Structured Review

Cell Signaling Technology Inc anti sp1
HDAC1 and HDAC2 promoter activity is regulated by <t>Sp1</t> and Sp3 in colon cancer cell lines. A ) Analysis of HDAC1 and HDAC2 mRNA expression by qRT-PCR in HCT116, HT29, and FHs 74 Int cells. HDAC1 and HDAC2 mRNA expression is shown relative to the control
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1) Product Images from "Overexpression of histone deacetylases in cancer cells is controlled by interplay of transcription factors and epigenetic modulators"

Article Title: Overexpression of histone deacetylases in cancer cells is controlled by interplay of transcription factors and epigenetic modulators

Journal:

doi: 10.1096/fj.14-250654

HDAC1 and HDAC2 promoter activity is regulated by Sp1 and Sp3 in colon cancer cell lines. A ) Analysis of HDAC1 and HDAC2 mRNA expression by qRT-PCR in HCT116, HT29, and FHs 74 Int cells. HDAC1 and HDAC2 mRNA expression is shown relative to the control
Figure Legend Snippet: HDAC1 and HDAC2 promoter activity is regulated by Sp1 and Sp3 in colon cancer cell lines. A ) Analysis of HDAC1 and HDAC2 mRNA expression by qRT-PCR in HCT116, HT29, and FHs 74 Int cells. HDAC1 and HDAC2 mRNA expression is shown relative to the control

Techniques Used: Activity Assay, Expressing, Quantitative RT-PCR

Sp1 or Sp3 silencing affects cell growth and colony formation in colon cancer cells. A ) Effect of Sp1 or Sp3 silencing on cell proliferation. HCT116 cells (1×10 4 ) with or without Sp1 or Sp3 KD were seeded and the cell numbers were counted every
Figure Legend Snippet: Sp1 or Sp3 silencing affects cell growth and colony formation in colon cancer cells. A ) Effect of Sp1 or Sp3 silencing on cell proliferation. HCT116 cells (1×10 4 ) with or without Sp1 or Sp3 KD were seeded and the cell numbers were counted every

Techniques Used:

Effect of Sp1 on recruitment of other coregulators on HDAC1 promoter. A ) Levels of p300 or other indicated proteins in control or Sp1 KD cells were determined by Western blot. NC, negative control of scramble shRNA. β-Actin was used as a loading
Figure Legend Snippet: Effect of Sp1 on recruitment of other coregulators on HDAC1 promoter. A ) Levels of p300 or other indicated proteins in control or Sp1 KD cells were determined by Western blot. NC, negative control of scramble shRNA. β-Actin was used as a loading

Techniques Used: Western Blot, Negative Control, shRNA

Sp1 and Sp3 affect HDAC1 and HDAC2 expression in colorectal cancer cell lines. A ) Levels of HDAC1 and other indicated proteins in control and Sp1 or Sp3 KD cells were determined by Western blot. NC, negative control of scramble shRNA. β-Actin
Figure Legend Snippet: Sp1 and Sp3 affect HDAC1 and HDAC2 expression in colorectal cancer cell lines. A ) Levels of HDAC1 and other indicated proteins in control and Sp1 or Sp3 KD cells were determined by Western blot. NC, negative control of scramble shRNA. β-Actin

Techniques Used: Expressing, Western Blot, Negative Control, shRNA

Sp1 and Sp3 are overexpressed and bind to HDAC1 and HDAC2 promoters in colon cancer. A ) Levels of Sp1 and Sp3 proteins from various colon cancer cell lines were determined by Western blotting. Vaco250 and Vaco330 are human adenoma cell lines and used
Figure Legend Snippet: Sp1 and Sp3 are overexpressed and bind to HDAC1 and HDAC2 promoters in colon cancer. A ) Levels of Sp1 and Sp3 proteins from various colon cancer cell lines were determined by Western blotting. Vaco250 and Vaco330 are human adenoma cell lines and used

Techniques Used: Western Blot

Effect of p300 KD on epigenetic markers and Sp1 binding on HDAC1 promoter. A ) Levels of HDAC1 and other indicated proteins in control or p300 KD cells were determined by Western blot. NC, negative control of scramble shRNA. β-Actin was used as
Figure Legend Snippet: Effect of p300 KD on epigenetic markers and Sp1 binding on HDAC1 promoter. A ) Levels of HDAC1 and other indicated proteins in control or p300 KD cells were determined by Western blot. NC, negative control of scramble shRNA. β-Actin was used as

Techniques Used: Binding Assay, Western Blot, Negative Control, shRNA

Effect of Sp1 on recruitment of other regulators on the HDAC2 promoter. ChIP assays were carried out in HCT116 cells with Sp1 KD by using indicated antibodies: ac-H3 ( A ), H3K4 trimethylation ( B ), p300 ( C ), and SET1 ( D ). The resulting DNA was subjected
Figure Legend Snippet: Effect of Sp1 on recruitment of other regulators on the HDAC2 promoter. ChIP assays were carried out in HCT116 cells with Sp1 KD by using indicated antibodies: ac-H3 ( A ), H3K4 trimethylation ( B ), p300 ( C ), and SET1 ( D ). The resulting DNA was subjected

Techniques Used: Chromatin Immunoprecipitation

2) Product Images from "Baicalin induces apoptosis in SW480 cells through downregulation of the SP1 transcription factor"

Article Title: Baicalin induces apoptosis in SW480 cells through downregulation of the SP1 transcription factor

Journal: Anti-Cancer Drugs

doi: 10.1097/CAD.0000000000000708

Baicalin inhibits SW480 cells growth and decreases the expression of sp1 in SW480 human colon cancer cells. Cells were treated with DMSO or 50–400 µg/ml baicalin for 24 and 48 h. (a) The photograph of inhibition effects of baicalin on the growth of SW480 cells. (b) Inhibition rate of baicalin, cell proliferation was measured using the CCK-8 kit, as described in ‘Materials and methods’ section. (c) Baicalin decreased the protein expression of sp1 in SW480 human colon cancer cells. Each bar represents the mean±SD of three independent experiments. P
Figure Legend Snippet: Baicalin inhibits SW480 cells growth and decreases the expression of sp1 in SW480 human colon cancer cells. Cells were treated with DMSO or 50–400 µg/ml baicalin for 24 and 48 h. (a) The photograph of inhibition effects of baicalin on the growth of SW480 cells. (b) Inhibition rate of baicalin, cell proliferation was measured using the CCK-8 kit, as described in ‘Materials and methods’ section. (c) Baicalin decreased the protein expression of sp1 in SW480 human colon cancer cells. Each bar represents the mean±SD of three independent experiments. P

Techniques Used: Expressing, Inhibition, CCK-8 Assay

sp1 as a determinant transcription factor of colorectal cancer during the analysis of four datasets. (a) GSE4107 gene expression according to; (b) GSE24514 gene expression according to; (c) GSE32323 gene expression according to; (b) GSE73883 gene expression according to. P
Figure Legend Snippet: sp1 as a determinant transcription factor of colorectal cancer during the analysis of four datasets. (a) GSE4107 gene expression according to; (b) GSE24514 gene expression according to; (c) GSE32323 gene expression according to; (b) GSE73883 gene expression according to. P

Techniques Used: Expressing

(a, b) sp1 inhibitor mithramycin-A inhibits SW480 cells growth and decreases the expression of sp1 , (c, d) mithramycin-A induces SW480 human colon cancer cell apoptosis. Each bar represents the mean±SD of three independent experiments. P
Figure Legend Snippet: (a, b) sp1 inhibitor mithramycin-A inhibits SW480 cells growth and decreases the expression of sp1 , (c, d) mithramycin-A induces SW480 human colon cancer cell apoptosis. Each bar represents the mean±SD of three independent experiments. P

Techniques Used: Expressing

3) Product Images from "SP1 and RARα regulate AGAP2 expression in cancer"

Article Title: SP1 and RARα regulate AGAP2 expression in cancer

Journal: Scientific Reports

doi: 10.1038/s41598-018-36888-x

SP1 role in AGAP2 expression. The −246 to + 36 region of AGAP2 was analysed for SP1 binding sites with the online JASPAR prediction tool. ( a ) Representation of the SP1 binding sites location within the AGAP2 −246/+36 DNA fragment. The actual logo used for the searches is shown in ( b ) and the scores for binding are provided as a table in ( c ). ( d ) Effect of mithramycin (MTR) treatment on luciferase activity. KU812 and DU145 were transfected with the AGAP2 −246/+36 luc and the β-galactosidase plasmids and the reporter activity was measured after 24 h in the presence of 200 nM MTR or just vehicle (Vhc). Represented in the graph are the mean ± SD of data from at least three independent experiments performed in triplicates (n = 9). Values were calculated as the ratio of the luciferase activity values over the respective β-galactosidase activity and made relative to the values obtained for Vhc treated cells (control). Treatment differences were analysed with Mann-Whitney U tests (* P
Figure Legend Snippet: SP1 role in AGAP2 expression. The −246 to + 36 region of AGAP2 was analysed for SP1 binding sites with the online JASPAR prediction tool. ( a ) Representation of the SP1 binding sites location within the AGAP2 −246/+36 DNA fragment. The actual logo used for the searches is shown in ( b ) and the scores for binding are provided as a table in ( c ). ( d ) Effect of mithramycin (MTR) treatment on luciferase activity. KU812 and DU145 were transfected with the AGAP2 −246/+36 luc and the β-galactosidase plasmids and the reporter activity was measured after 24 h in the presence of 200 nM MTR or just vehicle (Vhc). Represented in the graph are the mean ± SD of data from at least three independent experiments performed in triplicates (n = 9). Values were calculated as the ratio of the luciferase activity values over the respective β-galactosidase activity and made relative to the values obtained for Vhc treated cells (control). Treatment differences were analysed with Mann-Whitney U tests (* P

Techniques Used: Expressing, Binding Assay, Luciferase, Activity Assay, Transfection, MANN-WHITNEY

Mechanism of ATRA-mediated AGAP2 transcription. ( a ) Sheared chromatin of DU145 cells grown under AGAP2 expression conditions was used for immunoprecipitation using 2 μg of antibody (a rabbit IgG as negative control, a rabbit antibody against RNApol II as positive control, a rabbit anti-RARα, a rabbit anti-RXRα or a rabbit anti-PCAF antibody) and optimised primers were used to amplify a region specific to the AGAP2 promoter (see Supplementary Table 1 ). Data are represented as fold enrichment: fold enrich in signal relative to the IgG background signal. ( b ) Diagram representing the proposed mechanism of ATRA-mediated activation of AGAP2 transcription. In the presence of ATRA (lower panel), the heterodimer RARα/RXRα would recruit the lysine acetyl transferase PCAF to activate transcription and the recruitment of SP1 would also be enhanced.
Figure Legend Snippet: Mechanism of ATRA-mediated AGAP2 transcription. ( a ) Sheared chromatin of DU145 cells grown under AGAP2 expression conditions was used for immunoprecipitation using 2 μg of antibody (a rabbit IgG as negative control, a rabbit antibody against RNApol II as positive control, a rabbit anti-RARα, a rabbit anti-RXRα or a rabbit anti-PCAF antibody) and optimised primers were used to amplify a region specific to the AGAP2 promoter (see Supplementary Table 1 ). Data are represented as fold enrichment: fold enrich in signal relative to the IgG background signal. ( b ) Diagram representing the proposed mechanism of ATRA-mediated activation of AGAP2 transcription. In the presence of ATRA (lower panel), the heterodimer RARα/RXRα would recruit the lysine acetyl transferase PCAF to activate transcription and the recruitment of SP1 would also be enhanced.

Techniques Used: Expressing, Immunoprecipitation, Negative Control, Positive Control, Activation Assay

4) Product Images from "Long noncoding RNA UCA1 induced by SP1 promotes cell proliferation via recruiting EZH2 and activating AKT pathway in gastric cancer"

Article Title: Long noncoding RNA UCA1 induced by SP1 promotes cell proliferation via recruiting EZH2 and activating AKT pathway in gastric cancer

Journal: Cell Death & Disease

doi: 10.1038/cddis.2017.143

The transcription factor SP1 is involved in UCA1 upregulation. ( a ) The predicted top three positions of putative SP1-binding sites in −2.5 kb human UCA1 promoter from JASPAR database. ( b ) Upregulation of SP1 and slicing of SP1 by transfecting pcDNA-SP1 and SP1 siRNAs, respectively, which were determined by western blot in the protein expression level. ( c ) RT-PCR was used to determine the expression of UCA1 in cells transfected with pcDNA-SP1 and SP1 siRNAs, respectively. Error bars represent the mean±S.D. of triplicate experiments (* P
Figure Legend Snippet: The transcription factor SP1 is involved in UCA1 upregulation. ( a ) The predicted top three positions of putative SP1-binding sites in −2.5 kb human UCA1 promoter from JASPAR database. ( b ) Upregulation of SP1 and slicing of SP1 by transfecting pcDNA-SP1 and SP1 siRNAs, respectively, which were determined by western blot in the protein expression level. ( c ) RT-PCR was used to determine the expression of UCA1 in cells transfected with pcDNA-SP1 and SP1 siRNAs, respectively. Error bars represent the mean±S.D. of triplicate experiments (* P

Techniques Used: Binding Assay, Western Blot, Expressing, Reverse Transcription Polymerase Chain Reaction, Transfection

Schematic diagram illustrating signaling of EZH2 and its upstream activator and its downstream effectors in GC. SP1 directly binds to the core promoter of UCA1 so that it activates the expression of UCA1. UCA1 directly interacts with EZH2 and increase the expression of EZH2, which affects the activation of AKT/GSK-3B/cyclin D1 axis. EZH2 could also physically interact with the cyclin D1 promoter to promote the expression of cyclin D1. In addition, P-AKT could indirectly influence the expression of EZH2 so as to form a positive feedback loop with EZH2. With the accumulation of cyclin D1, G1/S transition was activated to promote cell proliferation and cancer progression
Figure Legend Snippet: Schematic diagram illustrating signaling of EZH2 and its upstream activator and its downstream effectors in GC. SP1 directly binds to the core promoter of UCA1 so that it activates the expression of UCA1. UCA1 directly interacts with EZH2 and increase the expression of EZH2, which affects the activation of AKT/GSK-3B/cyclin D1 axis. EZH2 could also physically interact with the cyclin D1 promoter to promote the expression of cyclin D1. In addition, P-AKT could indirectly influence the expression of EZH2 so as to form a positive feedback loop with EZH2. With the accumulation of cyclin D1, G1/S transition was activated to promote cell proliferation and cancer progression

