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Regulation of claudins in renal TJ by Na + ,K + ‐ATPase alpha 1 subunit
Author(s) -
Larre Maria Isabel,
Zhang Jue,
Xie Zijian
Publication year - 2017
Publication title -
the faseb journal
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 1.709
H-Index - 277
eISSN - 1530-6860
pISSN - 0892-6638
DOI - 10.1096/fasebj.31.1_supplement.1007.53
Subject(s) - claudin , paracellular transport , ouabain , tight junction , chemistry , microbiology and biotechnology , nephron , western blot , epithelial sodium channel , apical membrane , biology , permeability (electromagnetism) , biophysics , sodium , biochemistry , membrane , organic chemistry , renal function , gene
Tight junctions (TJs) are located at the uppermost portion of the lateral membrane of epithelial cells that seal the intracellular interspace between adjacent cells, transforming the epithelial cells into an effective permeable barrier. Tight junctions regulate the movements of substances through the paracellular pathway, and claudins are major determinants of TJ permeability. In the kidneys, the molecular composition of TJ determines the permeability and ion selectivity of different nephron segments along renal tubules. For example, while proximal tubules express claudin‐2, distal nephrons express the claudin‐4 isoform. Little is known about what controls the composition of TJ along the nephron. Previous reports have shown that ouabain, a cardiac glycoside that acts by inhibiting the Na + ,K + ‐ATPase pump activity, increases TJ sealing degree (10–100 nM), suggesting the participation of Na + ,K + ‐ATPase in TJ regulation. In this study, with newly developed rescue α1 mutant protocol, we were able to generate unique stable epithelial cell lines that express Na + ,K + ‐ATPase alpha‐1 with specific mutations in the N domain. The expression of Na + ,K + ‐ATPase alpha‐1 mutants (A425P and A420P) does not affect the activity of sodium pump but increases the level expression of Src phosphorylation at Y418 and also decrees the expression of caveolin‐1. This study is the first approach to investigated the role of Na + ,K + ‐ATPase alpha 1 on TJ protein expression and function on kidney epithelial cells. Results Using TER measurement, western blot and confocal microscopy we found that A420P and A425P mutation decreases the ion permeability and changes the molecular composition of TJ proteins when compared to wild type Na + ,K + ‐ATPase alpha‐1. The expression levels and cellular localization of Claudin‐2 and claudin‐4 were altered. Src inhibitor PP2 and pNaKtide were able to decrease the TER values in A425P cell line comparable to wild type. Thus, this work shows that the N domain of Na + /K + ‐ATPase alpha‐1 has an important role in tight junction function and this regulation implicates Na + ,K + ‐ATPase‐Src‐Caveolin‐Claudin pathway. Taken together, these observations have a significant relevance to the altered function of kidneys in physiological and pathological conditions, in which the transport of ion and nutrients are significantly altered.