Mice lacking protein kinase Cα show enhanced membrane localization and activity of the epithelial sodium channel (ENaC) in the renal cortical collecting duct

ENaC insertion into the apical membrane of collecting duct principal cells is dependent on recruitment by the phosphatidylinositol lipids PIP 2 and PIP 3 . The presence of PIP 2 at the membrane is determined by membrane targeting of myristolyated C kinase anchoring protein (MARCKS). Phosphorylation and membrane removal of MARCKS by protein kinase C (PKC) negatively regulates ENaC as shown using PKC activators in a cell culture model. To test whether PKCα influences ENaC activity in vivo , we tested the regulation of ENaC in PKCα −/− mice. Cortical collecting duct tubules were dissected and split open and principal cells were subjected to whole cell patch clamp. In the absence of PKCα, open probability (Po) of ENaC was increased three‐fold vs wild type SV129 mice (0.52 ± 0.04 vs 0.17 ± 0.02). The number of channels per patch was also increased. We observed an increase in membrane accumulation of α, β, and γ subunits of ENaC in the cortical collecting ducts of PKCα−/−. To confirm this increase, one kidney from each animal was perfused with biotin and membrane protein pulled down with streptavidin. The unbiotinylated kidney was used to assess total protein. While total ENaC protein did not change in PKCα−/− mice, membrane localization of both α and β ENaC was increased. These results suggest negative regulation of membrane accumulation and Po of ENaC by PKCα in vivo . This work was supported by NIH grants R37‐DK037963 and T32‐DK07656.