Techniques Used: Gas Chromatography, Expressing, Activation Assay

5) Product Images from "MicroRNA 373 Facilitates the Replication of Porcine Reproductive and Respiratory Syndrome Virus by Its Negative Regulation of Type I Interferon Induction"

Article Title: MicroRNA 373 Facilitates the Replication of Porcine Reproductive and Respiratory Syndrome Virus by Its Negative Regulation of Type I Interferon Induction

Journal:

doi: 10.1128/JVI.01311-16

PRRSV infection upregulated miR-373 expression in MARC-145 cells. (A) MARC-145 cells were infected with PRRSV at an MOI of 1 for the indicated times, and the levels of miR-373 expression were measured by qRT-PCR. (B) MARC-145 cells were infected with PRRSV at different MOIs for 24 h, and then the levels of miR-373 expression were measured by qRT-PCR. (C) MARC-145 cells were transfected with pGL-miR-373, phRL-TK, or pGL4.17, and 24 h later the cells were infected with PRRSV at different MOIs. Forty-eight hours later, the cells were subjected to dual-luciferase assays. (D) 293T cells were cotransfected with different truncated miR-373 promoter report plasmids and phRL-TK, and 48 h later the cells were harvested for dual-luciferase assays. (E) miR-373 promoter sequence of Macaca mulatta and human harbored one conserved putative GR binding site and three highly conserved putative Sp1 binding site. 293T cells were cotransfected with the indicated report plasmids and phRL-TK, and 48 h later the cells were harvested for dual-luciferase assays. (Left) Schematic representation of mutation constructs of the Macaca mulatta miR-373 promoter. (Right) Results of dual-luciferase assays. (F) MARC-145 cells were cotransfected with phRL-TK, pGL-miR-373, pcDNA-3.1-Flag (NC), Sp1-Flag (800 ng), siRNA control (SC), different concentrations of si-Sp1, or different doses of Mith, and 48 h later the miR-373 promoter activity was analyzed by dual-luciferase assays and the expression levels of pri-miR-373 and miR-373 were detected by qRT-PCR. Additionally, the expression levels of Sp1 were detected by qRT-PCR and Western blotting of the corresponding group. (G) EMSA was performed as described in Materials and Methods. Biotin-labeled 40-bp probes, which included the putative Sp1 binding site (underlined) of the Macaca mulatta miRNA-373 promoter, were used. Lane 1 shows labeled probes alone without nuclear extracts (N.E.) from MARC-145 cells, while lane 2 and lane 4 show labeled wild-type or mutant probes with N.E. Competition assays were conducted by adding an excess (200-fold) of unlabeled wild-type or mutant consensus sequence (lanes 3 and 5). (H) ChIP assays in MARC-145 cells were performed with anti-Sp1 antibody or anti-IgG isotype control antibody. The input DNA and immunoprecipitated DNA then were purified and analyzed by qRT-PCR and PCR using primers specific for the miR-373 promoter. (I) MARC-145 cells were infected with PRRSV at an MOI of 1 or mock infected for 24 h, and the expression levels of Sp1 were determined by qRT-PCR and Western blotting. (J) MARC-145 cells were cotransfected with phRL-TK, pGL-miR-373, pcDNA-3.1-Flag (NC), siRNA control (SC), different concentrations of Sp1-Flag, different concentrations of si-Sp1, or different doses of Mith, and 24 h later the cells were infected with PRRSV at an MOI of 0.1. Forty-eight hours later, miR-373 promoter activity was analyzed by dual-luciferase assays. (K and L) MARC-145 cells were transfected with pcDNA-3.1-Flag (NC), siRNA control (SC), different concentrations of Sp1-Flag, different concentrations of si-Sp1, or different doses of Mith, and 24 h later the cells were infected with PRRSV at an MOI of 0.1. Forty-eight hours later the expression levels of pri-miR-373 (K) and miR-373 (L) were detected by qRT-PCR. Results are expressed as means ± SD from three independent experiments. P values were calculated using Student's t test. An asterisk indicates a comparison with the indicated control. *, P < 0.05; **, P < 0.01.
Figure Legend Snippet: PRRSV infection upregulated miR-373 expression in MARC-145 cells. (A) MARC-145 cells were infected with PRRSV at an MOI of 1 for the indicated times, and the levels of miR-373 expression were measured by qRT-PCR. (B) MARC-145 cells were infected with PRRSV at different MOIs for 24 h, and then the levels of miR-373 expression were measured by qRT-PCR. (C) MARC-145 cells were transfected with pGL-miR-373, phRL-TK, or pGL4.17, and 24 h later the cells were infected with PRRSV at different MOIs. Forty-eight hours later, the cells were subjected to dual-luciferase assays. (D) 293T cells were cotransfected with different truncated miR-373 promoter report plasmids and phRL-TK, and 48 h later the cells were harvested for dual-luciferase assays. (E) miR-373 promoter sequence of Macaca mulatta and human harbored one conserved putative GR binding site and three highly conserved putative Sp1 binding site. 293T cells were cotransfected with the indicated report plasmids and phRL-TK, and 48 h later the cells were harvested for dual-luciferase assays. (Left) Schematic representation of mutation constructs of the Macaca mulatta miR-373 promoter. (Right) Results of dual-luciferase assays. (F) MARC-145 cells were cotransfected with phRL-TK, pGL-miR-373, pcDNA-3.1-Flag (NC), Sp1-Flag (800 ng), siRNA control (SC), different concentrations of si-Sp1, or different doses of Mith, and 48 h later the miR-373 promoter activity was analyzed by dual-luciferase assays and the expression levels of pri-miR-373 and miR-373 were detected by qRT-PCR. Additionally, the expression levels of Sp1 were detected by qRT-PCR and Western blotting of the corresponding group. (G) EMSA was performed as described in Materials and Methods. Biotin-labeled 40-bp probes, which included the putative Sp1 binding site (underlined) of the Macaca mulatta miRNA-373 promoter, were used. Lane 1 shows labeled probes alone without nuclear extracts (N.E.) from MARC-145 cells, while lane 2 and lane 4 show labeled wild-type or mutant probes with N.E. Competition assays were conducted by adding an excess (200-fold) of unlabeled wild-type or mutant consensus sequence (lanes 3 and 5). (H) ChIP assays in MARC-145 cells were performed with anti-Sp1 antibody or anti-IgG isotype control antibody. The input DNA and immunoprecipitated DNA then were purified and analyzed by qRT-PCR and PCR using primers specific for the miR-373 promoter. (I) MARC-145 cells were infected with PRRSV at an MOI of 1 or mock infected for 24 h, and the expression levels of Sp1 were determined by qRT-PCR and Western blotting. (J) MARC-145 cells were cotransfected with phRL-TK, pGL-miR-373, pcDNA-3.1-Flag (NC), siRNA control (SC), different concentrations of Sp1-Flag, different concentrations of si-Sp1, or different doses of Mith, and 24 h later the cells were infected with PRRSV at an MOI of 0.1. Forty-eight hours later, miR-373 promoter activity was analyzed by dual-luciferase assays. (K and L) MARC-145 cells were transfected with pcDNA-3.1-Flag (NC), siRNA control (SC), different concentrations of Sp1-Flag, different concentrations of si-Sp1, or different doses of Mith, and 24 h later the cells were infected with PRRSV at an MOI of 0.1. Forty-eight hours later the expression levels of pri-miR-373 (K) and miR-373 (L) were detected by qRT-PCR. Results are expressed as means ± SD from three independent experiments. P values were calculated using Student's t test. An asterisk indicates a comparison with the indicated control. *, P < 0.05; **, P < 0.01.

Techniques Used: Infection, Expressing, Quantitative RT-PCR, Transfection, Luciferase, Sequencing, Binding Assay, Mutagenesis, Construct, Activity Assay, Western Blot, Labeling, Chromatin Immunoprecipitation, Immunoprecipitation, Purification, Polymerase Chain Reaction

Model for the mechanisms by which PRRSV infection upregulated miR-373 expression and miR-373 facilitated PRRSV replication. PRRSV, nsp9, and N upregulated miR-373 expression, since they elevated the expression of Sp1, which was an important transcriptional factor for miR-373 expression. miR-373 enhanced PRRSV replication by its negative regulation of type I interferon induction and the type I interferon receptor signaling pathway, since miR-373 targeted NFIA, NFIB, IRAK1, IRAK4, IRF1, IFR9, IFNAR1, and IFNAR2.
Figure Legend Snippet: Model for the mechanisms by which PRRSV infection upregulated miR-373 expression and miR-373 facilitated PRRSV replication. PRRSV, nsp9, and N upregulated miR-373 expression, since they elevated the expression of Sp1, which was an important transcriptional factor for miR-373 expression. miR-373 enhanced PRRSV replication by its negative regulation of type I interferon induction and the type I interferon receptor signaling pathway, since miR-373 targeted NFIA, NFIB, IRAK1, IRAK4, IRF1, IFR9, IFNAR1, and IFNAR2.

Techniques Used: Infection, Expressing

6) Product Images from "SP1-induced lncRNA-ZFAS1 contributes to colorectal cancer progression via the miR-150-5p/VEGFA axis"

Article Title: SP1-induced lncRNA-ZFAS1 contributes to colorectal cancer progression via the miR-150-5p/VEGFA axis

Journal: Cell Death & Disease

doi: 10.1038/s41419-018-0962-6

The transcription factor SP1 is involved in ZFAS1 upregulation. a The predicted positions of puative SP1 binding motif in −2500 bp human ZFAS1 promoter. b Quantitative ChIP assays were performed to show direct binding of SP1 to endogenous ZFAS1 promoter regions. The primers designed for ChIP were provided in supplementary materials and methods. c A luciferase reporter assay was used by cotransfecting the full ZFAS1 promoter (ZFAS1-pGL3-F) or deleted ZFAS1 promoter fragment E2 (ZFAS1-pGL3-D) with SP1 expression plasmid or blank vector in 293T cells. Luciferase activities were expressed as relative to that of the pGL3 vector. d qPCR analysis of ZFAS1 expression levels following the treatment of siSP1-1, siSP1-2 in HCT116 and HCT8 cells. Data were shown as mean ± SD of three independent experiments. ** P
Figure Legend Snippet: The transcription factor SP1 is involved in ZFAS1 upregulation. a The predicted positions of puative SP1 binding motif in −2500 bp human ZFAS1 promoter. b Quantitative ChIP assays were performed to show direct binding of SP1 to endogenous ZFAS1 promoter regions. The primers designed for ChIP were provided in supplementary materials and methods. c A luciferase reporter assay was used by cotransfecting the full ZFAS1 promoter (ZFAS1-pGL3-F) or deleted ZFAS1 promoter fragment E2 (ZFAS1-pGL3-D) with SP1 expression plasmid or blank vector in 293T cells. Luciferase activities were expressed as relative to that of the pGL3 vector. d qPCR analysis of ZFAS1 expression levels following the treatment of siSP1-1, siSP1-2 in HCT116 and HCT8 cells. Data were shown as mean ± SD of three independent experiments. ** P

Techniques Used: Binding Assay, Chromatin Immunoprecipitation, Luciferase, Reporter Assay, Expressing, Plasmid Preparation, Real-time Polymerase Chain Reaction

7) Product Images from "miR-506 attenuates methylation of lncRNA MEG3 to inhibit migration and invasion of breast cancer cell lines via targeting SP1 and SP3"

Article Title: miR-506 attenuates methylation of lncRNA MEG3 to inhibit migration and invasion of breast cancer cell lines via targeting SP1 and SP3

Journal: Cancer Cell International

doi: 10.1186/s12935-018-0642-8

miR-506 overexpression downregulates SP1, SP3, DNMT1 and upregulates MEG3 expression. a RT-qPCR analysis showing the level of miR-506 in control (wild-type), mimic NC- or miR-506 mimic transfected MCF-7 cells. b Western blot analysis showing the protein level of DNMT1 and SP3 in control (wild-type), mimic NC- or miR-506 mimic-overexpressed MCF-7 cells. β-actin acts as internal control. c RT-qPCR analysis showing the mRNA level of DNMT1 and SP3 in control (wild-type), mimic NC- or miR-506 mimic-overexpressed MCF-7 cells. GAPDH acts as internal control. d Bioinformatic prediction of binding site at 3′-UTR of SP1 by miR-506. e Luciferase reporter assay showing miR-506 binds wild-type 3′-UTR of SP1, not mutant 3′-UTR of SP1. The relative luciferase activity was measured and the data were presented as mean ± SD. f Bioinformatic prediction of binding site at 3′-UTR of SP3 by miR-506. g Luciferase reporter assay showing miR-506 binds wild-type 3′-UTR of SP3, not mutant 3′-UTR of SP3. The relative luciferase activity was measured and the data were presented as mean ± SD. h RT-qPCR analysis showing the level of MEG3 in control (wild-type), mimic NC- or miR-506 mimic-overexpressed MCF-7 cells. GAPDH acts as internal control. i Methylation-specific PCR (MSP) analysis showing the methylation level of MEG3 promoter in wild-type (control), mimic NC- or miR-506 mimic-overexpressed MCF-7 cells. GAPDH acts as negative control. j Methylation-specific PCR (MSP) analysis showing the methylation level of MEG3 promoter in wild-type (control), mimic NC- or miR-506 mimic-overexpressed MDA-MB-231 cells. GAPDH acts as negative control. All data were presented as mean ± SD from three biological replicates (*P
Figure Legend Snippet: miR-506 overexpression downregulates SP1, SP3, DNMT1 and upregulates MEG3 expression. a RT-qPCR analysis showing the level of miR-506 in control (wild-type), mimic NC- or miR-506 mimic transfected MCF-7 cells. b Western blot analysis showing the protein level of DNMT1 and SP3 in control (wild-type), mimic NC- or miR-506 mimic-overexpressed MCF-7 cells. β-actin acts as internal control. c RT-qPCR analysis showing the mRNA level of DNMT1 and SP3 in control (wild-type), mimic NC- or miR-506 mimic-overexpressed MCF-7 cells. GAPDH acts as internal control. d Bioinformatic prediction of binding site at 3′-UTR of SP1 by miR-506. e Luciferase reporter assay showing miR-506 binds wild-type 3′-UTR of SP1, not mutant 3′-UTR of SP1. The relative luciferase activity was measured and the data were presented as mean ± SD. f Bioinformatic prediction of binding site at 3′-UTR of SP3 by miR-506. g Luciferase reporter assay showing miR-506 binds wild-type 3′-UTR of SP3, not mutant 3′-UTR of SP3. The relative luciferase activity was measured and the data were presented as mean ± SD. h RT-qPCR analysis showing the level of MEG3 in control (wild-type), mimic NC- or miR-506 mimic-overexpressed MCF-7 cells. GAPDH acts as internal control. i Methylation-specific PCR (MSP) analysis showing the methylation level of MEG3 promoter in wild-type (control), mimic NC- or miR-506 mimic-overexpressed MCF-7 cells. GAPDH acts as negative control. j Methylation-specific PCR (MSP) analysis showing the methylation level of MEG3 promoter in wild-type (control), mimic NC- or miR-506 mimic-overexpressed MDA-MB-231 cells. GAPDH acts as negative control. All data were presented as mean ± SD from three biological replicates (*P

