells before measuring the mRNA level by RT-qPCR assay. (B) miR-29a reduced the protein level of YY1 in A549 cells. The protein level of YY1 was measured by western blot analysis. (C) Quantification of the protein expression level of YY1 as analyzed by ImageJ software. (D) miR-29a reduced the protein level of N-cadherin in the A549 cells. The protein level of N-cadherin was measured by western blot analysis. (E) Quantification of the protein expression level of N-cadherin as analyzed by ImageJ software. *P

    Article Snippet: Membranes were incubated with primary antibodies: YY1 (cat. no. ab10923; Abcam, Cambridge, MA, USA), AKT (cat. no. C67E7; Cell Signaling Technology, Danvers, MA, USA), pAKT Ser473 (cat. no. 9271; Cell Signaling Technology) and N-cadherin (cat. no. D4R1H; Cell Signaling Technology) (diluted by 1:2,000) and β-actin antibody (cat. no. ZB2305; ZEGB-Bio Ltd., Beijing, China), respectively, at 4°C overnight.

    Techniques: Transfection, Quantitative RT-PCR, Western Blot, Expressing, Software

    A purified YY1 complex methylates histone H4. Anti-Flag immunoprecipitates obtained either from HeLa cells transfected with plasmids expressing Flag–DRBP76 ( A ) or from immunopurification of Flag–YY1 ( B ) were assayed for methylase activity in the presence of core histones. “Flag competitor” corresponds to the addition of excess Flag peptide immunogen. Negative controls include immunoprecipitates from mock transfected cells and anti-Flag immunopurified materials from cells transduced with adenovirus expressing the GFP. Purified recombinant SUV39H1 was used as a positive control for histone H3 methylation. Each blot was stained with Amido black to ensure proper protein transfer.

    Journal: Genes & Development

    Article Title: Targeted recruitment of a histone H4-specific methyltransferase by the transcription factor YY1

    doi: 10.1101/gad.1068003

    Figure Lengend Snippet: A purified YY1 complex methylates histone H4. Anti-Flag immunoprecipitates obtained either from HeLa cells transfected with plasmids expressing Flag–DRBP76 ( A ) or from immunopurification of Flag–YY1 ( B ) were assayed for methylase activity in the presence of core histones. “Flag competitor” corresponds to the addition of excess Flag peptide immunogen. Negative controls include immunoprecipitates from mock transfected cells and anti-Flag immunopurified materials from cells transduced with adenovirus expressing the GFP. Purified recombinant SUV39H1 was used as a positive control for histone H3 methylation. Each blot was stained with Amido black to ensure proper protein transfer.

    Article Snippet: Anti-DRBP76 and anti-YY1 antibodies were obtained from BD Biosciences and Santa Cruz Biotechnology, respectively.

    Techniques: Purification, Transfection, Expressing, Immu-Puri, Activity Assay, Transduction, Recombinant, Positive Control, Methylation, Staining

    A model of gene activation and repression by YY1 through recruitment of histone-modifying enzymes.

    Journal: Genes & Development

    Article Title: Targeted recruitment of a histone H4-specific methyltransferase by the transcription factor YY1

    doi: 10.1101/gad.1068003

    Figure Lengend Snippet: A model of gene activation and repression by YY1 through recruitment of histone-modifying enzymes.

    Article Snippet: Anti-DRBP76 and anti-YY1 antibodies were obtained from BD Biosciences and Santa Cruz Biotechnology, respectively.

    Techniques: Activation Assay

    DRBP76- and PRMT1-dependent activation by YY1. ( A ) Schematic diagram of effector and reporter plasmids. ( B,C ) Expression plasmids and reporter plasmids were transfected together into HeLa cells as indicated. Luciferase activities are the averages ± S.D. from three separate experiments. Western blots were performed to monitor protein expression.

    Journal: Genes & Development

    Article Title: Targeted recruitment of a histone H4-specific methyltransferase by the transcription factor YY1

    doi: 10.1101/gad.1068003

    Figure Lengend Snippet: DRBP76- and PRMT1-dependent activation by YY1. ( A ) Schematic diagram of effector and reporter plasmids. ( B,C ) Expression plasmids and reporter plasmids were transfected together into HeLa cells as indicated. Luciferase activities are the averages ± S.D. from three separate experiments. Western blots were performed to monitor protein expression.

    Article Snippet: Anti-DRBP76 and anti-YY1 antibodies were obtained from BD Biosciences and Santa Cruz Biotechnology, respectively.

