Fig. 3: MED15 T603 phosphorylation accelerates cellular senescence by exacerbating the SASP.

a Endogenous MED15 T603D mutant genotyping by PCR and Sanger sequencing in MCF7 cells. b Immunoblot analysis with the indicated antibodies in WT and MED15 T603D mutant MCF7 cells treated without or with 2 ng/mL TGFβ. β-Actin was blotted as a loading control. c qRT-PCR was used to determine the mRNA levels of SASP genes in WT and MED15 T603D mutant MCF7 cells. The values are presented as means ± SD. *P < 0.05, **P < 0.01, and ***P < 0.001. d CCK-8 assays were performed to assess the proliferation rates of WT and T603D mutant MCF7 cells. The values are presented as means ± SD. *P < 0.05 and ***P < 0.001. e β-Galactosidase activity was determined by β-gal staining in cultured WT and MED15 T603D mutant MCF7 cells treated without or with 5 ng/mL TGFβ. f Statistical analysis of the senescent cells in (e). The values are presented as means ± SD. ***P < 0.001. g Heatmap of DEGs identified by RNA-seq in WT and MED15 T603D mutant MCF7 cells. h Scatter plot of DEGs (FPKM, fold change ≥ 1.5, P < 0.05) identified by RNA-seq in WT and MED15 T603D mutant MCF7 cells. i GO analysis of DEGs upregulated by MED15 T603D mutant. j GSEA enrichment plot of the indicated gene sets. k Heatmaps of DEGs associated with TβR genes and SASP genes.