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Updated: May 21, 2026

Study of the Functions and Activities of Neuronal K-Cl Co-Transporter KCC2 Using Western Blotting
Published on: December 9, 2022
Role of cholesterol in functional association between K(+)-Cl(-) cotransporter-3a and Na+,K(+)-ATPase
Kyosuke Fujita1, Takuto Fujii, Takahiro Shimizu
1Department of Pharmaceutical Physiology, University of Toyama, Toyama 930-0194, Japan. d1061211@ems.u-toyama.ac.jp
Abstract:
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.
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