Fig. 5
a Protein phosphorylation of EGFR P-Tyr1068, Akt P-Ser473, Akt P-Thr308, MEK1/2 P-Ser217/Ser221, Erk1/2 P-Thr202/Tyr204, and S6 P-Ser240/Ser244 analyzed by RPPA. NFI (blank-corrected mean fluorescence intensity) ratio of phospho-protein/total protein was calculated, normalized to MCF7/EVC, and ratio in MCF7/EVC cells was set to 1. Up-/downregulation of protein phosphorylation in MCF7/PGRMC1 cells was calculated. *p ≤ 0.05, **p ≤ 0.01 (Student’s t test, n = 3). b Protein phosphorylation of EGFR P-Tyr1068, Akt P-Ser473, MEK1/2 P-Ser217/Ser221, and Erk1/2 P-Thr202/Tyr204 verified by western blot analysis. Cells were treated with EGF (10 ng/mL) for 10 min/37 °C. Representative blot of 3 independent analyses. Total protein expression of EGFR, Akt, MEK1/2, and Erk1/2 verified by western blot analysis. Representative blot of 3 independent analyses shown. c PGRMC1 mediates phosphorylation of EGFR and its downstream targets and upregulates E2 levels, ERα expression, and ERα-target genes. EGFR phosphorylation activates the MAPK signaling cascade (including MEK1/2-, ERK1/2-, and S6-phosphorylation) and PI3K signaling cascade (including Akt- and S6-phosphorylation). Phosphorylation of S6 induces transcription of genes, involved in the regulation of cell cycle progression, cell proliferation, and glucose homeostasis. ERα translocates into the nucleus upon ligand-dependent or ligand-independent activation and acts as a transcription factor to transcribe genes involved in tumor progression. d Overview of the influence of PGRMC1 in cholesterol and lipid metabolism. e MCF7/EVC and MCF7/PGRMC1 cells were treated with 100 μM, 50 μM, 25 μM, 12.5 μM, 6.25 μM, and 3.175 μM simvastatin and respective DMSO control. MDA-MB-231/EVC and MDA-MB-231/PGRMC1 cells were treated with 20 μM, 10 μM, 5 μM, 2.5 μM, 1.25 μm, and 0.625 μM simvastatin and respective DMSO control. Viability was analyzed by MTT assay at t = 24 h, t = 48 h, t = 72 h and 37 °C. Depicted are results after 48 h of treatment. Viability is normalized on the DMSO control. p values were adjusted using the Bonferroni correction (ndoses = 6; nreplicates = 9)