Phosphorylation of P-glycoprotein by PKA and PKC modulates swelling- activated Cl^- currents

Several proteins belonging to the ATP-binding cassette superfamily can affect ion channel function. These include the cystic fibrosis transmembrane conductance regulator, the sulfonylurea receptor, and the multidrug resistance protein P-glycoprotein (MDR1). We measured whole cell swelling- activated Cl^- currents (I(Cl,swell)) in parental cells and cells expressing wild-type MDR1 or a phosphorylation-defective mutant (Ser-661, Ser-667, and Ser-671 replaced by Ala). Stimulation of protein kinase C (PKC) with a phorbol ester reduced the rate of increase in I(Cl,swell) only in cells that express MDR1. PKC stimulation had no effect on steady-state I(Cl,swell). Stimulation of protein kinase A (PKA) with 8-bromoadenosine 3',5'-cyclic monophosphate reduced steady-state I(Cl,swell) only in MDR1-expressing cells. PKA stimulation had no effect on the rate of I(Cl,swell) activation. The effects of stimulation of PKA and PKC on I(Cl,swell) were additive (i.e., decrease in the rate of activation and reduction in steady-state I(Cl,swell). The effects of PKA and PKC stimulation were absent in cells expressing the phosphorylation-defective mutant. In summary, it is likely that phosphorylation of MDR1 by PKA and by PKC alters swelling-activated Cl^- channels by independent mechanisms and that Ser-661, Ser-667, and Ser-671 are involved in the responses of I(Cl,swell) to stimulation of PKA and PKC. These results support the notion that MDR1 phosphorylation affects I(Cl,swell).