HER2 overexpression regulates MED1 phosphorylation through MAP kinase pathway. A and B, whole-cell extracts from MCF-7, BT474, and MCF-7/HER2 cells were prepared and subjected to Western blot analyses with anti-MED1, p-MED1, HER2, or GAPDH antibodies. C, total RNA was extracted from MCF-7 and MCF-7/ HER2 cells, followed by real-time RT-PCR analyses of MED1 expression and normalization to GAPDH levels. ns, not significant. D and E, Western blot analyses of p-MED1 levels in BT474 (D) and MCF-7/HER2 (E) cells after control vehicle or 50 μmol/L AG825 treatment. F, MCF-7/HER2 cells were treated with 25 μmol/L PD98059 or vehicle for 24 hours, and subjected to Western blot analyses with anti–p-MED1 and anti-GAPDH (control) antibodies.