Gene/Protein Disease Symptom Drug Enzyme Compound
Pivot Concepts: Gene/Protein Disease Symptom Drug Enzyme Compound   Target Concepts: Gene/Protein Disease Symptom Drug Enzyme Compound

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Query: EC:3.6.4.1 (myosin ATPase)
1,140 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

Increased maximum velocity of shortening (Vmax), increased shortening ability (delta Lmax) and decreased relaxation rate have been reported for arterial smooth muscle from 16- to 18-week-old spontaneously, hypertensive rats (SHR) compared with age-matched normotensive Wistar-Kyoto rats (WKY). Vmax is dependent on actomyosin ATPase activity, and this activity is in turn dependent on the level of phosphorylation of the 20-kDa myosin light chain (MLC20) normally a function of calcium concentration. In this article, methods are described and data are presented from studies addressing possible intracellular regulatory mechanisms that might lead to the altered contractility of the SHR arterial muscle. In one study, myofibrillar protein was extracted from 16- to 18-week-old SHR and WKY caudal arterial muscle. The Mg(2+)-activated ATPase activity was measured under conditions where the Ca2+ concentration was controlled. In another study, the amount of myosin present and relative proportions of the myosin heavy chain (MHC) isoforms were determined by quantitative SDS-PAGE using heavy molecular weight standards and bovine serum albumin as the standard for concentration. In a third study, MLC20 phosphorylation levels in electrically stimulated arterial muscle were determined by urea glycerol gel electrophoresis and Western blot analyses. The SHR (n = 6) myofibrillar ATPase liberated 0.011 +/- 0.003 mumol Pi/mg myosin/min, which was significantly more than the 0.006 +/- 0.001 mumol Pi/mg myosin/min liberated by the WKY (n = 4) myofibrillar ATPase (P < 0.05). Consistent with the increased ATPase activity, phosphorylation of MLC20 was increased by 2.8 times as much in the SHR compared with the WKY electrically stimulated arterial muscle. However, there was no difference in MHC isoform pattern in the SHR compared with the WKY arterial muscle in contrast to the findings of at least one other laboratory. This discrepancy is discussed. The data reviewed in this article lead to the conclusions that an increased actin-activated myosin ATPase activity and MLC20 phosphorylation are likely responsible for the increased velocity of shortening previously reported in SHR arterial muscle and the increased ATPase activity is not a function of an increased myosin content or of altered MHC isoform pattern in the SHR muscle.
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PMID:Arterial muscle myosin heavy chains and light chains in spontaneous hypertension. 918 11

The small GTPase Rho is implicated in cytoskeletal rearrangements including stress fiber and focal adhesion formation and in the transcriptional activation of c-fos serum response element. In vitro, Rho-kinase, which is activated by Rho, phosphorylates not only myosin light chain (MLC) (thereby activating myosin ATPase) but also myosin phosphatase, thus inactivating myosin phosphatase. Rho-kinase is involved in the formation of stress fibers and focal adhesions in fibroblasts. Here we show that the expression of constitutively active Rho-kinase increased the level of MLC phosphorylation. The activity of Rho-kinase was necessary for maintaining the vinculin-containing focal adhesions, whereas organized actin stress fibers were not necessary for this. The microinjection of constitutively active Rho-kinase into fibroblasts induced the formation of focal adhesions to some extent under the conditions where organized actin stress fibers were disrupted. The expression of constitutively active Rho-kinase also stimulated the transcriptional activity of c-fos serum response element. These results suggest that Rho-kinase has distinct roles in divergent pathways downstream of Rho, which include MLC phosphorylation leading to stress fiber formation, focal adhesion formation, and gene expression.
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PMID:Cytoskeletal rearrangements and transcriptional activation of c-fos serum response element by Rho-kinase. 931 22

The Drosophila spaghetti squash (sqh) gene encodes the regulatory myosin light chain (RMLC) of nonmuscle myosin II. Biochemical analysis of vertebrate nonmuscle and smooth muscle myosin II has established that phosphorylation of certain amino acids of the RMLC greatly increases the actin-dependent myosin ATPase and motor activity of myosin in vitro. We have assessed the in vivo importance of these sites, which in Drosophila correspond to serine-21 and threonine-20, by creating a series of transgenes in which these specific amino acids were altered. The phenotypes of the transgenes were examined in an otherwise null mutant background during oocyte development in Drosophila females. Germ line cystoblasts entirely lacking a functional sqh gene show severe defects in proliferation and cytokinesis. The ring canals, cytoplasmic bridges linking the oocyte to the nurse cells in the egg chamber, are abnormal, suggesting a role of myosin II in their establishment or maintenance. In addition, numerous aggregates of myosin heavy chain accumulate in the sqh null cells. Mutant sqh transgene sqh-A20, A21 in which both serine-21 and threonine-20 have been replaced by alanines behaves in most respects identically to the null allele in this system, with the exception that no heavy chain aggregates are found. In contrast, expression of sqh-A21, in which only the primary phosphorylation target serine-21 site is altered, partially restores functionality to germ line myosin II, allowing cystoblast division and oocyte development, albeit with some cytokinesis failure, defects in the rapid cytoplasmic transport from nurse cells to cytoplasm characteristic of late stage oogenesis, and some damaged ring canals. Substituting a glutamate for the serine-21 (mutant sqh-E21) allows oogenesis to be completed with minimal defects, producing eggs that can develop normally to produce fertile adults. Flies expressing sqh-A20, in which only the secondary phosphorylation site is absent, appear to be entirely wild type. Taken together, this genetic evidence argues that phosphorylation at serine-21 is critical to RMLC function in activating myosin II in vivo, but that the function can be partially provided by phosphorylation at threonine-20.
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PMID:Myosin light chain-activating phosphorylation sites are required for oogenesis in Drosophila. 941 74