Techniques Used: Over Expression, Expressing, Quantitative RT-PCR, Transfection, Western Blot, Binding Assay, Luciferase, Reporter Assay, Mutagenesis, Activity Assay, Methylation, Polymerase Chain Reaction, Negative Control, Multiple Displacement Amplification

8) Product Images from "Adeno-associated virus-mediated brain delivery of 5-lipoxygenase modulates the AD-like phenotype of APP mice"

Article Title: Adeno-associated virus-mediated brain delivery of 5-lipoxygenase modulates the AD-like phenotype of APP mice

Journal: Molecular Neurodegeneration

doi: 10.1186/1750-1326-7-1

AAV1/2-5LO over-expression modulates CREB levels and transcription of the γ-secretase complex in the brains of Tg2576 mice . A . Levels of total CREB and its phosphorylated form at Ser133 and Sp1 in the cortex of Tg2576 receiving AAV1/2-5LO or empty vector (CTL) assayed by western blot analyses. B . Densitometric analyses of the immunoreactivities to the antibodies shown in the previous panel. C . Relative mRNA levels for PS1, Nicastrin, APH-1 and Pen-2 in the cortex of Tg2576 mice receiving AAV1/2-5LO or empty vector (CTL), as determined by real-time quantitative RT-PCR amplification. Values represent mean ± SEM (*p
Figure Legend Snippet: AAV1/2-5LO over-expression modulates CREB levels and transcription of the γ-secretase complex in the brains of Tg2576 mice . A . Levels of total CREB and its phosphorylated form at Ser133 and Sp1 in the cortex of Tg2576 receiving AAV1/2-5LO or empty vector (CTL) assayed by western blot analyses. B . Densitometric analyses of the immunoreactivities to the antibodies shown in the previous panel. C . Relative mRNA levels for PS1, Nicastrin, APH-1 and Pen-2 in the cortex of Tg2576 mice receiving AAV1/2-5LO or empty vector (CTL), as determined by real-time quantitative RT-PCR amplification. Values represent mean ± SEM (*p

Techniques Used: Over Expression, Mouse Assay, Plasmid Preparation, CTL Assay, Western Blot, Quantitative RT-PCR, Amplification

9) Product Images from "ETS1 and SP1 drive DHX15 expression in acute lymphoblastic leukaemia, et al. ETS1 and SP1 drive DHX15 expression in acute lymphoblastic leukaemia"

Article Title: ETS1 and SP1 drive DHX15 expression in acute lymphoblastic leukaemia, et al. ETS1 and SP1 drive DHX15 expression in acute lymphoblastic leukaemia

Journal: Journal of Cellular and Molecular Medicine

doi: 10.1111/jcmm.13525

EMSA and Ch IP analyses of ETS 1 and SP 1 binding to the DHX 15 promoter. A, EMSA of ETS1 (left) and SP1 (right). The 5′‐biotin end‐labelled probe was incubated in the absence (lane 0) or presence (lane 1) of nuclear extracts from Jurkat and NALM6 cells. A cold mutated probe (lane 2) and cold probe (lane 3) were used as competitors at concentrations that were in a 100‐fold molar excess to the biotin‐labelled probe. Supershift assays were performed with 4 μg of a specific antibody against ETS1 or SP1 (lane 4). B, Equal amounts of Jurkat and NALM6 chromatin were immunoprecipitated with antibodies for ETS1 and SP1 and subsequently quantified through agarose gel electrophoresis using a primer set specific for the basal region (−181 to −36 bp). Moreover, immunoprecipitated DNA was amplified using a primer set specific to the off‐target region (GAPDH) shown in the lower panel as a negative control
Figure Legend Snippet: EMSA and Ch IP analyses of ETS 1 and SP 1 binding to the DHX 15 promoter. A, EMSA of ETS1 (left) and SP1 (right). The 5′‐biotin end‐labelled probe was incubated in the absence (lane 0) or presence (lane 1) of nuclear extracts from Jurkat and NALM6 cells. A cold mutated probe (lane 2) and cold probe (lane 3) were used as competitors at concentrations that were in a 100‐fold molar excess to the biotin‐labelled probe. Supershift assays were performed with 4 μg of a specific antibody against ETS1 or SP1 (lane 4). B, Equal amounts of Jurkat and NALM6 chromatin were immunoprecipitated with antibodies for ETS1 and SP1 and subsequently quantified through agarose gel electrophoresis using a primer set specific for the basal region (−181 to −36 bp). Moreover, immunoprecipitated DNA was amplified using a primer set specific to the off‐target region (GAPDH) shown in the lower panel as a negative control

Techniques Used: Binding Assay, Incubation, Immunoprecipitation, Agarose Gel Electrophoresis, Amplification, Negative Control

10) Product Images from "PPARγ regulated CIDEA affects pro-apoptotic responses in glioblastoma"

Article Title: PPARγ regulated CIDEA affects pro-apoptotic responses in glioblastoma

Journal: Cell Death Discovery

doi: 10.1038/cddiscovery.2015.38

Inhibition of PPAR γ reduces NF κ B and SP1 binding on CIDEA promoter. ( a ) PPAR γ inhibitior T007 has no effect on NF κ B and SP1 expression. Blot is representative of two independent experiments. Blots were re-probed for GAPDH to establish equal loading. ( b ) ChIP-qPCR assays demonstrating decreased binding of NF κ B to its cognate sites on CIDEA promoter. DNA isolated from control and PPAR γ inhibitor treated A172 glioma cells pre and post immunoprecipitation with anti-NF κ B antibody, was amplified using specific primer sets. Binding affinity of NF κ B was found to be low at three different putative binding sites on CIDEA promoter on inhibition of PPAR γ . ( c ) PPAR γ inhibition decreases SP1 binding to its cognate site on CIDEA promoter at −26 to +120 position, as indicated by ChIP-qPCR assay. Graph ( b and c ) represents fold change as calculated from Ct values of two independent experiments for a single site.
Figure Legend Snippet: Inhibition of PPAR γ reduces NF κ B and SP1 binding on CIDEA promoter. ( a ) PPAR γ inhibitior T007 has no effect on NF κ B and SP1 expression. Blot is representative of two independent experiments. Blots were re-probed for GAPDH to establish equal loading. ( b ) ChIP-qPCR assays demonstrating decreased binding of NF κ B to its cognate sites on CIDEA promoter. DNA isolated from control and PPAR γ inhibitor treated A172 glioma cells pre and post immunoprecipitation with anti-NF κ B antibody, was amplified using specific primer sets. Binding affinity of NF κ B was found to be low at three different putative binding sites on CIDEA promoter on inhibition of PPAR γ . ( c ) PPAR γ inhibition decreases SP1 binding to its cognate site on CIDEA promoter at −26 to +120 position, as indicated by ChIP-qPCR assay. Graph ( b and c ) represents fold change as calculated from Ct values of two independent experiments for a single site.

Techniques Used: Inhibition, Binding Assay, Expressing, Chromatin Immunoprecipitation, Real-time Polymerase Chain Reaction, Isolation, Immunoprecipitation, Amplification

11) Product Images from "Matrix metalloproteinase MMP9 maintains epithelial barrier function and preserves mucosal lining in colitis associated cancer"

Article Title: Matrix metalloproteinase MMP9 maintains epithelial barrier function and preserves mucosal lining in colitis associated cancer

Journal: Oncotarget

doi: 10.18632/oncotarget.21841

MMP9 activates EGFR1 signaling The regulation of MUC2 was investigated by using CaCo2BBE cells overexpressing MMP9 and performing WBs (25μg/lane) using whole cell lysates probed with (A) anti-EGFR, (B) anti-Sp1 and (C) anti-STAT3. The loading control for the each blot were GAPDH or β-actin. Each blot was a representation of three individual experiments. Densitometry evaluations of the WB is represented by the adjacent bar graph with each bar represents mean ± S.E., *p
Figure Legend Snippet: MMP9 activates EGFR1 signaling The regulation of MUC2 was investigated by using CaCo2BBE cells overexpressing MMP9 and performing WBs (25μg/lane) using whole cell lysates probed with (A) anti-EGFR, (B) anti-Sp1 and (C) anti-STAT3. The loading control for the each blot were GAPDH or β-actin. Each blot was a representation of three individual experiments. Densitometry evaluations of the WB is represented by the adjacent bar graph with each bar represents mean ± S.E., *p

Techniques Used: Western Blot

12) Product Images from "TGFβ Signaling Regulates the Timing of CNS Myelination by Modulating Oligodendrocyte Progenitor Cell Cycle Exit through SMAD3/4/FoxO1/Sp1"

Article Title: TGFβ Signaling Regulates the Timing of CNS Myelination by Modulating Oligodendrocyte Progenitor Cell Cycle Exit through SMAD3/4/FoxO1/Sp1

Journal:

doi: 10.1523/JNEUROSCI.0363-14.2014

Proposed model of TGFβ signaling modulation of oligodendrogenesis during the SCWM myelination. TGFβ signaling in OPs controls the canonical downstream TGFβ-R effectors, SMAD2/3/4. Upon TGFβ-R's activation, the heteromeric SMAD2/3/4 complex establishes a nuclear localization, and cooperates with FoxO1 and Sp1 to modulate the transcription of c-myc and p21. Upon binding regulatory sequences, SMAD2/3/4, FoxO1, and Sp1 cooperate to repress the transcription of c-myc and activate p21 transcription, allowing OPs to withdraw from the cell cycle and progress down OL differentiation and maturation.
Figure Legend Snippet: Proposed model of TGFβ signaling modulation of oligodendrogenesis during the SCWM myelination. TGFβ signaling in OPs controls the canonical downstream TGFβ-R effectors, SMAD2/3/4. Upon TGFβ-R's activation, the heteromeric SMAD2/3/4 complex establishes a nuclear localization, and cooperates with FoxO1 and Sp1 to modulate the transcription of c-myc and p21. Upon binding regulatory sequences, SMAD2/3/4, FoxO1, and Sp1 cooperate to repress the transcription of c-myc and activate p21 transcription, allowing OPs to withdraw from the cell cycle and progress down OL differentiation and maturation.

Techniques Used: Activation Assay, Binding Assay

Related Articles

Blocking Assay:

Article Title: Adeno-associated virus-mediated brain delivery of 5-lipoxygenase modulates the AD-like phenotype of APP mice
Article Snippet: They were blocked with Odyssey blocking buffer for 1 hr; and then incubated with primary antibodies overnight at 4°C. .. Primary antibodies used were as follows: anti-APP N-terminal raised against amino acids 66-81 for total APP (22C11; Chemicon Int.), anti-BACE-1 (IBL America), anti-ADAM-10 (Chemicon Int.), anti-PS1 (Cell Signaling), anti-nicastrin (Cell Signaling), anti-Pen2 (Invitrogen), anti-APH-1 (Millipore); anti-sAPPα (2B3; IBL America); anti-sAPPβ (6A1, IBL America); anti-CTFs (EMD Biosciences Inc.); anti-neprilysin (Santa Curz); anti-IDE N-terminal (EMD Biosciences Inc.); anti-apoE (Santa Cruz); anti-CREB and anti-p-CREB (Cell Signaling); anti-Sp1 (Cell Signaling); anti-5-LO (BD Bioscience), anti-β actin (Santa Cruz).

Article Title: GRP78‐mediated antioxidant response and ABC transporter activity confers chemoresistance to pancreatic cancer cells
Article Snippet: Tissues were then blocked with Dako protein block and incubated with primary antibody overnight. .. Slides were stained with anti‐SP1 (Cell Signaling, Cat # 9389) and anti‐GRP78 (Cell Signaling, Cat # 3177) at 1 : 200 dilutions.

Real-time Polymerase Chain Reaction:

Article Title: SP1 and RARα regulate AGAP2 expression in cancer
Article Snippet: Afterwards, crosslinked proteins of interest were immunoprecipitated with 2 μg of either anti-RARα (C-20), anti-RXRα (D-20) (both from Santa Cruz Biotechnology), anti-PCAF (C14G9, Cell Signaling), or 1 μg of anti-SP1 (D4C3, Cell Signalling). .. Anti-Pol II (N-20, Santa Cruz Biotechnology) was used as positive control and rabbit IgG (Invitrogen) as negative control.

Article Title: SP1-induced lncRNA-ZFAS1 contributes to colorectal cancer progression via the miR-150-5p/VEGFA axis
Article Snippet: Briefly, the CRC cells were fixed with 1% formaldehyde solution for 15 min and incubated with 125 nM glycine for 5 min. DNA fragments ranging from 200 to 300 bp were generated using sonication. .. Antibodies including anti-SP1 (#9389, Cell Signaling Technology, USA) and IgG were employed for each immunoprecipitation. qPCR was used to analyze the precipitated DNA. .. RIP assay was performed with Magna RIPTM RNA-Binding Protein Immunoprecipation Kit (Millipore, Billerica, USA) and AGO2 antibody (proteintech, 10686–1-AP, China) in accordance with the manufacturer’s protocol.

