Activation of phosphodiesterase 5 and inhibition of guanylate cyclase by cGMP-dependent protein kinase in smooth muscle.
The regulation of cGMP-specific phosphodiesterase (PDE) 5 and soluble guanylate cyclase (GC) by cGMP- and cAMP-dependent protein kinases (PKG and PKA respectively) was examined in gastric smooth muscle. The NO donor, sodium nitroprusside (SNP), stimulated PDE5 phosphorylation and activity, which was blocked by the selective PKG inhibitor, KT5823, resulting in an elevation of cGMP levels. Activation of PKA either directly by Sp-5,6-dichloro-1-beta-d-ribofuranosyl benzimidazole 3',5'-cyclic monophosphothioate, or via isoproterenol- and forskolin-dependent increase in cAMP, also caused an increase in PDE5 phosphorylation and activity, but only in the presence of cGMP; consistent with the dependence of PDE5 phosphorylation and activity on cGMP binding to allosteric sites in the regulatory domain of PDE5. The selective PKA inhibitors, myristoylated protein kinase inhibitor and H-89, blocked the increase in PDE5 phosphorylation and activity induced by PKA. SNP also stimulated soluble GC phosphorylation and activity. KT5823 abolished phosphorylation and augmented soluble GC activity, implying feedback inhibition of soluble GC by PKG-dependent phosphorylation. Phosphorylation by PKG was direct and could be induced in vitro. Activation of PKA had no effect on soluble GC. Thus cGMP levels are regulated by PKG- and PKA-dependent activation of PDE5 and PKG-specific inhibition of soluble GC.
Murthy
K S
KS
Department of Physiology, Medical College of Virginia, Virginia Commonwealth University, Richmond, VA 23298-0711, USA. skarnam@hsc.vcu.edu
D015106:3',5'-Cyclic-GMP Phosphodiesterases; D000470:Alkaloids / Q000494:pharmacology; D000495:Allosteric Site; D000818:Animals; D015153:Blotting, Western; D002227:Carbazoles; D002417:Cattle; D002478:Cells, Cultured; D000242:Cyclic AMP / Q000378:metabolism; D017868:Cyclic AMP-Dependent Protein Kinases / Q000378:metabolism; D006152:Cyclic GMP / Q000378:metabolism; D017869:Cyclic GMP-Dependent Protein Kinases / Q000378:metabolism*; D054706:Cyclic Nucleotide Phosphodiesterases, Type 5; D004305:Dose-Response Relationship, Drug; D004789:Enzyme Activation; D004791:Enzyme Inhibitors / Q000494:pharmacology; D006162:Guanylate Cyclase / Q000037:antagonists & inhibitors*; D007211:Indoles; D007421:Intestine, Small / Q000201:enzymology*; D008387:Marine Toxins; D008954:Models, Biological; D009130:Muscle, Smooth / Q000201:enzymology*; D009599:Nitroprusside / Q000494:pharmacology; D010080:Oxazoles / Q000494:pharmacology; D010727:Phosphoric Diester Hydrolases / Q000378:metabolism*; D010766:Phosphorylation; D011233:Precipitin Tests; D017434:Protein Structure, Tertiary; D011817:Rabbits; D011863:Radioimmunoassay
D000470:Alkaloids; D002227:Carbazoles; D004791:Enzyme Inhibitors; D007211:Indoles; D008387:Marine Toxins; D010080:Oxazoles; C073601:KT 5823; D009599:Nitroprusside; C059041:calyculin A; D000242:Cyclic AMP; D017868:Cyclic AMP-Dependent Protein Kinases; D017869:Cyclic GMP-Dependent Protein Kinases; D010727:Phosphoric Diester Hydrolases; D015106:3',5'-Cyclic-GMP Phosphodiesterases; D054706:Cyclic Nucleotide Phosphodiesterases, Type 5; D006162:Guanylate Cyclase; D006152:Cyclic GMP