Phosphatidylinositol 3-kinase-mediated endocytosis of renal Na+,K+-ATPase α subunit in response to dopamine

Dopamine (DA) inhibition of Na+,K+-ATPase in proximal tubule cells is associated with increased endocytosis of its a and beta subunits into early and late endosomes via a clathrin vesicle-dependent pathway. In this report we evaluated intracellular signals that could trigger this mechanism, specifically the role of phosphatidylinositol 3-kinase (PI3-K), the activation of which initiates vesicular trafficking and targeting of proteins to specific cell compartments. DA stimulated PI3-K activity in a time-and dose-dependent manner, and this effect was markedly blunted by wortmannin and LY 294002. Endocytosis of the Na+,K+-ATPase alpha subunit in response to DA was also inhibited in dose-dependent manner by wortmannin and LY 294002. Activation of PI3-K generally occurs by association with tyrosine kinase receptors. However, in this study immunoprecipitation with a phosphotyrosine antibody did not reveal PI3-K activity. DA-stimulated endocytosis of Na+,K+-ATPase alpha subunits required protein kinase C, and the ability of DA to stimulate PI 3-K was blocked by specific protein kinase C inhibitors. Activation of PI3-K is mediated via the D-1 receptor subtype and the sequential activation of phospholipase A(2), arachidonic acid, and protein kinase C. The results indicate a key role for activation of PI 3-K in the endocytic sequence that leads to internalization of Na+,K+-ATPase alpha subunits in response to DA, and suggest a mechanism for the participation of protein kinase C in this process.