Involvement of Src and epidermal growth factor receptor in the signal-transducing function of Na+/K+-ATPase

Nontoxic concentrations of ouabain, causing partial inhibition of the cardi ac myocyte Na+/K+-ATPase, induce hypertrophy and several growth-related gen es through signal pathways that include the activation of Ras and p42/44 mi togen-activated protein kinase (MAPK). The aim of this work was to examine the ouabain-induced events upstream of the Ras/MAPK cascade. Treatment of m yocytes with genistein antagonized ouabain-induced activation of the MAPK, suggesting that protein tyrosine phosphorylation has a role. Tyrosine phosp horylation of several myocyte proteins was increased rapidly upon cell expo sure to ouabain, Lowering of extracellular K+ had a similar ouabain-like ef fect. Ouabain also increased protein tyrosine phosphorylation in A7r5, HeLa , and L929 cells. In cardiac myocytes and A7r5 cells, herbimycin A antagoni zed the ouabain-induced increase in protein tyrosine phosphorylation and MA PK activation. In both cell types, ouabain stimulated Src kinase activity, Src translocation to the Triton-insoluble fraction, Src association with th e epidermal growth factor receptor, and the tyrosine phosphorylation of thi s receptor on site(s) other than its major autophosphorylation site, Tyr(11 73). The findings suggest that (a) the ouabain-induced activation of Src an d the Src-induced phosphorylation of the growth factor receptor provide the scaffolding for the recruitment of adaptor proteins and Ras and the activa tion of Ras/MAPK cascade; and (b) the activation of such pathways may be a common feature of the signal-transducing function of Na+/K+-ATPase in most cells.