Angiotensin II stimulates p90rsk in vascular smooth muscle cells. A potential Na(+)-H+ exchanger kinase.
Angiotensin II is a multifunctional agonist for vascular smooth muscle cells (VSMCs), stimulating increases in signal events, cell growth, and ion flux. We previously defined protein kinase C (PKC)-dependent and -independent mechanisms by which angiotensin II stimulated activity of the Na(+)-H+ exchanger isoform-1 (NHE-1) and identified a 90-kD kinase that exhibited increased activity in VSMCs isolated from genetically hypertensive rats. To determine whether this 90-kD kinase was p90rsk (RSK), VSMCs were stimulated with 100 nmol/L angiotensin II, and NHE-1 kinase activity was measured by phosphorylation of recombinant NHE-1 (a glutathione S-transferase fusion protein containing amino acids 516 to 815 of the cytoplasmic carboxyl tail) in vitro. NHE-1 kinase (90 kD) activity was markedly decreased by immunodepletion of RSK. Characterization of RSK activation by angiotensin II revealed many similarities to the 90-kD NHE-1 kinase, including time course and NHE-1 domain phosphorylation, as well as regulation by extracellular signal-regulated kinases (ERK1/2), intracellular Ca2+, and PKC. Specifically, angiotensin II stimulated a rapid and transient (peak, 5 minutes) increase in RSK activity. Analysis of several NHE-1 fusion proteins revealed that only proteins containing amino acids 670 to 714 were phosphorylated by RSK. Inhibiting ERK1/2 (30 mumol/L PD098059 for 30 minutes) or chelating intracellular Ca2+ prevented RSK activation. In contrast, downregulating PKC (1 mumol/L phorbol dibutyrate for 24 hours) had little effect. These findings establish RSK as a putative NHE-1 kinase and potential mediator of increased Na(+)-H+ exchange in hypertension.
Takahashi
E
E
Department of Medicine, University of Washington, Seattle 98195, USA.
Abe
J
J
Berk
B C
BC
D000595:Amino Acid Sequence; D000804:Angiotensin II / Q000378:metabolism / Q000494:pharmacology*; D000818:Animals; D002118:Calcium / Q000378:metabolism; D008297:Male; D008969:Molecular Sequence Data; D009131:Muscle, Smooth, Vascular / Q000378:metabolism*; D010766:Phosphorylation; D017346:Protein Serine-Threonine Kinases / Q000378:metabolism*; D051381:Rats; D017207:Rats, Sprague-Dawley; D019893:Ribosomal Protein S6 Kinases; D015398:Signal Transduction / Q000187:drug effects*; D017923:Sodium-Hydrogen Exchangers / Q000378:metabolism*
D017923:Sodium-Hydrogen Exchangers; C069139:growth factor-activatable Na-H exchanger NHE-1; D000804:Angiotensin II; D017346:Protein Serine-Threonine Kinases; D019893:Ribosomal Protein S6 Kinases; D002118:Calcium