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Heterogeneity of Signal Transduction by Na/K‐ATPase α‐Isoforms: Role of Src Interaction
Author(s) -
Yu Hui,
Cui Xiaoyu,
Xie ZiJian
Publication year - 2016
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.30.1_supplement.1224.1
Subject(s) - proto oncogene tyrosine protein kinase src , gene isoform , signal transduction , mutant , biology , microbiology and biotechnology , conserved sequence , phosphotyrosine binding domain , peptide sequence , genetics , gene , sh2 domain
Among the four α isoforms expressed in mammalian cells, we have shown that the ubiquitous α1 Na/K‐ATPase possesses both ion‐pumping and Src‐dependent signaling functions. In contrast, we have reported that α2 Na/K‐ATPase pumps but does not conduct Src‐dependent signal transduction, which represents a major functional difference between α isoforms. We have identified two putative pairs of domain interactions between α1 Na/K‐ATPase and Src, but others have challenged the concept of a direct interaction between Na/K‐ATPase and Src. In support of our model, a sequence comparison revealed major differences between α1 and α2 in these two putative Src‐interacting domains, in contrast to the overall high (~80%) level of homology between the α1 and α2 polypeptides. In this study, we tested whether targeted point mutations in the two putative Src‐interacting domains could confer “α1‐like” Src‐regulating properties to α2. To this end, we generated stable mammalian cell lines that express almost exclusively a α2 polypeptide mutated to mimic α1 sequence in the putative Src‐interacting domains. Functional analyses of α2 and α2 mutant cell lines reveal the following. First, the mutant α2 was fully functional as an ion pump. Second, unlike α2, the mutated α2 gained α1‐like signaling function, was capable of Src regulation, and restored caveolin‐1 expression. Because these mutated sequences of α2 are highly conserved across different species, the lack of Src‐regulation by α2 appears to be a conserved property of this isoform, most likely to provide α2‐specific and physiologically important functions that Src interaction could alter. Third, as expected, ouabain activated Src/ERK in the α2 mutant, but not in α2‐expressing cells. Finally, the mutant α2 cells exhibited a growth phenotype comparable to that of α1, which was much faster than that of α2 cells. Taken together, these new findings not only reveal the structural requirements for the Na/K‐ATPase to function as a Src‐dependent receptor, but also provide strong evidence of isoform‐specific Src interaction involving the identified key amino acids.