Dominant negative c-Src inhibits angiotensin II induced activation of NHE3 in OKP cells.
BACKGROUND
Angiotensin II is a potent stimulator of the proximal tubule apical membrane Na/H antiporter, encoded by NHE3. The nonreceptor tyrosine kinase, c-Src, plays a key role in regulation of NHE3 by acidosis in the proximal tubule, and in signaling effects of angiotensin II in vascular smooth muscle.


METHODS
The present studies examined the role of c-Src in mediating angiotensin II-induced NHE3 activation in cultured OKP cells. c-Src was inhibited with herbimycin A, a tyrosine kinase inhibitor, and expression of a dominant negative c-Src, c-SrcK295M.


RESULTS
Herbimycin A blocked angiotensin II induced increases in Na/H antiporter activity. In two clonal cell lines expressing vector alone, angiotensin II increased Na/H antiporter activity, while in three clones expressing c-SrcK295M, angiotensin II had no effect. Cyclic AMP and protein kinase A have been proposed to be key mediators in regulation of NHE3 by angiotensin II. 10(-4) M 8-bromo cAMP induced a 40 to 50% inhibition of Na/H antiporter activity in cells expressing c-SrcK295M, similar to that seen in wild-type OKP cells. In addition, cells expressing c-SrcK295M responded normally to 10(-7) M dexamethasone with a 50 to 80% increase in Na/H antiporter activity.


CONCLUSIONS
These studies demonstrate that c-Src is required for angiotensin II-induced increases in NHE3 activity. Thus, c-Src plays a key role in antiporter activation by acidosis and angiotensin II.
Tsuganezawa
H
H
Department of Internal Medicine, University of Texas Southwestern Medical Center, Dallas, USA.
Preisig
P A
PA
Alpern
R J
RJ
D000804:Angiotensin II / Q000037:antagonists & inhibitors / Q000502:physiology*; D000818:Animals; D016227:Benzoquinones; D000081247:CSK Tyrosine-Protein Kinase; D002478:Cells, Cultured; D000242:Cyclic AMP / Q000502:physiology; D003907:Dexamethasone / Q000494:pharmacology; D007668:Kidney / Q000378:metabolism*; D047029:Lactams, Macrocyclic; D009893:Opossums; D011505:Protein-Tyrosine Kinases / Q000502:physiology*; D011809:Quinones / Q000494:pharmacology; D017828:Rifabutin / Q000031:analogs & derivatives; D017923:Sodium-Hydrogen Exchangers / Q000096:biosynthesis* / Q000235:genetics; D019061:src-Family Kinases
D016227:Benzoquinones; D047029:Lactams, Macrocyclic; D011809:Quinones; D017923:Sodium-Hydrogen Exchangers; D000804:Angiotensin II; D017828:Rifabutin; C020754:herbimycin; D003907:Dexamethasone; D000242:Cyclic AMP; D011505:Protein-Tyrosine Kinases; D000081247:CSK Tyrosine-Protein Kinase; D019061:src-Family Kinases