Phosphoinositide 3-kinase lipid products regulate ATP-dependent transport by sister of P-glycoprotein and multidrug resistance associated protein 2 in bile canalicular membrane vesicles

Bile acid transport and secretion in hepatocytes require phosphatidylinositol (PI) 3-kinase-dependent recruitment of ATP-dependent transporters to the bile canalicular membrane and are accompanied by increased canalicular PI 3-kinase activity. We report here that the lipid products of PI 3-kinase also regulate ATP-dependent transport of taurocholate and dinitrophenyl-glutathione directly in canalicular membranes. ATP-dependent transport of taurocholate and dinitrophenyl-glutathione in isolated canalicular vesicles from rat liver was reduced 50-70% by PI 3-kinase inhibitors, wortmannin, and LY294002, at concentrations that are specific for Type I PI 3-kinase, Inhibition was reversed by addition of lipid products of PI 3-kinase (PI 3,4-bisphosphate and, to a lesser extent, PI 3-phosphate and PI 3,4,5-trisphosphate) but not by PI 4,5-bisphosphate. A membrane-permeant synthetic 10-mer peptide that binds polyphosphoinositides and leads to activation of PI 3-kinase in macrophages doubled PI 3-kinase activity in canalicular membrane vesicles and enhanced taurocholate and dinitrophenylglutathione transport in canalicular membrane vesicles above maximal ATP-dependent transport. The effect of the peptide was blocked by wortmannin and LY294002. PI 3-kinase activity was also necessary for function of the transporters in vivo. ATP-dependent transport of taurocholate and PI 3-kinase activity were reduced in canalicular membrane vesicles isolated from rat liver that had been perfused with taurocholate and wortmannin. PI 3,4-bisphosphate enhanced ATP-dependent transport of taurocholate in these vesicles above control levels. Our results indicate that PI 3-kinase lipid products are necessary in vivo and in vitro for maximal ATP-dependent transport of bile acid and nonbile acid organic anions across the canalicular membrane. Our results demonstrate regulation of membrane ATP binding cassette transporters by PI 3-kinase lipid products.