Article Title: PPARγ regulated CIDEA affects pro-apoptotic responses in glioblastoma
Article Snippet: Paragraph title: ChIP and ChIP real-time PCR assays ... Anti-NFκ B (Santa Cruz Biotechnology) and anti-SP1 (Cell signaling) were used for immunoprecipitation and non-specific IgG antibody (Abcam) was used as control.

Article Title: TGFβ Signaling Regulates the Timing of CNS Myelination by Modulating Oligodendrocyte Progenitor Cell Cycle Exit through SMAD3/4/FoxO1/Sp1
Article Snippet: Immunoprecipitation was performed with anti-SMAD3, anti-SMAD4 (Santa Cruz Biotechnology), anti-Sp1, and anti-FoxO1 (Cell Signaling Technology) antibodies or a nonspecific rabbit IgG as control. .. The presence of DNA bound to each transcription factor (TF) from the indicated regions of the two promoters analyzed was determined by qPCR analysis as above.

Article Title: Gene expression analysis of human induced pluripotent stem cell-derived neurons carrying copy number variants of chromosome 15q11-q13.1
Article Snippet: For ChIP analysis of Sp1 binding, the following antibodies were used: mouse monoclonal anti-Sp1 (sc-17824, Santa Cruz Biotechnology, Inc, Dallas, TX, USA) and anti-Sp1 (#9389, Cell Signaling Technology, Beverly, MA, USA). .. For ChIP analysis of Sp1 binding, the following antibodies were used: mouse monoclonal anti-Sp1 (sc-17824, Santa Cruz Biotechnology, Inc, Dallas, TX, USA) and anti-Sp1 (#9389, Cell Signaling Technology, Beverly, MA, USA).

Incubation:

Article Title: SP1-induced lncRNA-ZFAS1 contributes to colorectal cancer progression via the miR-150-5p/VEGFA axis
Article Snippet: Briefly, the CRC cells were fixed with 1% formaldehyde solution for 15 min and incubated with 125 nM glycine for 5 min. DNA fragments ranging from 200 to 300 bp were generated using sonication. .. Antibodies including anti-SP1 (#9389, Cell Signaling Technology, USA) and IgG were employed for each immunoprecipitation. qPCR was used to analyze the precipitated DNA.

Article Title: miR-506 attenuates methylation of lncRNA MEG3 to inhibit migration and invasion of breast cancer cell lines via targeting SP1 and SP3
Article Snippet: Then, the membrane was washed with 1 × TBST for 3 times, 5 min each time, and incubated with secondary antibodies at room temperature for 1 h. Finally, the membrane was incubated with ECL and exposed. .. The following antibodies were used: anti-SP3 (Santa Cruz, USA), anti-DNMT1 (Cell Signaling Technology, USA), anti-SP1 (Cell Signaling Technology, USA), anti-β-actin (Proteintech).

Article Title: Adeno-associated virus-mediated brain delivery of 5-lipoxygenase modulates the AD-like phenotype of APP mice
Article Snippet: After three washing cycles with T-TBS, membranes were incubated with IRDye 800CW or IRDye 680CW-labeled secondary antibodies (LI-COR Bioscience) at 22°C for 1 hr. .. Primary antibodies used were as follows: anti-APP N-terminal raised against amino acids 66-81 for total APP (22C11; Chemicon Int.), anti-BACE-1 (IBL America), anti-ADAM-10 (Chemicon Int.), anti-PS1 (Cell Signaling), anti-nicastrin (Cell Signaling), anti-Pen2 (Invitrogen), anti-APH-1 (Millipore); anti-sAPPα (2B3; IBL America); anti-sAPPβ (6A1, IBL America); anti-CTFs (EMD Biosciences Inc.); anti-neprilysin (Santa Curz); anti-IDE N-terminal (EMD Biosciences Inc.); anti-apoE (Santa Cruz); anti-CREB and anti-p-CREB (Cell Signaling); anti-Sp1 (Cell Signaling); anti-5-LO (BD Bioscience), anti-β actin (Santa Cruz).

Article Title: ETS1 and SP1 drive DHX15 expression in acute lymphoblastic leukaemia, et al. ETS1 and SP1 drive DHX15 expression in acute lymphoblastic leukaemia
Article Snippet: The membranes were blocked by incubation with 5% skim milk in TBST buffer (10 mmol/L Tris‐HCl, pH 8.0, 150 mmol/L NaCl and 0.1% Tween‐20). .. After incubation with the anti‐ETS1 (14069, 1:1000 dilution, CST), anti‐SP1 (9389, 1:1000 dilution, CST), anti‐DHX15 (1:1500 dilution, Proteintech) or anti‐β‐actin (1:2000 dilution, TransGen) antibody, the detection was performed with HRP‐coupled secondary antibodies (Millipore). .. The intensity of each band was calculated using the Quantity One software (Bio‐Rad, Hercules, CA).

Article Title: GRP78‐mediated antioxidant response and ABC transporter activity confers chemoresistance to pancreatic cancer cells
Article Snippet: Tissues were then blocked with Dako protein block and incubated with primary antibody overnight. .. Slides were stained with anti‐SP1 (Cell Signaling, Cat # 9389) and anti‐GRP78 (Cell Signaling, Cat # 3177) at 1 : 200 dilutions.

Stripping Membranes:

Article Title: Matrix metalloproteinase MMP9 maintains epithelial barrier function and preserves mucosal lining in colitis associated cancer
Article Snippet: As described previously [ ], for WB analysis, colonic mucosal stripping was obtained from the TgM9 and WT mice (n=20 per group) with and without CAC. .. The antibodies used were anti-MMP9 (Abcam, Cambridge, MA), anti-EGFR (Cell Signaling, Beverly, MA), anti-Claudin-2 (Life Technologies, Rockford, IL), anti-Claudin-4 (Invitrogen, Rockford, IL), anti-Claudin-5 (Invitrogen), anti-TFF3 (Cloud-Clone Corp., Katy, TX), anti-Sp1 (Upstate Cell Signaling Solutions, Lake Placid, NY), anti-Occludin (Invitrogen), anti-STAT3 (Cell Signaling).

BIA-KA:

Article Title: ETS1 and SP1 drive DHX15 expression in acute lymphoblastic leukaemia, et al. ETS1 and SP1 drive DHX15 expression in acute lymphoblastic leukaemia
Article Snippet: The concentrations of the protein extracts were measured using the BCA Protein Assay Kit (Pierce, Rockford, IL, USA). .. After incubation with the anti‐ETS1 (14069, 1:1000 dilution, CST), anti‐SP1 (9389, 1:1000 dilution, CST), anti‐DHX15 (1:1500 dilution, Proteintech) or anti‐β‐actin (1:2000 dilution, TransGen) antibody, the detection was performed with HRP‐coupled secondary antibodies (Millipore).

Article Title: Role of Cdc6 in re-replication in cells expressing human papillomavirus E7 oncogene
Article Snippet: The protein concentration was measured using a BCA protein assay kit. .. The purity of the obtained fractions was confirmed using anti-β-tubulin (Sigma, T-4026, for the CEs), anti-Sp1 (Cell Signaling #9389, for the SNEs) or anti-Histone H3 (Cell Signaling #3688, for the CBEs).

Western Blot:

Article Title: Overexpression of histone deacetylases in cancer cells is controlled by interplay of transcription factors and epigenetic modulators
Article Snippet: After 48 h, firefly and Renilla luciferase activities were measured using the dual-luciferase reporter assay system (Promega). .. Antibodies for Western blot, immunoprecipitation, and chromatin immunoprecipitation (ChIP) were as follows: anti-HDAC1 and anti-HDAC2 (Pierce); anti-Sp1 (Cell Signaling Technology), anti-Sp3 and anti-p300 (Santa Cruz Biotechnology); anti-p21, anti-acetyl-lysine, anti-ac-H3, anti-H3, anti-H3K4me3, anti-SET1, and anti-RbBP5 (Milipore, Temecula, CA, USA); anti-Ash2L (Bethyl Laboratories, Montgomery, TX, USA); and anti-β-actin and anti-α-tubulin (Sigma-Aldrich, St. Louis, MO, USA). .. HDAC1 and acetylated HDAC1 antibodies were generated as described previously ( ).

Article Title: MicroRNA 373 Facilitates the Replication of Porcine Reproductive and Respiratory Syndrome Virus by Its Negative Regulation of Type I Interferon Induction
Article Snippet: The relative luciferase activity was determined by the dual-luciferase assays. .. For Western blot experiments, the primary antibodies anti-Flag (F1804; Sigma), anti-NFIA (AP20998c; Abgent), anti-NFIB (AP17409c; Abgent), anti-IRAK1 (4504; CST), anti-IRAK4 (4363; CST), anti-IRF1 (8478; CST), anti-IRF9 (28492; CST), anti-Ago2 (2897; CST), anti-Sp1 (9389; CST), anti-actin (4967; CST), anti-rabbit IgG-horseradish peroxidase (HRP)-linked antibody (7074; CST), anti-mouse IgG-HRP-linked antibody (7076; CST), and anti-PRRSV N monoclonal antibody (2D6) were used. .. MARC-145 cells were lysed, and the cellular proteins were separated by SDS-PAGE and transferred onto a polyvinylidene difluoride (PVDF) membrane.

Article Title: miR-506 attenuates methylation of lncRNA MEG3 to inhibit migration and invasion of breast cancer cell lines via targeting SP1 and SP3
Article Snippet: Paragraph title: Western blot ... The following antibodies were used: anti-SP3 (Santa Cruz, USA), anti-DNMT1 (Cell Signaling Technology, USA), anti-SP1 (Cell Signaling Technology, USA), anti-β-actin (Proteintech).

Article Title: ETS1 and SP1 drive DHX15 expression in acute lymphoblastic leukaemia, et al. ETS1 and SP1 drive DHX15 expression in acute lymphoblastic leukaemia
Article Snippet: Paragraph title: Western blotting ... After incubation with the anti‐ETS1 (14069, 1:1000 dilution, CST), anti‐SP1 (9389, 1:1000 dilution, CST), anti‐DHX15 (1:1500 dilution, Proteintech) or anti‐β‐actin (1:2000 dilution, TransGen) antibody, the detection was performed with HRP‐coupled secondary antibodies (Millipore).

Article Title: Matrix metalloproteinase MMP9 maintains epithelial barrier function and preserves mucosal lining in colitis associated cancer
Article Snippet: Paragraph title: Western blot ... The antibodies used were anti-MMP9 (Abcam, Cambridge, MA), anti-EGFR (Cell Signaling, Beverly, MA), anti-Claudin-2 (Life Technologies, Rockford, IL), anti-Claudin-4 (Invitrogen, Rockford, IL), anti-Claudin-5 (Invitrogen), anti-TFF3 (Cloud-Clone Corp., Katy, TX), anti-Sp1 (Upstate Cell Signaling Solutions, Lake Placid, NY), anti-Occludin (Invitrogen), anti-STAT3 (Cell Signaling).

Article Title: Serum deprivation/starvation leads to reactivation of HIV-1 in latently infected monocytes via activating ERK/JNK pathway
Article Snippet: Paragraph title: Western blot analysis ... The primary antibodies used were anti-AKT, anti-S473 phopsho-AKT, anti-GAPDH, anti-PARP, anti-Caspase-3, anti-ERK1/2, anti-phospho 44/42 ERK1/2, anti-phospho-SAPK/JNK, anti-SAPK/JNK, anti-SP1, anti-LC3B (Cell Signalling Technology), anti-p24 (Cat No. 6457, NIH), anti-IKBα (Santa Cruz Biotechnology) The secondary antibodies used were anti-rabbit/mouse-HRP conjugated (Jackson Immuno Research).

Immunohistochemistry:

Article Title: GRP78‐mediated antioxidant response and ABC transporter activity confers chemoresistance to pancreatic cancer cells
Article Snippet: Paragraph title: Immunohistochemistry ... Slides were stained with anti‐SP1 (Cell Signaling, Cat # 9389) and anti‐GRP78 (Cell Signaling, Cat # 3177) at 1 : 200 dilutions.

Gas Chromatography:

Article Title: Long noncoding RNA UCA1 induced by SP1 promotes cell proliferation via recruiting EZH2 and activating AKT pathway in gastric cancer
Article Snippet: Briefly, GC cells were cross-linked in 1% formaldehyde solution for 10 min at room temperature, followed by the addition of 125 mM of glycine for 5 min. DNA fragments ranging from 200 to 500 bp were yielded via sonication. .. Antibodies including anti-SP1, anti-EZH2 (Cell Signaling Technology, Danvers, MA, USA) and normal IgG were used for each immunoprecipitation.

Protease Inhibitor:

Article Title: ETS1 and SP1 drive DHX15 expression in acute lymphoblastic leukaemia, et al. ETS1 and SP1 drive DHX15 expression in acute lymphoblastic leukaemia
Article Snippet: 2.9 RIPA buffer supplemented with the ProteinSafe Protease Inhibitor Cocktail (100×) (TransGenBiotek, China) was used to isolate total proteins from the Jurkat and NALM6 cells. .. After incubation with the anti‐ETS1 (14069, 1:1000 dilution, CST), anti‐SP1 (9389, 1:1000 dilution, CST), anti‐DHX15 (1:1500 dilution, Proteintech) or anti‐β‐actin (1:2000 dilution, TransGen) antibody, the detection was performed with HRP‐coupled secondary antibodies (Millipore).

Immunolabeling:

Article Title: MicroRNA 373 Facilitates the Replication of Porcine Reproductive and Respiratory Syndrome Virus by Its Negative Regulation of Type I Interferon Induction
Article Snippet: For Western blot experiments, the primary antibodies anti-Flag (F1804; Sigma), anti-NFIA (AP20998c; Abgent), anti-NFIB (AP17409c; Abgent), anti-IRAK1 (4504; CST), anti-IRAK4 (4363; CST), anti-IRF1 (8478; CST), anti-IRF9 (28492; CST), anti-Ago2 (2897; CST), anti-Sp1 (9389; CST), anti-actin (4967; CST), anti-rabbit IgG-horseradish peroxidase (HRP)-linked antibody (7074; CST), anti-mouse IgG-HRP-linked antibody (7076; CST), and anti-PRRSV N monoclonal antibody (2D6) were used. .. MARC-145 cells were lysed, and the cellular proteins were separated by SDS-PAGE and transferred onto a polyvinylidene difluoride (PVDF) membrane.

