NO decreases phosphorylation of focal adhesion proteins via reduction of Ca in rat aortic smooth muscle cells.
Our laboratory has previously reported that the antimitogenic effect of nitric oxide (NO) in primary cultures of rat aortic smooth muscle cells may be attributed to activation of protein tyrosine phosphatase and dephosphorylation of protein phosphotyrosine [G.S. Dhaunsi, C. Matthews, K. Kaur, and A. Hassid, Am, J. Physiol. 272 (Heart Circ. Physiol. 41): H1342-H1349, 1997]. The goal of the current study was to investigate the role of cytoplasmic Ca in this process and to identify protein substrates that are dephosphorylated by treatment with NO. Treatment of primary rat aortic smooth muscle cell cultures with the NO donor S-nitroso-N-acetylpenicillamine (SNAP) decreased cytoplasmic Ca levels and elicited phosphotyrosine dephosphorylation. Both effects were mimicked by the extracellular and intracellular Ca chelators ethylene glycol-bis(beta-aminoethyl ether)-N,N,N',N'-tetraacetic acid (EGTA) and 1,2-bis(2-aminophenoxy)ethane-N,N,N',N'-tetraacetic acid (BAPTA), respectively, and by the Ca channel blocker nifedipine. Conversely, elevation of cytoplasmic Ca via the use of the Ca ionophore A-23187 or high extracellular K+ prevented or attenuated SNAP-induced dephosphorylation. Both BAPTA and nifedipine also decreased DNA synthesis, providing further evidence to link dephosphorylation to antimitogenesis. Two of the proteins dephosphorylated by treatment of cells with NO or EGTA were identified as the focal adhesion proteins, cortactin and paxillin. These results indicate that NO-induced dephosphorylation of protein phosphotyrosine is mediated by reduction of cytoplasmic Ca and suggest that dephosphorylation of focal adhesion proteins may be of relevance to the antimitogenic effect of NO.
Kaur
K
K
Department of Physiology and Biophysics, University of Tennessee, Memphis 38163, USA.
Yao
J
J
Pan
X
X
Matthews
C
C
Hassid
A
A
D000818:Animals; D000001:Calcimycin / Q000494:pharmacology; D002118:Calcium / Q000378:metabolism*; D002478:Cells, Cultured; D002614:Chelating Agents / Q000494:pharmacology; D051356:Cortactin; D003598:Cytoskeletal Proteins / Q000378:metabolism*; D004533:Egtazic Acid / Q000494:pharmacology; D004789:Enzyme Activation; D005260:Female; D007476:Ionophores / Q000494:pharmacology; D008840:Microfilament Proteins / Q000378:metabolism*; D009131:Muscle, Smooth, Vascular / Q000378:metabolism*; D009569:Nitric Oxide / Q000378:metabolism* / Q000494:pharmacology*; D051419:Paxillin; D010396:Penicillamine / Q000031:analogs & derivatives / Q000494:pharmacology; D010750:Phosphoproteins / Q000378:metabolism*; D010766:Phosphorylation; D017027:Protein Tyrosine Phosphatases / Q000378:metabolism; D051381:Rats; D017207:Rats, Sprague-Dawley
D002614:Chelating Agents; D051356:Cortactin; C495870:Cttn protein, rat; D003598:Cytoskeletal Proteins; D007476:Ionophores; D008840:Microfilament Proteins; D051419:Paxillin; D010750:Phosphoproteins; C496063:Pxn protein, rat; C110881:S-nitro-N-acetylpenicillamine; D009569:Nitric Oxide; D000001:Calcimycin; D004533:Egtazic Acid; D017027:Protein Tyrosine Phosphatases; D010396:Penicillamine; D002118:Calcium