Role of guanine nucleotide-binding proteins - Ras-family or trimeric proteins or both - in Ca2+ sensitization of smooth muscle

Ming Cui Gong, Kunihiko Iizuka, Graeme Nixon, J. Peter Browne, Alan Hall, John F. Eccleston, Motoyuki Sugai, Sei Kobayashi, Avril V. Somlyo, Andrew P. Somlyo

Research output: Contribution to journalArticlepeer-review

269 Scopus citations

Abstract

The purpose of this study was to identify guanine nucleotide-binding proteins (G proteins) involved in the agonist- and guanosine 5′-[γ-thio]triphosphate (GTP[γ-S])-induced increase in the Ca2+ sensitivity of 20-kDa myosin light chain (MLC20) phosphorylation and contraction in smooth muscle. A constitutively active, recombinant val14p21rhoA.GTP expressed in the baculovirus/Sf9 system, but not the protein expressed without posttranslational modification in Escherichia coli, induced at constant Ca2+ (pCa 6.4) a slow contraction associated with increased MLC20 phosphorylation from 19.8% to 29.5% (P < 0.05) in smooth muscle permeabilized with β-escin. The effect of val14p21rhoA.GTP was inhibited by ADP-ribosylation of the protein and was absent in smooth muscle extensively permeabilized with Triton X-100. ADP-ribosylation of endogenous p21rho with epidermal cell differentiation inhibitor (EDIN) inhibited Ca2+ sensitization induced by GTP [in rabbit mesenteric artery (RMA) and rabbit ileum smooth muscles], by carbachol (in rabbit ileum), and by endothelin (in RMA), but not by phenylephrine (in RMA), and only slowed the rate without reducing the amplitude of contractions induced in RMA by 1 μM GTP[γ-S] at constant Ca2+ concentrations. AIF4--induced Ca2+ sensitization was inhibited by both guanosine 5′-[β-thio]diphosphate (GDP[β-S]) and by EDIN. EDIN also inhibited, to a lesser extent, contractions induced by Ca2+ alone (pCa 6.4) in both RMA and rabbit ileum. ADP-ribosylation of trimeric G proteins with pertussis toxin did not inhibit Ca2+ sensitization. We conclude that p21rho may play a role in physiological Ca2+ sensitization as a cofactor with other messengers, rather than as a sole direct inhibitor of smooth muscle MLC20 phosphatase.

Original languageEnglish
Pages (from-to)1340-1345
Number of pages6
JournalProceedings of the National Academy of Sciences of the United States of America
Volume93
Issue number3
DOIs
StatePublished - Feb 6 1996

Keywords

  • ADP-ribosylation
  • Epidermal cell differentiation inhibitor
  • Pertussis toxin
  • Rho protein

ASJC Scopus subject areas

  • General

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