Actions downstream of cyclic GMP/protein kinase G can reverse protein kinase C-mediated phosphorylation of CPI-17 and Ca²⁺ sensitization in smooth muscle.
Ca(2+) sensitivity of smooth muscle contraction is modulated by several systems converging on myosin light chain phosphatase (MLCP). Rho-Rho kinase is considered to inhibit MLCP via phosphorylation, whereas protein kinase C (PKC) induced sensitization has been shown to be dependent on phosphorylation of the inhibitory protein CPI-17. We have explored the interaction of cGMP-dependent protein kinase (PKG) with Ca(2+) sensitization pathways using permeabilized mouse smooth muscle. Three conditions giving approximately 50% of maximal active force were compared in small intestinal preparations: 1). Ca(2+)-activated unsensitized muscle (pCa 5.9 with Rho kinase inhibitor Y27632); 2). Rho-Rho kinase-sensitized muscle (pCa 6.1 with guanosine 5'-3-O-(thio)triphosphate); and 3). PKC-sensitized muscle (pCa 6.0 with Y27632 and PKC activator phorbol 12,13-dibutyrate). 8-Br-cGMP relaxed the sensitized muscles but had marginal effects on unsensitized preparations, showing that PKG reverses both PKC and Rho-mediated Ca(2+) sensitization. CPI-17 was present in permeabilized intestinal tissue. In PKC-sensitized preparations, CPI-17 phosphorylation decreased in response to 8-Br-cGMP. The rate of PKC-mediated phosphorylation in the presence of the MLCP inhibitor microcystin-LR was not influenced by 8-Br-cGMP. PKC-induced Ca(2+) sensitization also was reversed in vascular smooth muscle tissues (portal vein and femoral artery). We conclude that actions downstream of cGMP/PKG can reverse PKC-mediated phosphorylation of CPI-17 and Ca(2+) sensitization in smooth muscle.
Bonnevier
Johan
J
Department of Physiological Sciences, Lund University, SE-221 84 Lund, Sweden.
Arner
Anders
A
D015124:8-Bromo Cyclic Adenosine Monophosphate / Q000494:pharmacology; D000577:Amides / Q000494:pharmacology; D000818:Animals; D001427:Bacterial Toxins / Q000494:pharmacology; D001808:Blood Vessels / Q000033:anatomy & histology / Q000378:metabolism; D002118:Calcium / Q000378:metabolism*; D006152:Cyclic GMP / Q000378:metabolism*; D017869:Cyclic GMP-Dependent Protein Kinases / Q000378:metabolism*; D004791:Enzyme Inhibitors / Q000494:pharmacology; D004928:Escin / Q000494:pharmacology; D005260:Female; D016244:Guanosine 5'-O-(3-Thiotriphosphate) / Q000494:pharmacology; D066298:In Vitro Techniques; D007421:Intestine, Small / Q000033:anatomy & histology / Q000378:metabolism; D047908:Intracellular Signaling Peptides and Proteins; D051379:Mice; D008815:Mice, Inbred Strains; D009119:Muscle Contraction / Q000502:physiology*; D009130:Muscle, Smooth / Q000187:drug effects / Q000378:metabolism*; D010539:Permeability; D015240:Phorbol 12,13-Dibutyrate / Q000494:pharmacology; D010766:Phosphorylation; D011493:Protein Kinase C / Q000378:metabolism*; D017346:Protein Serine-Threonine Kinases / Q000378:metabolism; D011725:Pyridines / Q000494:pharmacology; D054460:rho-Associated Kinases
D000577:Amides; D001427:Bacterial Toxins; D004791:Enzyme Inhibitors; D047908:Intracellular Signaling Peptides and Proteins; D011725:Pyridines; C108830:Y 27632; D015124:8-Bromo Cyclic Adenosine Monophosphate; D015240:Phorbol 12,13-Dibutyrate; D016244:Guanosine 5'-O-(3-Thiotriphosphate); D004928:Escin; D017346:Protein Serine-Threonine Kinases; D054460:rho-Associated Kinases; D017869:Cyclic GMP-Dependent Protein Kinases; D011493:Protein Kinase C; D006152:Cyclic GMP; D002118:Calcium