Cell Culture:

Article Title: TGFβ Signaling Regulates the Timing of CNS Myelination by Modulating Oligodendrocyte Progenitor Cell Cycle Exit through SMAD3/4/FoxO1/Sp1
Article Snippet: CNP -EGFP FACS-sorted cells were cultured under proliferating conditions and were treated with TGFβ1 or vehicle and 4 h later samples were processed as recommended by the manufacturer (EZ-ChIP; Millipore). .. Immunoprecipitation was performed with anti-SMAD3, anti-SMAD4 (Santa Cruz Biotechnology), anti-Sp1, and anti-FoxO1 (Cell Signaling Technology) antibodies or a nonspecific rabbit IgG as control.

Generated:

Article Title: SP1-induced lncRNA-ZFAS1 contributes to colorectal cancer progression via the miR-150-5p/VEGFA axis
Article Snippet: Briefly, the CRC cells were fixed with 1% formaldehyde solution for 15 min and incubated with 125 nM glycine for 5 min. DNA fragments ranging from 200 to 300 bp were generated using sonication. .. Antibodies including anti-SP1 (#9389, Cell Signaling Technology, USA) and IgG were employed for each immunoprecipitation. qPCR was used to analyze the precipitated DNA.

Article Title: Matrix metalloproteinase MMP9 maintains epithelial barrier function and preserves mucosal lining in colitis associated cancer
Article Snippet: The antibodies used were anti-MMP9 (Abcam, Cambridge, MA), anti-EGFR (Cell Signaling, Beverly, MA), anti-Claudin-2 (Life Technologies, Rockford, IL), anti-Claudin-4 (Invitrogen, Rockford, IL), anti-Claudin-5 (Invitrogen), anti-TFF3 (Cloud-Clone Corp., Katy, TX), anti-Sp1 (Upstate Cell Signaling Solutions, Lake Placid, NY), anti-Occludin (Invitrogen), anti-STAT3 (Cell Signaling). .. Goat anti-mouse secondary antibody (Bio-Rad, Hercules, CA) or goat anti-rabbit secondary antibody (Bio-Rad) were used.

Imaging:

Article Title: Adeno-associated virus-mediated brain delivery of 5-lipoxygenase modulates the AD-like phenotype of APP mice
Article Snippet: Primary antibodies used were as follows: anti-APP N-terminal raised against amino acids 66-81 for total APP (22C11; Chemicon Int.), anti-BACE-1 (IBL America), anti-ADAM-10 (Chemicon Int.), anti-PS1 (Cell Signaling), anti-nicastrin (Cell Signaling), anti-Pen2 (Invitrogen), anti-APH-1 (Millipore); anti-sAPPα (2B3; IBL America); anti-sAPPβ (6A1, IBL America); anti-CTFs (EMD Biosciences Inc.); anti-neprilysin (Santa Curz); anti-IDE N-terminal (EMD Biosciences Inc.); anti-apoE (Santa Cruz); anti-CREB and anti-p-CREB (Cell Signaling); anti-Sp1 (Cell Signaling); anti-5-LO (BD Bioscience), anti-β actin (Santa Cruz). .. Primary antibodies used were as follows: anti-APP N-terminal raised against amino acids 66-81 for total APP (22C11; Chemicon Int.), anti-BACE-1 (IBL America), anti-ADAM-10 (Chemicon Int.), anti-PS1 (Cell Signaling), anti-nicastrin (Cell Signaling), anti-Pen2 (Invitrogen), anti-APH-1 (Millipore); anti-sAPPα (2B3; IBL America); anti-sAPPβ (6A1, IBL America); anti-CTFs (EMD Biosciences Inc.); anti-neprilysin (Santa Curz); anti-IDE N-terminal (EMD Biosciences Inc.); anti-apoE (Santa Cruz); anti-CREB and anti-p-CREB (Cell Signaling); anti-Sp1 (Cell Signaling); anti-5-LO (BD Bioscience), anti-β actin (Santa Cruz).

Protein Concentration:

Article Title: Role of Cdc6 in re-replication in cells expressing human papillomavirus E7 oncogene
Article Snippet: The protein concentration was measured using a BCA protein assay kit. .. The purity of the obtained fractions was confirmed using anti-β-tubulin (Sigma, T-4026, for the CEs), anti-Sp1 (Cell Signaling #9389, for the SNEs) or anti-Histone H3 (Cell Signaling #3688, for the CBEs).

Sequencing:

Article Title: TGFβ Signaling Regulates the Timing of CNS Myelination by Modulating Oligodendrocyte Progenitor Cell Cycle Exit through SMAD3/4/FoxO1/Sp1
Article Snippet: Immunoprecipitation was performed with anti-SMAD3, anti-SMAD4 (Santa Cruz Biotechnology), anti-Sp1, and anti-FoxO1 (Cell Signaling Technology) antibodies or a nonspecific rabbit IgG as control. .. Immunoprecipitation was performed with anti-SMAD3, anti-SMAD4 (Santa Cruz Biotechnology), anti-Sp1, and anti-FoxO1 (Cell Signaling Technology) antibodies or a nonspecific rabbit IgG as control.

Sonication:

Article Title: SP1 and RARα regulate AGAP2 expression in cancer
Article Snippet: The chromatin was sheared to 100–600 bp by sonication using an AFA Focused-Ultrasonicator (S220- Series from CovarisTM ) at 6 × 60 second on-off pulses at 4 °C. .. Afterwards, crosslinked proteins of interest were immunoprecipitated with 2 μg of either anti-RARα (C-20), anti-RXRα (D-20) (both from Santa Cruz Biotechnology), anti-PCAF (C14G9, Cell Signaling), or 1 μg of anti-SP1 (D4C3, Cell Signalling).

Article Title: Long noncoding RNA UCA1 induced by SP1 promotes cell proliferation via recruiting EZH2 and activating AKT pathway in gastric cancer
Article Snippet: Briefly, GC cells were cross-linked in 1% formaldehyde solution for 10 min at room temperature, followed by the addition of 125 mM of glycine for 5 min. DNA fragments ranging from 200 to 500 bp were yielded via sonication. .. Antibodies including anti-SP1, anti-EZH2 (Cell Signaling Technology, Danvers, MA, USA) and normal IgG were used for each immunoprecipitation.

Article Title: SP1-induced lncRNA-ZFAS1 contributes to colorectal cancer progression via the miR-150-5p/VEGFA axis
Article Snippet: Briefly, the CRC cells were fixed with 1% formaldehyde solution for 15 min and incubated with 125 nM glycine for 5 min. DNA fragments ranging from 200 to 300 bp were generated using sonication. .. Antibodies including anti-SP1 (#9389, Cell Signaling Technology, USA) and IgG were employed for each immunoprecipitation. qPCR was used to analyze the precipitated DNA.

Article Title: Gene expression analysis of human induced pluripotent stem cell-derived neurons carrying copy number variants of chromosome 15q11-q13.1
Article Snippet: Instead of a two-step cell lysis, cells were lysed once for 15 minutes in an SDS Cell Lysis Buffer (Millipore, Billerica, MA, USA) before sonication. .. For ChIP analysis of Sp1 binding, the following antibodies were used: mouse monoclonal anti-Sp1 (sc-17824, Santa Cruz Biotechnology, Inc, Dallas, TX, USA) and anti-Sp1 (#9389, Cell Signaling Technology, Beverly, MA, USA).

Article Title: Role of Cdc6 in re-replication in cells expressing human papillomavirus E7 oncogene
Article Snippet: The final chromatin pellet was resuspended in 1× Laemmli buffer without dithiothreitol and bromophenol blue for 10min at 70°C and sonicated for 15s in a 4710 Series Ultrasonic Homogenizer using a microtip at 25% amplitude (Cole-Parmer Instrument Co., Chicago, IL). .. The purity of the obtained fractions was confirmed using anti-β-tubulin (Sigma, T-4026, for the CEs), anti-Sp1 (Cell Signaling #9389, for the SNEs) or anti-Histone H3 (Cell Signaling #3688, for the CBEs).

Binding Assay:

Article Title: TGFβ Signaling Regulates the Timing of CNS Myelination by Modulating Oligodendrocyte Progenitor Cell Cycle Exit through SMAD3/4/FoxO1/Sp1
Article Snippet: Immunoprecipitation was performed with anti-SMAD3, anti-SMAD4 (Santa Cruz Biotechnology), anti-Sp1, and anti-FoxO1 (Cell Signaling Technology) antibodies or a nonspecific rabbit IgG as control. .. Immunoprecipitation was performed with anti-SMAD3, anti-SMAD4 (Santa Cruz Biotechnology), anti-Sp1, and anti-FoxO1 (Cell Signaling Technology) antibodies or a nonspecific rabbit IgG as control.

Article Title: Gene expression analysis of human induced pluripotent stem cell-derived neurons carrying copy number variants of chromosome 15q11-q13.1
Article Snippet: A rabbit polyclonal anti-YY-1 (sc-281, Santa Cruz Biotechnology, Inc, Dallas, TX, USA) was used at 5 μg per immunoprecipitation reaction. .. For ChIP analysis of Sp1 binding, the following antibodies were used: mouse monoclonal anti-Sp1 (sc-17824, Santa Cruz Biotechnology, Inc, Dallas, TX, USA) and anti-Sp1 (#9389, Cell Signaling Technology, Beverly, MA, USA). .. Novex Protein A DynaBeads magnetic beads (Life Technologies, Grand Island, NY, USA) were used during immunoprecipitation.

Cellular Antioxidant Activity Assay:

Article Title: Gene expression analysis of human induced pluripotent stem cell-derived neurons carrying copy number variants of chromosome 15q11-q13.1
Article Snippet: For ChIP analysis of Sp1 binding, the following antibodies were used: mouse monoclonal anti-Sp1 (sc-17824, Santa Cruz Biotechnology, Inc, Dallas, TX, USA) and anti-Sp1 (#9389, Cell Signaling Technology, Beverly, MA, USA). .. For ChIP analysis of Sp1 binding, the following antibodies were used: mouse monoclonal anti-Sp1 (sc-17824, Santa Cruz Biotechnology, Inc, Dallas, TX, USA) and anti-Sp1 (#9389, Cell Signaling Technology, Beverly, MA, USA).

Molecular Weight:

Article Title: Adeno-associated virus-mediated brain delivery of 5-lipoxygenase modulates the AD-like phenotype of APP mice
Article Snippet: Samples were electrophoretically separated using 10% Bis-Tris gels or 3-8% Tris-acetate gel (Bio-Rad, Richmond, CA), according to the molecular weight of the target molecule, and then transferred onto nitrocellulose membranes (Bio-Rad). .. Primary antibodies used were as follows: anti-APP N-terminal raised against amino acids 66-81 for total APP (22C11; Chemicon Int.), anti-BACE-1 (IBL America), anti-ADAM-10 (Chemicon Int.), anti-PS1 (Cell Signaling), anti-nicastrin (Cell Signaling), anti-Pen2 (Invitrogen), anti-APH-1 (Millipore); anti-sAPPα (2B3; IBL America); anti-sAPPβ (6A1, IBL America); anti-CTFs (EMD Biosciences Inc.); anti-neprilysin (Santa Curz); anti-IDE N-terminal (EMD Biosciences Inc.); anti-apoE (Santa Cruz); anti-CREB and anti-p-CREB (Cell Signaling); anti-Sp1 (Cell Signaling); anti-5-LO (BD Bioscience), anti-β actin (Santa Cruz).

In Vivo:

Article Title: Matrix metalloproteinase MMP9 maintains epithelial barrier function and preserves mucosal lining in colitis associated cancer
Article Snippet: In vivo WB analysis was performed with 30μg/ well of cell lysates. .. The antibodies used were anti-MMP9 (Abcam, Cambridge, MA), anti-EGFR (Cell Signaling, Beverly, MA), anti-Claudin-2 (Life Technologies, Rockford, IL), anti-Claudin-4 (Invitrogen, Rockford, IL), anti-Claudin-5 (Invitrogen), anti-TFF3 (Cloud-Clone Corp., Katy, TX), anti-Sp1 (Upstate Cell Signaling Solutions, Lake Placid, NY), anti-Occludin (Invitrogen), anti-STAT3 (Cell Signaling).

Purification:

Article Title: SP1 and RARα regulate AGAP2 expression in cancer
Article Snippet: Afterwards, crosslinked proteins of interest were immunoprecipitated with 2 μg of either anti-RARα (C-20), anti-RXRα (D-20) (both from Santa Cruz Biotechnology), anti-PCAF (C14G9, Cell Signaling), or 1 μg of anti-SP1 (D4C3, Cell Signalling). .. Anti-Pol II (N-20, Santa Cruz Biotechnology) was used as positive control and rabbit IgG (Invitrogen) as negative control.

Article Title: Gene expression analysis of human induced pluripotent stem cell-derived neurons carrying copy number variants of chromosome 15q11-q13.1
Article Snippet: For ChIP analysis of Sp1 binding, the following antibodies were used: mouse monoclonal anti-Sp1 (sc-17824, Santa Cruz Biotechnology, Inc, Dallas, TX, USA) and anti-Sp1 (#9389, Cell Signaling Technology, Beverly, MA, USA). .. Novex Protein A DynaBeads magnetic beads (Life Technologies, Grand Island, NY, USA) were used during immunoprecipitation.

Polymerase Chain Reaction:

Article Title: TGFβ Signaling Regulates the Timing of CNS Myelination by Modulating Oligodendrocyte Progenitor Cell Cycle Exit through SMAD3/4/FoxO1/Sp1
Article Snippet: Immunoprecipitation was performed with anti-SMAD3, anti-SMAD4 (Santa Cruz Biotechnology), anti-Sp1, and anti-FoxO1 (Cell Signaling Technology) antibodies or a nonspecific rabbit IgG as control. .. The presence of DNA bound to each transcription factor (TF) from the indicated regions of the two promoters analyzed was determined by qPCR analysis as above.

