srf  (Santa Cruz Biotechnology)

Journal: International journal of biological sciences

Article Title: The Antitumor and Sorafenib-resistant Reversal Effects of Ursolic Acid on Hepatocellular Carcinoma via Targeting ING5.

doi: 10.7150/ijbs.97720

Figure Lengend Snippet: Figure 5. UA abrogated ING5 transcription in hepatocellular carcinoma cells by reducing SRF and YY1 expression and disassociating the SRF-YY1 complex. (A) SRF and YY1 expressions was analyzed in hepatocellular carcinoma (HCC) and normal liver tissues using TCGA-LIHC and TCGA-CPTAC data from UALCAN (***P < 0.001). (B) The prognostic significance of SRF and YY1 mRNA expression in HCC was studied using Xiantao (https://www.xiantaozi.com/) (HR, hazard ratio). (C) The correlations among SRF, YY1 and ING5 were assessed by R software. (D) HepG2 cells were exposed to the culture medium containing 2% and 12% FBS for 36 h, respectively (left). HepG2 cells were transfected with SRF siRNA for 24 h following exposure to medium containing 12% FBS for 36 h (middle). HepG2 cells were transfected with YY1 siRNA for 24 h following exposure to the medium containing 12% FBS for 36 h (right). The expression levels of SRF, ING5 and PCNA were determined by western blotting. (E) HepG2 cells were exposed to the culture medium containing 2% and 12% FBS for 36 h. HepG2 cells were transfected with ING5 shRNA, SRF or YY1 siRNA following exposure to the medium containing 12% FBS for 36 h. HepG2 cells were transfected with ING5 overexpression plasmid following exposure to the medium containing 2% FBS for 36 h. CCK-8 assays were performed for assessing the proliferation of HepG2 cells (n=3, *P < 0.05, **P < 0.01, ***P < 0.001). (F) qRT‒PCR assays were performed to assess the expression of ING5 mRNA in HepG2 cells treated with 5 μM UA for 24 h (n=3, *P < 0.05 vs. the control group). (G) HepG2 cells were individually transfected with the pGL3-basic reporter

Article Snippet: ACC1, ACLY and T antigen overexpression and shRNA plasmids (HonorGene, Changsha, China) and SRF (sc-36563, Santa Cruz, CA, USA) and YY1 (sc-36863, Santa Cruz, CA, USA) siRNA plasmids were transfected into cells using a Lipofectamine 3000 transfection kit (L3000015, Invitrogen, USA).

Techniques: Expressing, Software, Transfection, Western Blot, shRNA, Over Expression, Plasmid Preparation, CCK-8 Assay, Control

Journal: International journal of biological sciences

Article Title: The Antitumor and Sorafenib-resistant Reversal Effects of Ursolic Acid on Hepatocellular Carcinoma via Targeting ING5.

doi: 10.7150/ijbs.97720

Figure Lengend Snippet: Figure 7. T antigen upregulated ING5 expression by inhibiting ubiquitin-mediated degradation and promoting the T antigen-SRF-YY1-ING5 complex-associated transcription. (A) Mouse primary HCC cells (upper) and Hepa1-6 cells (lower) were treated with CHX (0.5 μg/ml) and subjected to western blot analysis of ING5 expression. (B) Mouse primary HCC cells and Hepa1-6 cells were exposed to MG132 (10 μg/ml) for 6 h, and T antigen and ING5 expression was measured by Western blotting. (C) Cells were treated with MG132 (10 μg/ml) for 6 h, and then ING5 ubiquitination was assessed by Co-IP assays. (D) qRT‒PCR assays were performed to assess ING5 mRNA expression after silencing or overexpressing T antigen. (E) Cells were individually transfected with the pGL3-ING5 containing WT or MUT SRF/YY1 binding site for 36 h, and subjected to a dual-luciferase reporter assay. (F) ChIP assays with anti-SRF, anti-YY1 or anti-T antigen antibodies were used to measure their binding to the promoter of ING5. (G) Co-IP assays with anti-T antigen, anti-SRF, anti-YY1, and anti-ING5 antibodies were conducted to observe their interactions. Data are presented mean ± SD, and Student’s t test was used for significance tests. n.s.=not significant, *P < 0.01, **P < 0.001 vs. vector group. WT, wild-type. MUT, mutant. UB, ubiquitin. IP, immunoprecipitation.

Article Snippet: ACC1, ACLY and T antigen overexpression and shRNA plasmids (HonorGene, Changsha, China) and SRF (sc-36563, Santa Cruz, CA, USA) and YY1 (sc-36863, Santa Cruz, CA, USA) siRNA plasmids were transfected into cells using a Lipofectamine 3000 transfection kit (L3000015, Invitrogen, USA).

