| Article ID | Journal | Published Year | Pages | File Type |
|---|---|---|---|---|
| 1929380 | Biochemical and Biophysical Research Communications | 2012 | 5 Pages |
K+–Cl− cotransporter-3a (KCC3a) is associated with Na+,K+-ATPase α1-subunit (α1NaK) in lipid rafts of gastric acid-secreting cells and positively regulates Na+,K+-ATPase activity. Here, effects of cholesterol on association of KCC3a with α1NaK in lipid rafts were studied in LLC-PK1 cells stably expressing KCC3a. In the cells, lipid rafts destructed by methyl-β-cyclodextrin (MβCD) could be reconstructed by exogenous addition of cholesterol accompanying a shift of both KCC3a and α1NaK from non-rafts to rafts. The KCC3a-increased Na+,K+-ATPase activity was abolished by MβCD, and recovered by repletion of cholesterol without changing expression levels of KCC3a and α1NaK in the cells. KCC3a was co-immunoprecipitated with α1NaK even after destruction of lipid rafts by MβCD, indicating that molecular association of KCC3a with α1NaK still retains in the non-raft environment. Our results suggest that cholesterol is essential for eliciting up-regulation of Na+,K+-ATPase activity by KCC3a in the KCC3a–α1NaK complex.
► Regulatory mechanism of Na+ pump by K+–Cl− cotransporter-3a (KCC3a) was examined. ► Functional association of Na+ pump and KCC3a was eliminated by destruction of raft. ► Molecular association of Na+ pump and KCC3a retained after destruction of raft. ► Cholesterol is essential for eliciting up-regulation of Na+ pump by KCC3a.
