Translocation of rhoA Associated with Ca2+ Sensitization of Smooth Muscle

• Published in: The Journal of biological chemistry 1997-04, Vol.272 (16), p.10704-10709
• Main Authors: Gong, M C, Fujihara, H, Somlyo, A V, Somlyo, A P
• Format: Article
• Language: English
• Subjects: Animals; Calcium - pharmacology; cdc42 GTP-Binding Protein, Saccharomyces cerevisiae; Cell Cycle Proteins - isolation & purification; Cell Cycle Proteins - metabolism; Cytosol - metabolism; GTP-Binding Proteins - isolation & purification; GTP-Binding Proteins - metabolism; Guanosine 5'-O-(3-Thiotriphosphate) - pharmacology; Guanosine Triphosphate - metabolism; Ileum; In Vitro Techniques; Isometric Contraction - drug effects; Isometric Contraction - physiology; Kinetics; Muscle, Smooth - drug effects; Muscle, Smooth - physiology; Muscle, Smooth, Vascular - drug effects; Muscle, Smooth, Vascular - physiology; NAD - metabolism; Portal Vein; Rabbits; rhoA GTP-Binding Protein; Type C Phospholipases - pharmacology
• ISSN: 0021-9258; 1083-351X
• DOI: 10.1074/jbc.272.16.10704

We determined the relationship between the localization of rhoA and Ca 2+ sensitization of force in smooth muscle. In α-toxin-permeabilized rabbit portal vein at p Ca 6.5, the particulate hydrophobic fraction of rhoA (10 ± 1.6% of the total) was significantly increased by phenylephrine to 18 ± 5.5% at 5 min, by AlF 4 − to 26 ± 8.4% at 20 min, and dose-dependently up to 62 ± 9.5% by guanosine 5′- O -(3-thiotriphosphate) (GTPγS; 0.3–50 μ m ). Translocation of rhoA was selective (Rac1 and Cdc42 were not translocated) and was quantitatively correlated (up to ∼50%; r = 0.91, p < 0.05) with Ca 2+ sensitization; high GTPγS concentrations (≥10 μ m ) further increased translocation without increasing force. The initial recruitment of rhoA to the membrane paralleled the time course of contraction, but sensitization could be reversed without a decrease in particulate rhoA . High [Ca 2+ ] ( p Ca 4.5) also increased particulate rhoA to 31 ± 5.8%. Membrane-associated rhoA in unstimulated portal vein was a good substrate for in vitro ADP-ribosylation, whereas the large amount translocated by GTPγS was not. We conclude that 1) translocation of rhoA plays a causal role in Ca 2+ sensitization, and 2) membrane-bound rhoA can exist in two or more states.