Techniques: Expressing, Ubiquitin Proteomics, Western Blot, Co-Immunoprecipitation Assay, Transfection, Binding Assay, Luciferase, Reporter Assay, Plasmid Preparation, Mutagenesis, Immunoprecipitation

Journal: International journal of biological sciences

Article Title: The Antitumor and Sorafenib-resistant Reversal Effects of Ursolic Acid on Hepatocellular Carcinoma via Targeting ING5.

doi: 10.7150/ijbs.97720

Figure Lengend Snippet: Figure 5. UA abrogated ING5 transcription in hepatocellular carcinoma cells by reducing SRF and YY1 expression and disassociating the SRF-YY1 complex. (A) SRF and YY1 expressions was analyzed in hepatocellular carcinoma (HCC) and normal liver tissues using TCGA-LIHC and TCGA-CPTAC data from UALCAN (***P < 0.001). (B) The prognostic significance of SRF and YY1 mRNA expression in HCC was studied using Xiantao (https://www.xiantaozi.com/) (HR, hazard ratio). (C) The correlations among SRF, YY1 and ING5 were assessed by R software. (D) HepG2 cells were exposed to the culture medium containing 2% and 12% FBS for 36 h, respectively (left). HepG2 cells were transfected with SRF siRNA for 24 h following exposure to medium containing 12% FBS for 36 h (middle). HepG2 cells were transfected with YY1 siRNA for 24 h following exposure to the medium containing 12% FBS for 36 h (right). The expression levels of SRF, ING5 and PCNA were determined by western blotting. (E) HepG2 cells were exposed to the culture medium containing 2% and 12% FBS for 36 h. HepG2 cells were transfected with ING5 shRNA, SRF or YY1 siRNA following exposure to the medium containing 12% FBS for 36 h. HepG2 cells were transfected with ING5 overexpression plasmid following exposure to the medium containing 2% FBS for 36 h. CCK-8 assays were performed for assessing the proliferation of HepG2 cells (n=3, *P < 0.05, **P < 0.01, ***P < 0.001). (F) qRT‒PCR assays were performed to assess the expression of ING5 mRNA in HepG2 cells treated with 5 μM UA for 24 h (n=3, *P < 0.05 vs. the control group). (G) HepG2 cells were individually transfected with the pGL3-basic reporter

Article Snippet: ACC1, ACLY and T antigen overexpression and shRNA plasmids (HonorGene, Changsha, China) and SRF (sc-36563, Santa Cruz, CA, USA) and YY1 (sc-36863, Santa Cruz, CA, USA) siRNA plasmids were transfected into cells using a Lipofectamine 3000 transfection kit (L3000015, Invitrogen, USA).

Techniques: Expressing, Software, Transfection, Western Blot, shRNA, Over Expression, Plasmid Preparation, CCK-8 Assay, Control

Journal: BMB Reports

Article Title: Quercetin induces dual specificity phosphatase 5 via serum response factor

doi: 10.5483/BMBRep.2023-0051

Figure Lengend Snippet: SRF transcription factor modulated the expression of DUSP5 gene. (A) A representative Western blot image demonstrates the protein expression levels of DUSP5 and p-ERK1. The transfection efficiency of SRF siRNA was assessed using an anti-SRF antibody. (B) HCT116 cells were transiently co-transfected with the human DUSP5 promoter region and siRNA. The induction of the DUSP5 promoter was quantified using a luciferase assay. The data presented are relative to the DMSO-treated sample, which is assigned a value of 1.0. (C) SRF knock-down HCT116 cells were treated with DMSO or quercetin. Cell viability was determined using an MTS assay. The DMSO-treated samples were designated as 100%. (D) A chromatin immunoprecipitation (ChIP) assay was performed to quantify the enhancement of the DUSP5 gene using quantitative real-time PCR (qRT-PCR). The fold enrichment of ChIP was calculated using the comparative Ct method and normalized to normal rabbit IgG. All bar graphs represent the mean data ± standard deviation (SD) from three independent experiments. Statistical significance was determined using *P < 0.05, **P < 0.01, and ***P < 0.001. (E) The molecular mechanism underlying the effects of quercetin on SRF and DUSP5 expression can be summarized by a schematic diagram. Quercetin induces the upregulation of DUSP5 through the activation of SRF. This increased expression of DUSP5 potentially affects the dephosphorylation of ERK. However, it should be noted that quercetin itself can mediate the phosphorylation of ERK, which ultimately contributes to the anticancer activity exhibited by quercetin.

Article Snippet: Serum response factor (SRF) small interfering RNA (siRNA) (sc-36563, Santa Cruz) or negative control siRNA (Bioneer, Daejeon, Korea) were transfected in HCT116 cells using PepMuteTM siRNA Transfection Reagent (SignaGen).

Techniques: Expressing, Western Blot, Transfection, Luciferase, Knockdown, MTS Assay, Chromatin Immunoprecipitation, Real-time Polymerase Chain Reaction, Quantitative RT-PCR, Standard Deviation, Activation Assay, De-Phosphorylation Assay, Phospho-proteomics, Activity Assay