title: Mitogen-induced activation of Na+/H+ exchange in vascular smooth muscle cells involves janus kinase 2 and Ca2+/calmodulin.
abstract: The sodium/proton exchanger type 1 (NHE-1) plays an important role in the proliferation of vascular smooth muscle cells (VSMC). We have examined the regulation of NHE-1 by two potent mitogens, serotonin (5-HT, 5-hydroxytryptamine) and angiotensin II (Ang II), in cultured VSMC derived from rat aorta. 5-HT and Ang II rapidly activated NHE-1 via their G protein-coupled receptors (5-HT(2A) and AT(1)) as assessed by proton microphysiometry of quiescent cells and by measurements of intracellular pH on a FLIPR (fluorometric imaging plate reader). Activation of NHE-1 was blocked by inhibitors of phospholipase C, CaM, and Jak2 but not by pertussis toxin or inhibitors of protein kinase C. Immunoprecipitation/immunoblot studies showed that 5-HT and Ang II induce phosphorylation of Jak2 and induce the formation of signal transduction complexes that included Jak2, CaM, and NHE-1. The cell-permeable Ca(2+) chelator BAPTA-AM blocked activation of Jak2, complex formation between Jak2 and CaM, and tyrosine phosphorylation of CaM, demonstrating that elevated intracellular Ca(2+) is essential for those events. Thus, mitogen-induced activation of NHE-1 in VSMC is dependent upon elevated intracellular Ca(2+) and is mediated by the Jak2-dependent tyrosine phosphorylation of CaM and subsequent increased binding of CaM to NHE-1, similar to the pathway previously described for the bradykinin B(2) receptor in inner medullary collecting duct cells of the kidney [Mukhin, Y. V., et al. (2001) J. Biol. Chem. 276, 17339-17346]. We propose that this pathway represents a fundamental mechanism for the rapid regulation of NHE-1 by G(q/11) protein-coupled receptors in multiple cell types.
authors.affiliation: Medical and Research Services, Ralph H. Johnson Veterans Affairs Medical Center, and Department of Medicine (Nephrology Division), Medical University of South Carolina, Charleston, South Carolina 29425, USA. garnovsk@musc.edu
authors.forename: Maria N
authors.initials: MN
authors.lastname: Garnovskaya
authors.forename: Yurii V
authors.initials: YV
authors.lastname: Mukhin
authors.forename: Justin H
authors.initials: JH
authors.lastname: Turner
authors.forename: Tamara M
authors.initials: TM
authors.lastname: Vlasova
authors.forename: Michael E
authors.initials: ME
authors.lastname: Ullian
authors.forename: John R
authors.initials: JR
authors.lastname: Raymond
mesh_terms: D000804:Angiotensin II / Q000494:pharmacology; D000818:Animals; D002118:Calcium / Q000032:analysis / Q000737:chemistry / Q000378:metabolism*; D002147:Calmodulin / Q000378:metabolism*; D002478:Cells, Cultured; D004791:Enzyme Inhibitors / Q000494:pharmacology; D005470:Fluorometry / Q000379:methods; D053614:Janus Kinase 2; D008934:Mitogens / Q000494:pharmacology*; D009131:Muscle, Smooth, Vascular / Q000166:cytology* / Q000187:drug effects / Q000378:metabolism*; D010766:Phosphorylation; D011233:Precipitin Tests; D011505:Protein-Tyrosine Kinases / Q000378:metabolism*; D011518:Proto-Oncogene Proteins; D051381:Rats; D017207:Rats, Sprague-Dawley; D012701:Serotonin / Q000494:pharmacology; D015398:Signal Transduction; D017923:Sodium-Hydrogen Exchangers / Q000378:metabolism*
chemical_list: D002147:Calmodulin; D004791:Enzyme Inhibitors; D008934:Mitogens; D011518:Proto-Oncogene Proteins; D017923:Sodium-Hydrogen Exchangers; D000804:Angiotensin II; D012701:Serotonin; D011505:Protein-Tyrosine Kinases; C507927:Jak2 protein, rat; D053614:Janus Kinase 2; D002118:Calcium