title: A central role for Pyk2-Src interaction in coupling diverse stimuli to increased epithelial NBC activity.
abstract: Regulation of renal Na-HCO cotransporter (NBC1) activity by cholinergic agonists, ANG II, and acute acidosis (CO(2)) requires both Src family kinase (SFK) and classic MAPK pathway activation. The nonreceptor tyrosine kinase proline-rich tyrosine kinase 2 (Pyk2) couples discrete G protein-coupled receptor and growth factor receptor signaling to SFK activation. We examined the role of Pyk2-SFK interaction in coupling these stimuli to increased NBC1 activity in opossum kidney cells. Carbachol increased tyrosine autophosphorylation of endogenous Pyk2 and ectopically expressed wild-type Pyk2 and were abrogated by kinase-dead mutant (Pyk2-KD) overexpression. Pyk2 phosphorylation was calcium/calmodulin dependent, and Pyk2 associated with Src by means of SH2 domain interaction. Pyk2 phosphorylation and Pyk2-Src interaction by carbachol were mimicked by both ANG II and CO(2). To correlate Pyk2 autophosphorylation and Pyk2-Src interaction with NBC activity, cotransporter activity was measured in untransfected cells and in cells overexpressing Pyk2-KD in the presence or absence of carbachol, ANG II, or CO(2). In Pyk2-KD-overexpressing cells, the effect of carbachol, ANG II, and CO(2) was abolished. We conclude that Pyk2 plays a central role in coupling carbachol, ANG II, and CO(2) to increased NBC activity. This coupling is mediated by Pyk2 autophosphorylation and Pyk2-Src interaction.
authors.affiliation: Section of Nephrology, Department of Medicine, University of Illinois at Chicago, 60612, USA.
authors.forename: Doris Joy D
authors.initials: DJ
authors.lastname: Espiritu
authors.forename: Angelito A
authors.initials: AA
authors.lastname: Bernardo
authors.forename: R Brooks
authors.initials: RB
authors.lastname: Robey
authors.forename: Jose A L
authors.initials: JA
authors.lastname: Arruda
mesh_terms: D000138:Acidosis / Q000503:physiopathology; D000804:Angiotensin II / Q000494:pharmacology; D000818:Animals; D015153:Blotting, Western; D002245:Carbon Dioxide / Q000494:pharmacology; D002460:Cell Line; D004847:Epithelial Cells / Q000378:metabolism*; D005470:Fluorometry; D051418:Focal Adhesion Kinase 2; D019204:GTP-Binding Proteins / Q000502:physiology; D006863:Hydrogen-Ion Concentration; D007202:Indicators and Reagents; D048052:Mitogen-Activated Protein Kinase 3; D020928:Mitogen-Activated Protein Kinases / Q000378:metabolism; D009893:Opossums / Q000502:physiology*; D010275:Parasympathetic Nervous System / Q000502:physiology; D010277:Parasympathomimetics / Q000494:pharmacology; D010766:Phosphorylation; D011233:Precipitin Tests; D011505:Protein-Tyrosine Kinases / Q000502:physiology*; D015398:Signal Transduction / Q000502:physiology; D027982:Sodium-Bicarbonate Symporters / Q000378:metabolism*; D013268:Stimulation, Chemical; D014443:Tyrosine / Q000378:metabolism
chemical_list: D007202:Indicators and Reagents; D010277:Parasympathomimetics; D027982:Sodium-Bicarbonate Symporters; D000804:Angiotensin II; D002245:Carbon Dioxide; D014443:Tyrosine; D011505:Protein-Tyrosine Kinases; D051418:Focal Adhesion Kinase 2; D048052:Mitogen-Activated Protein Kinase 3; D020928:Mitogen-Activated Protein Kinases; D019204:GTP-Binding Proteins