Ca²⁺Sensitivity of Smooth Muscle and Nonmuscle Myosin II: Modulated by G Proteins, Kinases, and Myosin Phosphatase

Abstract

Ca2+ sensitivity of smooth muscle and nonmuscle myosin II reflects the ratio of activities of myosin light-chain kinase (MLCK) to myosin light-chain phosphatase (MLCP) and is a major, regulated determinant of numerous cellular processes. We conclude that the majority of phenotypes attributed to the monomeric G protein RhoA and mediated by its effector, Rho-kinase (ROK), reflect Ca2+ sensitization: inhibition of myosin II dephosphorylation in the presence of basal (Ca2+ dependent or independent) or increased MLCK activity. We outline the pathway from receptors through trimeric G proteins (Galphaq, Galpha12, Galpha13) to activation, by guanine nucleotide exchange factors (GEFs), from GDP. RhoA. GDI to GTP. RhoA and hence to ROK through a mechanism involving association of GEF, RhoA, and ROK in multimolecular complexes at the lipid cell membrane. Specific domains of GEFs interact with trimeric G proteins, and some GEFs are activated by Tyr kinases whose inhibition can inhibit Rho signaling. Inhibition of MLCP, directly by ROK or by phosphorylation of the phosphatase inhibitor CPI-17, increases phosphorylation of the myosin II regulatory light chain and thus the activity of smooth muscle and nonmuscle actomyosin ATPase and motility. We summarize relevant effects of p21-activated kinase, LIM-kinase, and focal adhesion kinase. Mechanisms of Ca2+ desensitization are outlined with emphasis on the antagonism between cGMP-activated kinase and the RhoA/ROK pathway. We suggest that the RhoA/ROK pathway is constitutively active in a number of organs under physiological conditions; its aberrations play major roles in several disease states, particularly impacting on Ca2+ sensitization of smooth muscle in hypertension and possibly asthma and on cancer neoangiogenesis and cancer progression. It is a potentially important therapeutic target and a subject for translational research.

Pre-contraction with the thromboxane-mimetic U46619 enhances P2X receptor-mediated contractions in isolated porcine splenic artery

“…Calcium sensitisation has been proposed as a mechanism by which synergy can occur between two receptors to enhance contractile responses. Protein kinase C and Rho kinase are implicated in calcium sensitising pathways through their effects on myosin phosphatase activity [7,14]. However, in this study, inhibition of protein kinase C with calphostin C had no effect on either the direct or the enhanced contractions to α,β-methylene ATP.…”

“…Caldesmon inhibits the myosin ATPase. Therefore, inhibition of caldesmon removes this inhibitory effect and therefore promotes contraction of the smooth muscle [14]. Similarly, Rho kinase causes vasoconstriction through inhibition of the myosin phosphatase, thereby preventing dephosphorylation of myosin light chains and so enhancing vasoconstriction.…”

“…Similarly, Rho kinase causes vasoconstriction through inhibition of the myosin phosphatase, thereby preventing dephosphorylation of myosin light chains and so enhancing vasoconstriction. Both pathways are thought to play a role in the process of calcium sensitisation whereby large contractile responses can occur with small increases in intracellular calcium [14]. Therefore, the enhancement of the P2X-mediated vasoconstriction may be through activation of these signalling pathways such that the increase in intracellular calcium through activation of the P2X receptor causes a larger contraction.…”

Comparative pharmacological analysis of Rho-kinase inhibitors and identification of molecular components of Ca2+ sensitization in the rat lower urinary tract

“…Accumulating evidence reveals the physiological importance of the Ca 2+ -independent RhoA/ Rho-kinase pathway in the regulation of smooth muscle tone by alteration of the sensitivity of contractile proteins for Ca 2+ [15]. RhoA regulates smooth muscle contraction by cycling between a GDP-bound inactive form (coupled to a guanine dissociation inhibitor, RhoGDI) and a GTP-bound active form [16][17][18].…”

Conventional myosins – unconventional functions

“…While vertebrate myosin RLCs can be phosphorylated on Ser19 (or equivalent), Thr18 may also be phosphorylated-and the diphosphorylated product maximises filament stability (Ikebe et al 1988). Myosin light chain kinase (MLCK) operates in a calciumcalmodulin-dependent manner and can activate myosin 2 through phosphorylation at both sites (Somlyo and Somlyo 2003). By contrast, Rho kinase (ROCK) (Amano et al 1996), p21-activated kinase (PAK) (Chew et al 1998), citron kinase (Yamashiro et al 2003), zipper-interacting protein (ZIP) kinase (Ihara and MacDonald 2007) and myotonic dystrophy related-kinase cdc42 binding kinase (MRCK) (Leung et al 1998) catalyse the same reactions but operate through diverse calcium-independent pathways.…”