| Autor: |
Nickerson AJ; Department of Medicine., Mutchler SM; Department of Medicine., Sheng S; Department of Medicine., Cox NA; Department of Medicine., Ray EC; Department of Medicine., Kashlan OB; Department of Medicine.; Department of Computational and Systems Biology., Carattino MD; Department of Medicine.; Department of Cell Biology, and., Marciszyn AL; Department of Medicine., Winfrey A; Department of Medicine., Gingras S; Department of Immunology, University of Pittsburgh, Pittsburgh, Pennsylvania, USA., Kirabo A; Department of Medicine, Vanderbilt University Medical Center, Nashville, Tennessee, USA., Hughey RP; Department of Medicine.; Department of Cell Biology, and., Kleyman TR; Department of Medicine.; Department of Cell Biology, and.; Department of Pharmacology and Chemical Biology, University of Pittsburgh, Pittsburgh, Pennsylvania, USA. |
| Abstrakt: |
Epithelial Na+ channels (ENaCs) control extracellular fluid volume by facilitating Na+ absorption across transporting epithelia. In vitro studies showed that Cys-palmitoylation of the γENaC subunit is a major regulator of channel activity. We tested whether γ subunit palmitoylation sites are necessary for channel function in vivo by generating mice lacking the palmitoylated cysteines (γC33A,C41A) using CRISPR/Cas9 technology. ENaCs in dissected kidney tubules from γC33A,C41A mice had reduced open probability compared with wild-type (WT) littermates maintained on either standard or Na+-deficient diets. Male mutant mice also had higher aldosterone levels than WT littermates following Na+ restriction. However, γC33A,C41A mice did not have reduced amiloride-sensitive Na+ currents in the distal colon or benzamil-induced natriuresis compared to WT mice. We identified a second, larger conductance cation channel in the distal nephron with biophysical properties distinct from ENaC. The activity of this channel was higher in Na+-restricted γC33A,C41A versus WT mice and was blocked by benzamil, providing a possible compensatory mechanism for reduced prototypic ENaC function. We conclude that γ subunit palmitoylation sites are required for prototypic ENaC activity in vivo but are not necessary for amiloride/benzamil-sensitive Na+ transport in the distal nephron or colon. |