FACS:

Article Title: TGFβ Signaling Regulates the Timing of CNS Myelination by Modulating Oligodendrocyte Progenitor Cell Cycle Exit through SMAD3/4/FoxO1/Sp1
Article Snippet: CNP -EGFP FACS-sorted cells were cultured under proliferating conditions and were treated with TGFβ1 or vehicle and 4 h later samples were processed as recommended by the manufacturer (EZ-ChIP; Millipore). .. Immunoprecipitation was performed with anti-SMAD3, anti-SMAD4 (Santa Cruz Biotechnology), anti-Sp1, and anti-FoxO1 (Cell Signaling Technology) antibodies or a nonspecific rabbit IgG as control.

Lysis:

Article Title: Gene expression analysis of human induced pluripotent stem cell-derived neurons carrying copy number variants of chromosome 15q11-q13.1
Article Snippet: Instead of a two-step cell lysis, cells were lysed once for 15 minutes in an SDS Cell Lysis Buffer (Millipore, Billerica, MA, USA) before sonication. .. For ChIP analysis of Sp1 binding, the following antibodies were used: mouse monoclonal anti-Sp1 (sc-17824, Santa Cruz Biotechnology, Inc, Dallas, TX, USA) and anti-Sp1 (#9389, Cell Signaling Technology, Beverly, MA, USA).

Mouse Assay:

Article Title: Matrix metalloproteinase MMP9 maintains epithelial barrier function and preserves mucosal lining in colitis associated cancer
Article Snippet: As described previously [ ], for WB analysis, colonic mucosal stripping was obtained from the TgM9 and WT mice (n=20 per group) with and without CAC. .. The antibodies used were anti-MMP9 (Abcam, Cambridge, MA), anti-EGFR (Cell Signaling, Beverly, MA), anti-Claudin-2 (Life Technologies, Rockford, IL), anti-Claudin-4 (Invitrogen, Rockford, IL), anti-Claudin-5 (Invitrogen), anti-TFF3 (Cloud-Clone Corp., Katy, TX), anti-Sp1 (Upstate Cell Signaling Solutions, Lake Placid, NY), anti-Occludin (Invitrogen), anti-STAT3 (Cell Signaling).

Chromatin Immunoprecipitation:

Article Title: Overexpression of histone deacetylases in cancer cells is controlled by interplay of transcription factors and epigenetic modulators
Article Snippet: After 48 h, firefly and Renilla luciferase activities were measured using the dual-luciferase reporter assay system (Promega). .. Antibodies for Western blot, immunoprecipitation, and chromatin immunoprecipitation (ChIP) were as follows: anti-HDAC1 and anti-HDAC2 (Pierce); anti-Sp1 (Cell Signaling Technology), anti-Sp3 and anti-p300 (Santa Cruz Biotechnology); anti-p21, anti-acetyl-lysine, anti-ac-H3, anti-H3, anti-H3K4me3, anti-SET1, and anti-RbBP5 (Milipore, Temecula, CA, USA); anti-Ash2L (Bethyl Laboratories, Montgomery, TX, USA); and anti-β-actin and anti-α-tubulin (Sigma-Aldrich, St. Louis, MO, USA). .. HDAC1 and acetylated HDAC1 antibodies were generated as described previously ( ).

Article Title: SP1 and RARα regulate AGAP2 expression in cancer
Article Snippet: Paragraph title: Chromatin immunoprecipitation ... Afterwards, crosslinked proteins of interest were immunoprecipitated with 2 μg of either anti-RARα (C-20), anti-RXRα (D-20) (both from Santa Cruz Biotechnology), anti-PCAF (C14G9, Cell Signaling), or 1 μg of anti-SP1 (D4C3, Cell Signalling).

Article Title: Long noncoding RNA UCA1 induced by SP1 promotes cell proliferation via recruiting EZH2 and activating AKT pathway in gastric cancer
Article Snippet: Paragraph title: ChIP assay ... Antibodies including anti-SP1, anti-EZH2 (Cell Signaling Technology, Danvers, MA, USA) and normal IgG were used for each immunoprecipitation.

Article Title: SP1-induced lncRNA-ZFAS1 contributes to colorectal cancer progression via the miR-150-5p/VEGFA axis
Article Snippet: Paragraph title: Chromatin immunoprecipitation (ChIP) assays ... Antibodies including anti-SP1 (#9389, Cell Signaling Technology, USA) and IgG were employed for each immunoprecipitation. qPCR was used to analyze the precipitated DNA.

Article Title: PPARγ regulated CIDEA affects pro-apoptotic responses in glioblastoma
Article Snippet: Paragraph title: ChIP and ChIP real-time PCR assays ... Anti-NFκ B (Santa Cruz Biotechnology) and anti-SP1 (Cell signaling) were used for immunoprecipitation and non-specific IgG antibody (Abcam) was used as control.

Article Title: TGFβ Signaling Regulates the Timing of CNS Myelination by Modulating Oligodendrocyte Progenitor Cell Cycle Exit through SMAD3/4/FoxO1/Sp1
Article Snippet: Paragraph title: Chromatin immunoprecipitation. ... Immunoprecipitation was performed with anti-SMAD3, anti-SMAD4 (Santa Cruz Biotechnology), anti-Sp1, and anti-FoxO1 (Cell Signaling Technology) antibodies or a nonspecific rabbit IgG as control.

Article Title: Gene expression analysis of human induced pluripotent stem cell-derived neurons carrying copy number variants of chromosome 15q11-q13.1
Article Snippet: A rabbit polyclonal anti-YY-1 (sc-281, Santa Cruz Biotechnology, Inc, Dallas, TX, USA) was used at 5 μg per immunoprecipitation reaction. .. For ChIP analysis of Sp1 binding, the following antibodies were used: mouse monoclonal anti-Sp1 (sc-17824, Santa Cruz Biotechnology, Inc, Dallas, TX, USA) and anti-Sp1 (#9389, Cell Signaling Technology, Beverly, MA, USA). .. Novex Protein A DynaBeads magnetic beads (Life Technologies, Grand Island, NY, USA) were used during immunoprecipitation.

SDS Page:

Article Title: miR-506 attenuates methylation of lncRNA MEG3 to inhibit migration and invasion of breast cancer cell lines via targeting SP1 and SP3
Article Snippet: The lysates were boiled at 100 °C for 5 min and centrifuged at 10,000 rpm for 1 min. About 50 ug of total protein were loaded onto SDS-PAGE gel. .. The following antibodies were used: anti-SP3 (Santa Cruz, USA), anti-DNMT1 (Cell Signaling Technology, USA), anti-SP1 (Cell Signaling Technology, USA), anti-β-actin (Proteintech).

Plasmid Preparation:

Article Title: GRP78‐mediated antioxidant response and ABC transporter activity confers chemoresistance to pancreatic cancer cells
Article Snippet: Slides were stained with anti‐SP1 (Cell Signaling, Cat # 9389) and anti‐GRP78 (Cell Signaling, Cat # 3177) at 1 : 200 dilutions. .. Slides were stained with anti‐SP1 (Cell Signaling, Cat # 9389) and anti‐GRP78 (Cell Signaling, Cat # 3177) at 1 : 200 dilutions.

Software:

Article Title: Matrix metalloproteinase MMP9 maintains epithelial barrier function and preserves mucosal lining in colitis associated cancer
Article Snippet: The antibodies used were anti-MMP9 (Abcam, Cambridge, MA), anti-EGFR (Cell Signaling, Beverly, MA), anti-Claudin-2 (Life Technologies, Rockford, IL), anti-Claudin-4 (Invitrogen, Rockford, IL), anti-Claudin-5 (Invitrogen), anti-TFF3 (Cloud-Clone Corp., Katy, TX), anti-Sp1 (Upstate Cell Signaling Solutions, Lake Placid, NY), anti-Occludin (Invitrogen), anti-STAT3 (Cell Signaling). .. Goat anti-mouse secondary antibody (Bio-Rad, Hercules, CA) or goat anti-rabbit secondary antibody (Bio-Rad) were used.

Article Title: TGFβ Signaling Regulates the Timing of CNS Myelination by Modulating Oligodendrocyte Progenitor Cell Cycle Exit through SMAD3/4/FoxO1/Sp1
Article Snippet: Immunoprecipitation was performed with anti-SMAD3, anti-SMAD4 (Santa Cruz Biotechnology), anti-Sp1, and anti-FoxO1 (Cell Signaling Technology) antibodies or a nonspecific rabbit IgG as control. .. Immunoprecipitation was performed with anti-SMAD3, anti-SMAD4 (Santa Cruz Biotechnology), anti-Sp1, and anti-FoxO1 (Cell Signaling Technology) antibodies or a nonspecific rabbit IgG as control.

SYBR Green Assay:

Article Title: Gene expression analysis of human induced pluripotent stem cell-derived neurons carrying copy number variants of chromosome 15q11-q13.1
Article Snippet: For ChIP analysis of Sp1 binding, the following antibodies were used: mouse monoclonal anti-Sp1 (sc-17824, Santa Cruz Biotechnology, Inc, Dallas, TX, USA) and anti-Sp1 (#9389, Cell Signaling Technology, Beverly, MA, USA). .. Novex Protein A DynaBeads magnetic beads (Life Technologies, Grand Island, NY, USA) were used during immunoprecipitation.

In Vitro:

Article Title: Matrix metalloproteinase MMP9 maintains epithelial barrier function and preserves mucosal lining in colitis associated cancer
Article Snippet: WB analysis for in vitro model was performed using whole cell lysates (30μg/ well) of CaCo2BBE cells with and without MMP9. .. The antibodies used were anti-MMP9 (Abcam, Cambridge, MA), anti-EGFR (Cell Signaling, Beverly, MA), anti-Claudin-2 (Life Technologies, Rockford, IL), anti-Claudin-4 (Invitrogen, Rockford, IL), anti-Claudin-5 (Invitrogen), anti-TFF3 (Cloud-Clone Corp., Katy, TX), anti-Sp1 (Upstate Cell Signaling Solutions, Lake Placid, NY), anti-Occludin (Invitrogen), anti-STAT3 (Cell Signaling).

CCK-8 Assay:

Article Title: Baicalin induces apoptosis in SW480 cells through downregulation of the SP1 transcription factor
Article Snippet: Baicalin and mithramycin-A were procured from Sigma-Aldrich (St Louis, Missouri, USA). .. The annexin V-FITC and CCK-8 kits for detecting apoptosis were obtained from Beyotime (Shanghai, China), the RNAiso Plus reagent kit and the PrimeScript RT reagent kit (Perfect Real Time) were obtained from TAKARA, BIO (Kusatsu, Japan), whereas anti-sp1 , anti-C-PARP, anti-C-caspase-3, and tubulin antibodies were obtained from Cell Signaling Technology (Danvers, Massachusetts, USA). .. The gene expression profiles of patients with colorectal cancer were identified from the PubMed GEO datasets using the keywords ‘colorectal cancer’ and ‘gene expression profiling’.

Immunoprecipitation:

Article Title: Overexpression of histone deacetylases in cancer cells is controlled by interplay of transcription factors and epigenetic modulators
Article Snippet: After 48 h, firefly and Renilla luciferase activities were measured using the dual-luciferase reporter assay system (Promega). .. Antibodies for Western blot, immunoprecipitation, and chromatin immunoprecipitation (ChIP) were as follows: anti-HDAC1 and anti-HDAC2 (Pierce); anti-Sp1 (Cell Signaling Technology), anti-Sp3 and anti-p300 (Santa Cruz Biotechnology); anti-p21, anti-acetyl-lysine, anti-ac-H3, anti-H3, anti-H3K4me3, anti-SET1, and anti-RbBP5 (Milipore, Temecula, CA, USA); anti-Ash2L (Bethyl Laboratories, Montgomery, TX, USA); and anti-β-actin and anti-α-tubulin (Sigma-Aldrich, St. Louis, MO, USA). .. HDAC1 and acetylated HDAC1 antibodies were generated as described previously ( ).

Article Title: SP1 and RARα regulate AGAP2 expression in cancer
Article Snippet: The chromatin was sheared to 100–600 bp by sonication using an AFA Focused-Ultrasonicator (S220- Series from CovarisTM ) at 6 × 60 second on-off pulses at 4 °C. .. Afterwards, crosslinked proteins of interest were immunoprecipitated with 2 μg of either anti-RARα (C-20), anti-RXRα (D-20) (both from Santa Cruz Biotechnology), anti-PCAF (C14G9, Cell Signaling), or 1 μg of anti-SP1 (D4C3, Cell Signalling). .. Anti-Pol II (N-20, Santa Cruz Biotechnology) was used as positive control and rabbit IgG (Invitrogen) as negative control.

Article Title: Long noncoding RNA UCA1 induced by SP1 promotes cell proliferation via recruiting EZH2 and activating AKT pathway in gastric cancer
Article Snippet: Briefly, GC cells were cross-linked in 1% formaldehyde solution for 10 min at room temperature, followed by the addition of 125 mM of glycine for 5 min. DNA fragments ranging from 200 to 500 bp were yielded via sonication. .. Antibodies including anti-SP1, anti-EZH2 (Cell Signaling Technology, Danvers, MA, USA) and normal IgG were used for each immunoprecipitation. .. Immunoprecipitated and input DNAs were subjected to qRT-PCR analysis.

Article Title: SP1-induced lncRNA-ZFAS1 contributes to colorectal cancer progression via the miR-150-5p/VEGFA axis
Article Snippet: Briefly, the CRC cells were fixed with 1% formaldehyde solution for 15 min and incubated with 125 nM glycine for 5 min. DNA fragments ranging from 200 to 300 bp were generated using sonication. .. Antibodies including anti-SP1 (#9389, Cell Signaling Technology, USA) and IgG were employed for each immunoprecipitation. qPCR was used to analyze the precipitated DNA. .. RIP assay was performed with Magna RIPTM RNA-Binding Protein Immunoprecipation Kit (Millipore, Billerica, USA) and AGO2 antibody (proteintech, 10686–1-AP, China) in accordance with the manufacturer’s protocol.