    Techniques: Activation Assay, Expressing, Transfection, Luciferase, Western Blot

    Interaction of endogenous YY1 and DRBP76. ( A ) Anti-YY1 and anti-Flag immunoprecipitates from HeLa whole-cell extracts were separated by SDS-PAGE, transferred onto a membrane, and probed with an anti-DRBP76 antibody. “Mock IP” indicates a reaction carried out identically but without the primary antibody. ( B ) HeLa nuclear extracts were fractionated first on a P11 phosphocellulose column, and the active fraction, as monitored by Western blots and EMSAs, was subsequently fractionated on a Q-Sepharose column. Active fractions eluted from the second column were pooled and loaded onto a nickel affinity column. Final active fractions (1–5) were pooled, separated by SDS-PAGE, and analyzed by silver staining and separately by Coomassie blue staining followed by mass spectrometry. For simplicity, the Western blots and EMSA results obtained from the initial two purification steps are not shown here.

    Journal: Genes & Development

    Article Title: Targeted recruitment of a histone H4-specific methyltransferase by the transcription factor YY1

    doi: 10.1101/gad.1068003

    Figure Lengend Snippet: Interaction of endogenous YY1 and DRBP76. ( A ) Anti-YY1 and anti-Flag immunoprecipitates from HeLa whole-cell extracts were separated by SDS-PAGE, transferred onto a membrane, and probed with an anti-DRBP76 antibody. “Mock IP” indicates a reaction carried out identically but without the primary antibody. ( B ) HeLa nuclear extracts were fractionated first on a P11 phosphocellulose column, and the active fraction, as monitored by Western blots and EMSAs, was subsequently fractionated on a Q-Sepharose column. Active fractions eluted from the second column were pooled and loaded onto a nickel affinity column. Final active fractions (1–5) were pooled, separated by SDS-PAGE, and analyzed by silver staining and separately by Coomassie blue staining followed by mass spectrometry. For simplicity, the Western blots and EMSA results obtained from the initial two purification steps are not shown here.

    Article Snippet: Anti-DRBP76 and anti-YY1 antibodies were obtained from BD Biosciences and Santa Cruz Biotechnology, respectively.

    Techniques: SDS Page, Western Blot, Affinity Column, Silver Staining, Staining, Mass Spectrometry, Purification

    Mapping of the DRBP76-binding domain in YY1. Schematic diagram of full-length or various truncated GST–YY1 fusion proteins ( top panel). For simplicity, the GST portions of the fusion proteins are not shown. The ability of each GST–YY1 fusion protein to bind DRBP76 is indicated (+ or −). The shaded bar represents the deduced DRBP76-binding domain. Autoradiographs of in vitro translated DRBP76 protein captured by GST–YY1 fusion proteins are shown in the bottom panel. The input lane was loaded with one-tenth the amount of 35 S-labeled proteins used in the binding reactions. The gel was stained with Coomassie blue prior to autoradiography to show approximately equal amounts of GST fusion proteins in each lane.

    Journal: Genes & Development

    Article Title: Targeted recruitment of a histone H4-specific methyltransferase by the transcription factor YY1

    doi: 10.1101/gad.1068003

    Figure Lengend Snippet: Mapping of the DRBP76-binding domain in YY1. Schematic diagram of full-length or various truncated GST–YY1 fusion proteins ( top panel). For simplicity, the GST portions of the fusion proteins are not shown. The ability of each GST–YY1 fusion protein to bind DRBP76 is indicated (+ or −). The shaded bar represents the deduced DRBP76-binding domain. Autoradiographs of in vitro translated DRBP76 protein captured by GST–YY1 fusion proteins are shown in the bottom panel. The input lane was loaded with one-tenth the amount of 35 S-labeled proteins used in the binding reactions. The gel was stained with Coomassie blue prior to autoradiography to show approximately equal amounts of GST fusion proteins in each lane.

    Article Snippet: Anti-DRBP76 and anti-YY1 antibodies were obtained from BD Biosciences and Santa Cruz Biotechnology, respectively.

    Techniques: Binding Assay, In Vitro, Labeling, Staining, Autoradiography

    YY1 binds to PRMT1. HeLa cells were either transfected with plasmids encoding the indicated proteins ( A,B ) or transduced with recombinant adenovirus expressing Flag–YY1, Flag, or GFP ( C,D ). Cell extracts were immunoprecipitated with an anti-Flag antibody ( A – C ) or purified through an anti-Flag affinity column ( D ) and immunoblotted with an anti-PRMT1 antibody. “Mock IP” indicates reactions carried out identically but without the anti-Flag antibody. “Flag competitor” corresponds to the addition of excess Flag peptide immunogen.

