Role of cholesterol in functional association between K^+-Cl^- cotransporter-3a and Na^+, K^+-ATPase
Role of cholesterol in functional association between K^+-Cl^- cotransporter-3a and Na^+, K^+-ATPase
复制标题
胆固醇在 K^ -Cl^- 协同转运蛋白-3a 和 Na^ , K^ -ATPase 功能关联中的作用
DOI：
10.1016/j.bbrc.2012.06.089
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发表时间：
2012
影响因子：
3.1
通讯作者：
Kyosuke Fujita
中科院分区：
文献类型：
--
作者：
清水貴浩;家原貴大;佐藤かお理;藤井拓人;酒井秀紀;岡田泰伸;Kyosuke Fujita
K+ -Cl - cotransporter-3a （KCC3a）与胃酸分泌细胞脂筏中Na+，K+- atp酶α1亚基（α1NaK）相关，并积极调节Na+，K+- atp酶活性。本研究在稳定表达KCC3a的LLC-PK1细胞中研究胆固醇对脂筏中KCC3a与α1NaK关联的影响。在细胞中，被甲基β-环糊精（m -β cd）破坏的脂质筏可以通过外源性胆固醇的加入而重建，同时KCC3a和α1NaK从非筏变为筏。在不改变细胞中KCC3a和α1NaK表达水平的情况下，KCC3a增加的Na+、K+- atp酶活性被MβCD消除，并通过补充胆固醇恢复。即使在m - β cd破坏脂筏后，KCC3a仍与α1NaK共免疫沉淀，这表明在非脂筏环境下，KCC3a与α1NaK的分子关联仍然存在。我们的研究结果表明，在KCC3a -α1NaK复合物中，胆固醇对KCC3a上调Na+，K+- atp酶活性至关重要。
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.