Article Title: PPARγ regulated CIDEA affects pro-apoptotic responses in glioblastoma
Article Snippet: ChIP was performed on glioma cells treated with PPARγ inhibitor for 48 h by Chip-IT Enzymatic DNA shearing Kit (Active Motif, Carlsbad, CA, USA) as described previously. .. Anti-NFκ B (Santa Cruz Biotechnology) and anti-SP1 (Cell signaling) were used for immunoprecipitation and non-specific IgG antibody (Abcam) was used as control. .. After reverse cross-linking and DNA purification, DNA from input (1:10 diluted) or immune-precipitated (IP) samples were quantified by real-time PCR using ABI 7500 real-time thermal cycler with Power SYBR green PCR Master Mix (Life Technologies, Invitrogen) for 40 cycles.

Article Title: TGFβ Signaling Regulates the Timing of CNS Myelination by Modulating Oligodendrocyte Progenitor Cell Cycle Exit through SMAD3/4/FoxO1/Sp1
Article Snippet: CNP -EGFP FACS-sorted cells were cultured under proliferating conditions and were treated with TGFβ1 or vehicle and 4 h later samples were processed as recommended by the manufacturer (EZ-ChIP; Millipore). .. Immunoprecipitation was performed with anti-SMAD3, anti-SMAD4 (Santa Cruz Biotechnology), anti-Sp1, and anti-FoxO1 (Cell Signaling Technology) antibodies or a nonspecific rabbit IgG as control. .. The presence of DNA bound to each transcription factor (TF) from the indicated regions of the two promoters analyzed was determined by qPCR analysis as above.

Article Title: Gene expression analysis of human induced pluripotent stem cell-derived neurons carrying copy number variants of chromosome 15q11-q13.1
Article Snippet: A rabbit polyclonal anti-YY-1 (sc-281, Santa Cruz Biotechnology, Inc, Dallas, TX, USA) was used at 5 μg per immunoprecipitation reaction. .. For ChIP analysis of Sp1 binding, the following antibodies were used: mouse monoclonal anti-Sp1 (sc-17824, Santa Cruz Biotechnology, Inc, Dallas, TX, USA) and anti-Sp1 (#9389, Cell Signaling Technology, Beverly, MA, USA).

Fractionation:

Article Title: Role of Cdc6 in re-replication in cells expressing human papillomavirus E7 oncogene
Article Snippet: Paragraph title: Subcellular fractionation ... The purity of the obtained fractions was confirmed using anti-β-tubulin (Sigma, T-4026, for the CEs), anti-Sp1 (Cell Signaling #9389, for the SNEs) or anti-Histone H3 (Cell Signaling #3688, for the CBEs).

Staining:

Article Title: GRP78‐mediated antioxidant response and ABC transporter activity confers chemoresistance to pancreatic cancer cells
Article Snippet: Tissues were then blocked with Dako protein block and incubated with primary antibody overnight. .. Slides were stained with anti‐SP1 (Cell Signaling, Cat # 9389) and anti‐GRP78 (Cell Signaling, Cat # 3177) at 1 : 200 dilutions. .. Slides were washed with PBS, incubated with secondary anti‐rabbit antibody, and conjugated to horseradish peroxidase, for 30 min.

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    Cell Signaling Technology Inc anti sp1
    HDAC1 and HDAC2 promoter activity is regulated by <t>Sp1</t> and Sp3 in colon cancer cell lines. A ) Analysis of HDAC1 and HDAC2 mRNA expression by qRT-PCR in HCT116, HT29, and FHs 74 Int cells. HDAC1 and HDAC2 mRNA expression is shown relative to the control
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    HDAC1 and HDAC2 promoter activity is regulated by Sp1 and Sp3 in colon cancer cell lines. A ) Analysis of HDAC1 and HDAC2 mRNA expression by qRT-PCR in HCT116, HT29, and FHs 74 Int cells. HDAC1 and HDAC2 mRNA expression is shown relative to the control

    Journal:

    Article Title: Overexpression of histone deacetylases in cancer cells is controlled by interplay of transcription factors and epigenetic modulators

    doi: 10.1096/fj.14-250654

    Figure Lengend Snippet: HDAC1 and HDAC2 promoter activity is regulated by Sp1 and Sp3 in colon cancer cell lines. A ) Analysis of HDAC1 and HDAC2 mRNA expression by qRT-PCR in HCT116, HT29, and FHs 74 Int cells. HDAC1 and HDAC2 mRNA expression is shown relative to the control

    Article Snippet: Antibodies for Western blot, immunoprecipitation, and chromatin immunoprecipitation (ChIP) were as follows: anti-HDAC1 and anti-HDAC2 (Pierce); anti-Sp1 (Cell Signaling Technology), anti-Sp3 and anti-p300 (Santa Cruz Biotechnology); anti-p21, anti-acetyl-lysine, anti-ac-H3, anti-H3, anti-H3K4me3, anti-SET1, and anti-RbBP5 (Milipore, Temecula, CA, USA); anti-Ash2L (Bethyl Laboratories, Montgomery, TX, USA); and anti-β-actin and anti-α-tubulin (Sigma-Aldrich, St. Louis, MO, USA).

    Techniques: Activity Assay, Expressing, Quantitative RT-PCR

    Sp1 or Sp3 silencing affects cell growth and colony formation in colon cancer cells. A ) Effect of Sp1 or Sp3 silencing on cell proliferation. HCT116 cells (1×10 4 ) with or without Sp1 or Sp3 KD were seeded and the cell numbers were counted every

    Journal:

    Article Title: Overexpression of histone deacetylases in cancer cells is controlled by interplay of transcription factors and epigenetic modulators

    doi: 10.1096/fj.14-250654

    Figure Lengend Snippet: Sp1 or Sp3 silencing affects cell growth and colony formation in colon cancer cells. A ) Effect of Sp1 or Sp3 silencing on cell proliferation. HCT116 cells (1×10 4 ) with or without Sp1 or Sp3 KD were seeded and the cell numbers were counted every

    Article Snippet: Antibodies for Western blot, immunoprecipitation, and chromatin immunoprecipitation (ChIP) were as follows: anti-HDAC1 and anti-HDAC2 (Pierce); anti-Sp1 (Cell Signaling Technology), anti-Sp3 and anti-p300 (Santa Cruz Biotechnology); anti-p21, anti-acetyl-lysine, anti-ac-H3, anti-H3, anti-H3K4me3, anti-SET1, and anti-RbBP5 (Milipore, Temecula, CA, USA); anti-Ash2L (Bethyl Laboratories, Montgomery, TX, USA); and anti-β-actin and anti-α-tubulin (Sigma-Aldrich, St. Louis, MO, USA).

    Techniques:

    Effect of Sp1 on recruitment of other coregulators on HDAC1 promoter. A ) Levels of p300 or other indicated proteins in control or Sp1 KD cells were determined by Western blot. NC, negative control of scramble shRNA. β-Actin was used as a loading

    Journal:

    Article Title: Overexpression of histone deacetylases in cancer cells is controlled by interplay of transcription factors and epigenetic modulators

    doi: 10.1096/fj.14-250654

    Figure Lengend Snippet: Effect of Sp1 on recruitment of other coregulators on HDAC1 promoter. A ) Levels of p300 or other indicated proteins in control or Sp1 KD cells were determined by Western blot. NC, negative control of scramble shRNA. β-Actin was used as a loading

    Article Snippet: Antibodies for Western blot, immunoprecipitation, and chromatin immunoprecipitation (ChIP) were as follows: anti-HDAC1 and anti-HDAC2 (Pierce); anti-Sp1 (Cell Signaling Technology), anti-Sp3 and anti-p300 (Santa Cruz Biotechnology); anti-p21, anti-acetyl-lysine, anti-ac-H3, anti-H3, anti-H3K4me3, anti-SET1, and anti-RbBP5 (Milipore, Temecula, CA, USA); anti-Ash2L (Bethyl Laboratories, Montgomery, TX, USA); and anti-β-actin and anti-α-tubulin (Sigma-Aldrich, St. Louis, MO, USA).

    Techniques: Western Blot, Negative Control, shRNA

    Sp1 and Sp3 affect HDAC1 and HDAC2 expression in colorectal cancer cell lines. A ) Levels of HDAC1 and other indicated proteins in control and Sp1 or Sp3 KD cells were determined by Western blot. NC, negative control of scramble shRNA. β-Actin

    Journal:

    Article Title: Overexpression of histone deacetylases in cancer cells is controlled by interplay of transcription factors and epigenetic modulators

    doi: 10.1096/fj.14-250654

    Figure Lengend Snippet: Sp1 and Sp3 affect HDAC1 and HDAC2 expression in colorectal cancer cell lines. A ) Levels of HDAC1 and other indicated proteins in control and Sp1 or Sp3 KD cells were determined by Western blot. NC, negative control of scramble shRNA. β-Actin

    Article Snippet: Antibodies for Western blot, immunoprecipitation, and chromatin immunoprecipitation (ChIP) were as follows: anti-HDAC1 and anti-HDAC2 (Pierce); anti-Sp1 (Cell Signaling Technology), anti-Sp3 and anti-p300 (Santa Cruz Biotechnology); anti-p21, anti-acetyl-lysine, anti-ac-H3, anti-H3, anti-H3K4me3, anti-SET1, and anti-RbBP5 (Milipore, Temecula, CA, USA); anti-Ash2L (Bethyl Laboratories, Montgomery, TX, USA); and anti-β-actin and anti-α-tubulin (Sigma-Aldrich, St. Louis, MO, USA).

    Techniques: Expressing, Western Blot, Negative Control, shRNA

    Sp1 and Sp3 are overexpressed and bind to HDAC1 and HDAC2 promoters in colon cancer. A ) Levels of Sp1 and Sp3 proteins from various colon cancer cell lines were determined by Western blotting. Vaco250 and Vaco330 are human adenoma cell lines and used

    Journal:

    Article Title: Overexpression of histone deacetylases in cancer cells is controlled by interplay of transcription factors and epigenetic modulators

    doi: 10.1096/fj.14-250654

    Figure Lengend Snippet: Sp1 and Sp3 are overexpressed and bind to HDAC1 and HDAC2 promoters in colon cancer. A ) Levels of Sp1 and Sp3 proteins from various colon cancer cell lines were determined by Western blotting. Vaco250 and Vaco330 are human adenoma cell lines and used

    Article Snippet: Antibodies for Western blot, immunoprecipitation, and chromatin immunoprecipitation (ChIP) were as follows: anti-HDAC1 and anti-HDAC2 (Pierce); anti-Sp1 (Cell Signaling Technology), anti-Sp3 and anti-p300 (Santa Cruz Biotechnology); anti-p21, anti-acetyl-lysine, anti-ac-H3, anti-H3, anti-H3K4me3, anti-SET1, and anti-RbBP5 (Milipore, Temecula, CA, USA); anti-Ash2L (Bethyl Laboratories, Montgomery, TX, USA); and anti-β-actin and anti-α-tubulin (Sigma-Aldrich, St. Louis, MO, USA).

    Techniques: Western Blot

    Effect of p300 KD on epigenetic markers and Sp1 binding on HDAC1 promoter. A ) Levels of HDAC1 and other indicated proteins in control or p300 KD cells were determined by Western blot. NC, negative control of scramble shRNA. β-Actin was used as

    Journal:

    Article Title: Overexpression of histone deacetylases in cancer cells is controlled by interplay of transcription factors and epigenetic modulators

    doi: 10.1096/fj.14-250654

    Figure Lengend Snippet: Effect of p300 KD on epigenetic markers and Sp1 binding on HDAC1 promoter. A ) Levels of HDAC1 and other indicated proteins in control or p300 KD cells were determined by Western blot. NC, negative control of scramble shRNA. β-Actin was used as

    Article Snippet: Antibodies for Western blot, immunoprecipitation, and chromatin immunoprecipitation (ChIP) were as follows: anti-HDAC1 and anti-HDAC2 (Pierce); anti-Sp1 (Cell Signaling Technology), anti-Sp3 and anti-p300 (Santa Cruz Biotechnology); anti-p21, anti-acetyl-lysine, anti-ac-H3, anti-H3, anti-H3K4me3, anti-SET1, and anti-RbBP5 (Milipore, Temecula, CA, USA); anti-Ash2L (Bethyl Laboratories, Montgomery, TX, USA); and anti-β-actin and anti-α-tubulin (Sigma-Aldrich, St. Louis, MO, USA).

    Techniques: Binding Assay, Western Blot, Negative Control, shRNA

    Effect of Sp1 on recruitment of other regulators on the HDAC2 promoter. ChIP assays were carried out in HCT116 cells with Sp1 KD by using indicated antibodies: ac-H3 ( A ), H3K4 trimethylation ( B ), p300 ( C ), and SET1 ( D ). The resulting DNA was subjected

    Journal:

    Article Title: Overexpression of histone deacetylases in cancer cells is controlled by interplay of transcription factors and epigenetic modulators

    doi: 10.1096/fj.14-250654

    Figure Lengend Snippet: Effect of Sp1 on recruitment of other regulators on the HDAC2 promoter. ChIP assays were carried out in HCT116 cells with Sp1 KD by using indicated antibodies: ac-H3 ( A ), H3K4 trimethylation ( B ), p300 ( C ), and SET1 ( D ). The resulting DNA was subjected

    Article Snippet: Antibodies for Western blot, immunoprecipitation, and chromatin immunoprecipitation (ChIP) were as follows: anti-HDAC1 and anti-HDAC2 (Pierce); anti-Sp1 (Cell Signaling Technology), anti-Sp3 and anti-p300 (Santa Cruz Biotechnology); anti-p21, anti-acetyl-lysine, anti-ac-H3, anti-H3, anti-H3K4me3, anti-SET1, and anti-RbBP5 (Milipore, Temecula, CA, USA); anti-Ash2L (Bethyl Laboratories, Montgomery, TX, USA); and anti-β-actin and anti-α-tubulin (Sigma-Aldrich, St. Louis, MO, USA).

