Downregulation of SRC-2 in MCF-7 cells induces distinct changes in global gene expression profiles.

<p>(A). Quantification of SRC-2 mRNA expression in shRNA lentivirus-infected MCF-7 cells. mRNA levels of SRC-2 in a MCF-7 cell line infected with shRNA targeting SRC-2 (SRC-2 shRNA) were compared to the expression in a control shRNA MCF-7 cell line (Ctr shRNA), and in a MCF-7 cell line transduced with shRNA lentivirus targeting SRC-3 (SRC-3 shRNA). The mRNA expression of SRC-2 is relative to TBP mRNA. The results are representative of at least three independent experiments. (B). Western blotting analyses of SRC-2-depleted MCF-7 cells. MCF-7 cells infected with shRNA lentivirus targeting SRC-2 (SRC-2 shRNA) or a negative control shRNA empty vector (Ctr shRNA), were grown in phenol red-free DMEM supplemented with charcoaled stripped FBS (5%) and 17β-estradiol (10 nM) for two days. The Ctr shRNA cells were then treated with either Vehicle or 8-CPT-cAMP (150 µM), IBMX (50 µM) and forskolin (10 µM) (cAMP) for 24 hours. Immunoblotting was performed with anti-TIF2 antibody and anti-GAPDH antibody. The results shown are representative of at least three independent experiments. (C). Microarray analyses of five RNA samples isolated from five individual cell samples of shRNA control MCF-7 cells (Ctr shRNA), SRC-2 KD MCF-7 cells (SRC-2 shRNA) and control shRNA cells treated with cAMP elevating agents, as described in A. A Venn diagram shows the number of individual and overlapping sets of genes differentially expressed after SRC-2 KD (SRC-2 shRNA) and after treatment with cAMP elevating agents. To examine which genes were similarly differentially expressed between the two treated groups when compared to control, a SAM analysis with overlapping genes was performed. The fold change cut-off value ≥1.5, and q-value = 0, was used to determine differentially expressed genes.