Y-box binding factor-1 is phosphorylated by the MAP kinase pathway. (a) Stimulation of SUM149 cells (SS) with serum, epidermal growth factor (EGF) and phorbal 12-myristate 13-acetate (PMA) (15 min) results in the phosphorylation of Y-box binding factor-1 (YB-1) at the serine 102 residue (S102). There is no change in total YB-1 levels. Phosphorylation of ERK indicates activation of the MAP kinase pathway. Total ERK and vinculin indicate equal loading. (b) Inhibition of MAP kinase signaling with PD098059 results in the loss of growth-factor induced P-YB-1S102 (n = 3). (c) Treating SUM149 cells with Iressa (2 μM) results in a decrease in P-YB-1S102. (d) SUM149, MDA-MB-231, HCC1937 and MDA-MB-468 breast cancer cell lines were compared for expression level of P-RSKS380, P-AKTS473, P-ERKThr202/Tyr204 and P-YB-1S102. The MDA-MB-231 cells express high levels of P-YB-1 in the absence of P-AKTS473; however, they do express P-RSKS380. (e) Immortalized human mammary epithelial cells (HTRY) express P-RSKS380, P-ERKThr202/Tyr204 and P-YB-1S102, but not P-AKT. DMSO, dimethylsulfoxide.