Role of SGK1 in nitric oxide inhibition of ENaC in Na+-transporting epithelia.

摘要

Several studies have shown that nitric oxide (NO) inhibits Na(+) transport in renal and alveolar monolayers. However, the mechanisms by which NO alters epithelial Na(+) channel (ENaC) activity is unclear. Therefore, we examined the effect of applying the NO donor drug l-propanamine 3,2-hydroxy-2-nitroso-1-propylhidrazino (PAPA-NONOate) to cultured renal epithelial cells. A6 and M1 cells were maintained on permeable supports in medium containing 1.5 microM dexamethasone and 10% bovine serum. After 1.5 microM PAPA-NONOate was applied, amiloride-sensitive short-circuit current measurements decreased 29% in A6 cells and 44% in M1 cells. This differed significantly from the 3% and 19% decreases in A6 and M1 cells, respectively, treated with control donor compound (P < 0.0005). Subsequent application of PAPA-NONOate to amiloride-treated control (no NONOate) A6 and M1 cells did not further decrease transepithelial current. In single-channel patch-clamp studies, NONOate significantly decreased ENaC open probability (P(o)) from 0.186 +/- 0.043 to 0.045 +/- 0.009 (n = 7; P < 0.05) without changing the unitary current. We also showed that aldosterone significantly decreased NO production in primary cultures of alveolar type II (ATII) epithelial cells. Because inducible nitric oxide synthase (iNOS) coimmunoprecipitated with the serum- and glucocorticoid-inducible kinase (SGK1) and both proteins colocalized in the cytoplasm (as shown in our studies in mouse ATII cells), SGK1 may also be important in regulating NO production in the alveolar epithelium. Our study also identified iNOS as a novel SGK1 phosphorylated protein (at S733 and S903 residues in miNOS) suggesting that one way in which SGK1 could increase Na(+) transport is by altering iNOS production of NO.