    Journal: Genes & Development

    Article Title: Targeted recruitment of a histone H4-specific methyltransferase by the transcription factor YY1

    doi: 10.1101/gad.1068003

    Figure Lengend Snippet: YY1 binds to PRMT1. HeLa cells were either transfected with plasmids encoding the indicated proteins ( A,B ) or transduced with recombinant adenovirus expressing Flag–YY1, Flag, or GFP ( C,D ). Cell extracts were immunoprecipitated with an anti-Flag antibody ( A – C ) or purified through an anti-Flag affinity column ( D ) and immunoblotted with an anti-PRMT1 antibody. “Mock IP” indicates reactions carried out identically but without the anti-Flag antibody. “Flag competitor” corresponds to the addition of excess Flag peptide immunogen.

    Article Snippet: Anti-DRBP76 and anti-YY1 antibodies were obtained from BD Biosciences and Santa Cruz Biotechnology, respectively.

    Techniques: Transfection, Transduction, Recombinant, Expressing, Immunoprecipitation, Purification, Affinity Column

    YY1-dependent recruitment of PRMT1. ( A,B ) Cross-linked chromatin from NIH3T3 cells nontransduced or transduced with YY1- or GFP-expressing adenovirus was immunoprecipitated with the indicated antibodies and analyzed by PCR using primers specific for DNA surrounding YY1 sites in the c- myc and c- fos promoters. Identical results were obtained from multiple experiments. ( C ) Semiquantitative RT–PCR was performed to analyze expression of the c- myc gene in YY1-expressing cells. PCR products of cDNA samples are shown.

    Journal: Genes & Development

    Article Title: Targeted recruitment of a histone H4-specific methyltransferase by the transcription factor YY1

    doi: 10.1101/gad.1068003

    Figure Lengend Snippet: YY1-dependent recruitment of PRMT1. ( A,B ) Cross-linked chromatin from NIH3T3 cells nontransduced or transduced with YY1- or GFP-expressing adenovirus was immunoprecipitated with the indicated antibodies and analyzed by PCR using primers specific for DNA surrounding YY1 sites in the c- myc and c- fos promoters. Identical results were obtained from multiple experiments. ( C ) Semiquantitative RT–PCR was performed to analyze expression of the c- myc gene in YY1-expressing cells. PCR products of cDNA samples are shown.

    Article Snippet: Anti-DRBP76 and anti-YY1 antibodies were obtained from BD Biosciences and Santa Cruz Biotechnology, respectively.

    Techniques: Transduction, Expressing, Immunoprecipitation, Polymerase Chain Reaction, Reverse Transcription Polymerase Chain Reaction

    YY1 physically associates with DRBP76. ( A ) Silver-stained SDS-PAGE of the Flag–YY1 complex. “Control” indicates an anti-Flag immunopurified sample prepared from HeLa cells transduced with adenovirus expressing GFP. In addition to Flag–YY1, arrows indicate proteins copurified, in approximately stoichiometrically equivalent amounts, with Flag–YY1. ( B ) Amino acid sequence of DRBP76 and ILF3. Each dash (–) indicates an amino acid in ILF3 that is identical to the corresponding one in DRBP76. Peptide sequences obtained by microsequencing are underlined. ( C ) An immunoblot of the purified Flag–YY1 complex using an anti-DRBP76 antibody.

    Journal: Genes & Development

    Article Title: Targeted recruitment of a histone H4-specific methyltransferase by the transcription factor YY1

    doi: 10.1101/gad.1068003

    Figure Lengend Snippet: YY1 physically associates with DRBP76. ( A ) Silver-stained SDS-PAGE of the Flag–YY1 complex. “Control” indicates an anti-Flag immunopurified sample prepared from HeLa cells transduced with adenovirus expressing GFP. In addition to Flag–YY1, arrows indicate proteins copurified, in approximately stoichiometrically equivalent amounts, with Flag–YY1. ( B ) Amino acid sequence of DRBP76 and ILF3. Each dash (–) indicates an amino acid in ILF3 that is identical to the corresponding one in DRBP76. Peptide sequences obtained by microsequencing are underlined. ( C ) An immunoblot of the purified Flag–YY1 complex using an anti-DRBP76 antibody.

    Article Snippet: Anti-DRBP76 and anti-YY1 antibodies were obtained from BD Biosciences and Santa Cruz Biotechnology, respectively.

    Techniques: Staining, SDS Page, Transduction, Expressing, Sequencing, Purification