    Techniques: Chromatin Immunoprecipitation

    Overexpression of Sp1 impairs miR-29c-induced inhibition of EMT The mRNA and protein expression of TGF-β1-induced EMT-associated markers including TTF, E-cadherin, vimentin and α-SMA were analyzed in ( A – B ) 95C and ( C – D ) A549 cells with ectopically expressing miR-29c mimics or pcDNA-Sp1. * p

    Journal: Oncotarget

    Article Title: A regulatory loop involving miR-29c and Sp1 elevates the TGF-β1 mediated epithelial-to-mesenchymal transition in lung cancer

    doi: 10.18632/oncotarget.13137

    Figure Lengend Snippet: Overexpression of Sp1 impairs miR-29c-induced inhibition of EMT The mRNA and protein expression of TGF-β1-induced EMT-associated markers including TTF, E-cadherin, vimentin and α-SMA were analyzed in ( A – B ) 95C and ( C – D ) A549 cells with ectopically expressing miR-29c mimics or pcDNA-Sp1. * p

    Article Snippet: E-Cadherin, TIF-1, vimentin, a-SMA and sp1 antibodies were obtained from Cell Signaling Tech (Denver, MA).

    Techniques: Over Expression, Inhibition, Expressing

    Summary diagram describes the miR-29c/Sp1 network that regulates TGF-β1 expression and TGF-β1-induced EMT

    Journal: Oncotarget

    Article Title: A regulatory loop involving miR-29c and Sp1 elevates the TGF-β1 mediated epithelial-to-mesenchymal transition in lung cancer

    doi: 10.18632/oncotarget.13137

    Figure Lengend Snippet: Summary diagram describes the miR-29c/Sp1 network that regulates TGF-β1 expression and TGF-β1-induced EMT

    Article Snippet: E-Cadherin, TIF-1, vimentin, a-SMA and sp1 antibodies were obtained from Cell Signaling Tech (Denver, MA).

    Techniques: Expressing

    Inhibition of miR-29c significantly elevates the migration and invasion ( A ) The expression of mir-29c and Sp1 in TGF-β1-treated cells. Transwell assay was performed and ( B – C ) the migration and invasion of 95C and A549 cells transfected with miR-29c inhibitors were determined. ( D ) The evaluation of transfection efficiency of miR-29c inhibitors. * p

    Journal: Oncotarget

    Article Title: A regulatory loop involving miR-29c and Sp1 elevates the TGF-β1 mediated epithelial-to-mesenchymal transition in lung cancer

    doi: 10.18632/oncotarget.13137

    Figure Lengend Snippet: Inhibition of miR-29c significantly elevates the migration and invasion ( A ) The expression of mir-29c and Sp1 in TGF-β1-treated cells. Transwell assay was performed and ( B – C ) the migration and invasion of 95C and A549 cells transfected with miR-29c inhibitors were determined. ( D ) The evaluation of transfection efficiency of miR-29c inhibitors. * p

    Article Snippet: E-Cadherin, TIF-1, vimentin, a-SMA and sp1 antibodies were obtained from Cell Signaling Tech (Denver, MA).

    Techniques: Inhibition, Migration, Expressing, Transwell Assay, Transfection

    Sp1 could restore the miR-29c-induced inhibition of metastasis ( A – B ) Overexpression of Sp1 by transfection of pcDNA-Sp1 into 95C and A549 cells. Transwell assay was performed and ( C – D ) the migration and invasion of 95C and A549 cells transfected with miR-29c mimics or pcDNA-Sp1 were determined.

    Journal: Oncotarget

    Article Title: A regulatory loop involving miR-29c and Sp1 elevates the TGF-β1 mediated epithelial-to-mesenchymal transition in lung cancer

    doi: 10.18632/oncotarget.13137

    Figure Lengend Snippet: Sp1 could restore the miR-29c-induced inhibition of metastasis ( A – B ) Overexpression of Sp1 by transfection of pcDNA-Sp1 into 95C and A549 cells. Transwell assay was performed and ( C – D ) the migration and invasion of 95C and A549 cells transfected with miR-29c mimics or pcDNA-Sp1 were determined.

    Article Snippet: E-Cadherin, TIF-1, vimentin, a-SMA and sp1 antibodies were obtained from Cell Signaling Tech (Denver, MA).

    Techniques: Inhibition, Over Expression, Transfection, Transwell Assay, Migration

    MiR-29c could inhibit Sp1/TGF-β1 expression and lung cancer progression ( A ) The TGF-β1 expression in overexpression of miR-29c mimics or negative control was analyzed by Q-PCR in 95C cells with or without the treatment of TGF-β1. ( B – C ) The TGF-β1 expression in overexpression of siRNA-Sp1or negative control was analyzed by Q-PCR in 95C cells with or without the treatment of TGF-β1. ( D ) The relative luciferase activities in 95C cells were determined after the pGL-3-TGFB1 plasmids were transfected with miR-29c mimics. ( E – F ) The relative luciferase activities in 95C cells were determined after the pGL-3-TGFB1 plasmids were transfected with siRNA-Sp1 or pcDNA-Sp1. ( G ) The relative luciferase activities in 95C cells were determined after the pGL-3-TGFB1 plasmids were transfected with pcDNA-Sp1 and miR-29c mimics. ( H – I ) Nude mice (6 per group) were subcutaneously injected of 3 × 10 6 A549 cells and the miR-29c mimics, inhibitor or negative control (10 nM per injection) were delivered via intra-tumoral injection for six times, three days apart. The tumor volume (mm 3 ) and body weight (g) were measured. ( J – K ) The levelS of Sp1 and TGF-β1 in tumor tissues were estimated. * p

    Journal: Oncotarget

    Article Title: A regulatory loop involving miR-29c and Sp1 elevates the TGF-β1 mediated epithelial-to-mesenchymal transition in lung cancer

    doi: 10.18632/oncotarget.13137

    Figure Lengend Snippet: MiR-29c could inhibit Sp1/TGF-β1 expression and lung cancer progression ( A ) The TGF-β1 expression in overexpression of miR-29c mimics or negative control was analyzed by Q-PCR in 95C cells with or without the treatment of TGF-β1. ( B – C ) The TGF-β1 expression in overexpression of siRNA-Sp1or negative control was analyzed by Q-PCR in 95C cells with or without the treatment of TGF-β1. ( D ) The relative luciferase activities in 95C cells were determined after the pGL-3-TGFB1 plasmids were transfected with miR-29c mimics. ( E – F ) The relative luciferase activities in 95C cells were determined after the pGL-3-TGFB1 plasmids were transfected with siRNA-Sp1 or pcDNA-Sp1. ( G ) The relative luciferase activities in 95C cells were determined after the pGL-3-TGFB1 plasmids were transfected with pcDNA-Sp1 and miR-29c mimics. ( H – I ) Nude mice (6 per group) were subcutaneously injected of 3 × 10 6 A549 cells and the miR-29c mimics, inhibitor or negative control (10 nM per injection) were delivered via intra-tumoral injection for six times, three days apart. The tumor volume (mm 3 ) and body weight (g) were measured. ( J – K ) The levelS of Sp1 and TGF-β1 in tumor tissues were estimated. * p

    Article Snippet: E-Cadherin, TIF-1, vimentin, a-SMA and sp1 antibodies were obtained from Cell Signaling Tech (Denver, MA).

    Techniques: Expressing, Over Expression, Negative Control, Polymerase Chain Reaction, Luciferase, Transfection, Mouse Assay, Injection

    MiR-29c targets Sp1 and is down-regulated in high-metastatic lung cancer cell lines ( A – B ) The level of miR-29c and Sp1 in lung cancer tissues and ( C – D ) cell lines including BEAS-2B, the paired low-metastatic 95C and high-metastatic 95D and A549 were determined by Q-PCR and Western blotting. 95C cell line was transfected with miR-29c mimics or negative control. ( E – F ) the mRNA and protein level of Sp1 were determined. ( G – H ) The luciferase reporter was performed to confirm the direct target sites. ** p

    Journal: Oncotarget

    Article Title: A regulatory loop involving miR-29c and Sp1 elevates the TGF-β1 mediated epithelial-to-mesenchymal transition in lung cancer

    doi: 10.18632/oncotarget.13137

    Figure Lengend Snippet: MiR-29c targets Sp1 and is down-regulated in high-metastatic lung cancer cell lines ( A – B ) The level of miR-29c and Sp1 in lung cancer tissues and ( C – D ) cell lines including BEAS-2B, the paired low-metastatic 95C and high-metastatic 95D and A549 were determined by Q-PCR and Western blotting. 95C cell line was transfected with miR-29c mimics or negative control. ( E – F ) the mRNA and protein level of Sp1 were determined. ( G – H ) The luciferase reporter was performed to confirm the direct target sites. ** p

    Article Snippet: E-Cadherin, TIF-1, vimentin, a-SMA and sp1 antibodies were obtained from Cell Signaling Tech (Denver, MA).

    Techniques: Polymerase Chain Reaction, Western Blot, Transfection, Negative Control, Luciferase

    p16 modulates the expression of different Sp1 targets. A and C , total RNA was prepared from the indicated cells, and quantitative RT-PCR was performed using specific primers for the indicated genes. B , EMSA was performed using nuclear extracts from the

    Journal:

    Article Title: The Cyclin-dependent Kinase Inhibitor p16INK4

    doi: 10.1074/jbc.M113.512640

    Figure Lengend Snippet: p16 modulates the expression of different Sp1 targets. A and C , total RNA was prepared from the indicated cells, and quantitative RT-PCR was performed using specific primers for the indicated genes. B , EMSA was performed using nuclear extracts from the

    Article Snippet: ChIP experiments were performed using antibodies against Sp1 (Cell Signaling Technology), p16 (BD Biosciences), and CDK4 (C-22).

    Techniques: Expressing, Quantitative RT-PCR

    p16 positively regulates the expression of miR-141 and miR-146b-5p through the Sp1 transcription factor. A , nucleotide sequences of the miR-141 and miR-146b-5p promoters, −39 to +9 nucleotides and −93 to −26 nucleotides, respectively,

    Journal:

    Article Title: The Cyclin-dependent Kinase Inhibitor p16INK4

    doi: 10.1074/jbc.M113.512640

    Figure Lengend Snippet: p16 positively regulates the expression of miR-141 and miR-146b-5p through the Sp1 transcription factor. A , nucleotide sequences of the miR-141 and miR-146b-5p promoters, −39 to +9 nucleotides and −93 to −26 nucleotides, respectively,

    Article Snippet: ChIP experiments were performed using antibodies against Sp1 (Cell Signaling Technology), p16 (BD Biosciences), and CDK4 (C-22).

    Techniques: Expressing

    p16, Sp1, and CDK4 proteins form a heterocomplex that binds the miR-141 and miR-146b-5p promoters. A , whole cell extracts were prepared from HFSN1 cells and immunoprecipitated ( IP ) with anti-p16, anti-Sp1, or anti-CDK4 antibodies (mouse IgG was used as

    Journal:

    Article Title: The Cyclin-dependent Kinase Inhibitor p16INK4

    doi: 10.1074/jbc.M113.512640

    Figure Lengend Snippet: p16, Sp1, and CDK4 proteins form a heterocomplex that binds the miR-141 and miR-146b-5p promoters. A , whole cell extracts were prepared from HFSN1 cells and immunoprecipitated ( IP ) with anti-p16, anti-Sp1, or anti-CDK4 antibodies (mouse IgG was used as

    Article Snippet: ChIP experiments were performed using antibodies against Sp1 (Cell Signaling Technology), p16 (BD Biosciences), and CDK4 (C-22).

    Techniques: Immunoprecipitation

    p16 interacts with Sp1 through the fourth ankyrin repeat. A , p16-negative U2OS cells were transfected with the indicated constructs, and then whole cell extracts were prepared and used for immunoprecipitation ( IP ) with anti-p16 antibody (mouse IgG was

    Journal:

    Article Title: The Cyclin-dependent Kinase Inhibitor p16INK4

    doi: 10.1074/jbc.M113.512640

    Figure Lengend Snippet: p16 interacts with Sp1 through the fourth ankyrin repeat. A , p16-negative U2OS cells were transfected with the indicated constructs, and then whole cell extracts were prepared and used for immunoprecipitation ( IP ) with anti-p16 antibody (mouse IgG was

    Article Snippet: ChIP experiments were performed using antibodies against Sp1 (Cell Signaling Technology), p16 (BD Biosciences), and CDK4 (C-22).

    Techniques: Transfection, Construct, Immunoprecipitation

    Sp1 binds the miR-141 and miR-146b-5p promoters and activates their transcription in a p16-dependent manner. A , ChIP assay. Chromatin was purified from HFSN1C and HFSN1p16sh, and then immunoprecipitated ( IP ) using anti-Sp1 antibody. miR-141 and miR-146b-5p

    Journal:

    Article Title: The Cyclin-dependent Kinase Inhibitor p16INK4

    doi: 10.1074/jbc.M113.512640

    Figure Lengend Snippet: Sp1 binds the miR-141 and miR-146b-5p promoters and activates their transcription in a p16-dependent manner. A , ChIP assay. Chromatin was purified from HFSN1C and HFSN1p16sh, and then immunoprecipitated ( IP ) using anti-Sp1 antibody. miR-141 and miR-146b-5p

    Article Snippet: ChIP experiments were performed using antibodies against Sp1 (Cell Signaling Technology), p16 (BD Biosciences), and CDK4 (C-22).

    Techniques: Chromatin Immunoprecipitation, Purification, Immunoprecipitation

    p16-CDK4-Sp1-dependent up-regulation of miR-141 and miR-146b-5p and their role in apoptosis following UV damage. A , cells were either mock-treated or challenged with UV light (10 Jm −2 ) and then re-incubated for the indicated periods of time. Total

    Journal:

    Article Title: The Cyclin-dependent Kinase Inhibitor p16INK4

    doi: 10.1074/jbc.M113.512640

    Figure Lengend Snippet: p16-CDK4-Sp1-dependent up-regulation of miR-141 and miR-146b-5p and their role in apoptosis following UV damage. A , cells were either mock-treated or challenged with UV light (10 Jm −2 ) and then re-incubated for the indicated periods of time. Total

    Article Snippet: ChIP experiments were performed using antibodies against Sp1 (Cell Signaling Technology), p16 (BD Biosciences), and CDK4 (C-22).

    Techniques: Incubation