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Src‐family tyrosine kinase (SFK) phosphorylates With‐No‐ Lysine Kinase4 (WNK4) and modulates the inhibitory effect of WNK4 on ROMK channels.
Author(s) -
Lin DaoHong,
Yue Peng,
Yarborough OrLando,
Lifton Richard,
Rinehart Jesse,
Wang WenHui
Publication year - 2013
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.27.1_supplement.911.2
Subject(s) - phosphorylation , tyrosine protein kinase csk , proto oncogene tyrosine protein kinase src , chemistry , src family kinase , sgk1 , kinase , microbiology and biotechnology , biology , biochemistry , sh2 domain
The aim of the study is to determine whether SFK phosphorylates WNK4 and roles of tyrosine (Tyr) phosphorylation site in modulating inhibitory effect of WNK4 on ROMK. Western blots demonstrated that c‐Src increased WNK4's Tyr phosphorylation and that c‐Src‐induced Tyr phosphorylation sites included Tyr1092, Tyr1094 and Tyr1143. Mass spectrometry further confirmed that c‐Src increased the phosphorylation of WNK4 at Tyr1143. SFK and PTP1D were coimmunoprecipitaed with WNK4. Mutation of Tyr1092 and Tyr1143 decreased the association of c‐Src and PTP‐1D to WNK4, respectively. Also, the inhibitory effect of WNK4 on ROMK was largely abolished in WNK4Y1092F–transfected cells and was absent in cells transfected with WNK4Y1092/1094F. Expression of c‐Src inhibited SGK1‐induced phosphorylation of WNK4 at Ser1196 but had no effect in the cells transfected with WNK4Y1092/1094F. WNK4Y1143F inhibited ROMK as potently as those of wt WNK4 and the inhibitory effect of WNK4Y1143F was not abolished by SGK1 which reversed the inhibitory effect of wt WNK4 on ROMK. However, SGK1 reversed WNK4Y1143F‐induced inhibition of ROMK in the cells treated with SFK inhibitors, suggesting that elimination of PTP‐1D‐WNK4 association may enhance the effect of SFK on WNK4. We conclude that WNK4 is the substrate of SFK and that the association of c‐Src and PTP‐1D with WNK4 at Tyr1092 and Tyr1143 plays a role in modulating the effect of WNK4 on ROMK.