EFFECT OF A PEPTIDE INHIBITOR OF PROTEIN-KINASE-C ON G-PROTEIN-MEDIATED INCREASE IN MYOFILAMENT CA2+-SENSITIVITY IN RABBIT ARTERIAL SKINNED MUSCLE

1 To investigate the role of protein kinase C in the increase mediated by guanosine 5'-triphosphate (GTP)-binding proteins (G-proteins) in the sensitivity of the contractile proteins to Ca2+ in vascular smooth muscle, the effect of a novel peptide inhibitor of protein kinase C (PKC19-36) on Ca2+-induced contraction and myosin light chain (MLC) phosphorylation was studied in the presence and absence of guanosine 5'-O-(3-thiotriphosphate) (GTP(gamma)S) in beta-escin-skinned smooth muscle strips of rabbit mesenteric artery. For comparison, the effects were also observed of PKC19-36 On the action of phorbol 12,13-dibutylate (PDBu, an activator of PKC) on the two Ca2+-induced responses. 2 In beta-escin-skinned strips treated with ionomycin, Ca2+ (0.1-3 mu M) concentration-dependently produced contraction in parallel with an increase in MLC-phosphorylation, GTPVS (10 mu M) and PDBu (0.1 mu M) each shifted both the Ca2+-force and Ca2+-MLC-phosphorylation relationships to the left without a significant change in either maximum response. The relationship between force and MLC-phosphorylation was not modified by either GTP(gamma)S or PDBu, indicating that the sensitivity of MLC-phosphorylation to Ca2+ is enhanced by both GTP(gamma)S and PDBu. 3 PKC19-36 itself modified neither the contraction nor MLC-phosphorylation induced by Ca2+ but it did block the PDBu-induced enhancement of these two Ca2+-induced responses. By contrast, PKC19-36 did not modify the GTP(gamma)S-induced enhancement of the two Ca2+-induced responses. Guanosine 5'-O-(2-thiodiphosphate) (GDP beta S) attenuated the GTP(gamma)S-induced enhancement of the Ca2+-induced contraction. 4 These results suggest that GTP(gamma)S increases Calf-induced MLC-phosphorylation through the activation of a PKC-independent mechanism and thus causes an increase in the sensitivity of the contractile proteins to Ca2+ in beta-escin-skinned smooth muscle of rabbit mesenteric artery.