This review focuses on experiments in which the single turnover of myosin-bound ADP is used to characterize the regulation of the cross-bridge cycle by myosin light chain phosphorylation in mammalian smooth muscle. Under isometric conditions, at rest, when the myosin light chain is not phosphorylated, myosin cycles very slowly (about 0.004 s-1), while phosphorylation of the light chain results in a 50-fold increase in cycling rate of 0.2 s-1. Experiments consistently show that some myosin does not increase its cycling rate although its light chain is phosphorylated. Studies at low levels of myosin light chain phosphorylation show that phosphorylation also induces an increase in the cycling rate of unphosphorylated myosin. The fast cycling phosphorylated myosin is the main determinant of suprabasal myosin ATPase activity, while the cycling rate of cooperatively activated unphosphorylated myosin is slow and appears to depend on the extent of phosphorylation of the entire thick filament. Single turnover experiments measuring the rate of phosphorylation and dephosphorylation of myosin light chain show that the turnover of light chain phosphate can be very rapid (0.3-0.4 s-1) at suprabasal calcium concentrations. The expected effect of such a rapid turnover of light chain phosphorylation on the turnover of myosin-bound ADP is not observed. The effects of low levels of myosin light chain phosphorylation on the single turnover of myosin suggest that the same small pool of myosin remains phosphorylated for relatively long periods of time rather than the entire pool of myosin spending a small fraction of its cycle time in the phosphorylated state.
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PMID:Control of cross-bridge cycling by myosin light chain phosphorylation in mammalian smooth muscle. 988 63

Smooth muscle contraction has a relatively high requirement for free magnesium (Mg2+). In this study we examined the effect of Mg2+ concentration ([Mg2+]) on Ca2+-dependent stress development and stress maintenance, myosin ATPase activity, and myosin light chain (MLC) phosphorylation levels in Triton X-100 detergent-skinned fibers of the swine carotid media. Increasing [Mg2+] in a stepwise fashion from 0.1 to 6 mM 1) decreased the magnitude and Ca2+ sensitivity of stress development but augmented the amount of stress maintained without proportional MLC phosphorylation, 2) produced a greater decrease in the Ca2+ sensitivity of MLC phosphorylation than that of stress development, and 3) decreased myosin ATPase activity. These findings demonstrate that Mg2+ differentially modulates the MLC phosphorylation-dependent development of stress and the MLC phosphorylation-independent maintenance of stress. We suggest that increases in [Mg2+] enhance stress maintenance by increasing [MgADP], thus increasing the number of cross bridges in a force-generating state, and by a direct effect on the pathway responsible for Ca2+-dependent, MLC phosphorylation-independent contractions.
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PMID:Effect of Mg2+ on stress, myosin phosphorylation, and ATPase activity in detergent-skinned swine carotid media. 1033 Feb 23

The basally located actin cytoskeleton has been demonstrated previously to regulate Cl- secretion from intestinal epithelia via its effects on the Na+-K+-2Cl- cotransporter (NKCC1). In nontransporting epithelia, inhibition of myosin light chain kinase (MLCK) prevents cell-shrinkage-induced activation of NKCC1. The aim of this study was to investigate the role of myosin in the regulation of secretagogue-stimulated Cl- secretion in intestinal epithelia. The human intestinal epithelial cell line T84 was used for these studies. Prevention of myosin light chain phosphorylation with the MLCK inhibitor ML-9 or ML-7 and inhibition of myosin ATPase with butanedione monoxime (BDM) attenuated cAMP but not Ca2+-mediated Cl- secretion. Both ML-9 and BDM diminished cAMP activation of NKCC1. Neither apical Cl- channel activity, basolateral K+ channel activity, nor Na+-K+-ATPase were affected by these agents. Cytochalasin D prevented such attenuation. cAMP-induced rearrangement of basal actin microfilaments was prevented by both ML-9 and BDM. The phosphorylation of mosin light chain and subsequent contraction of basal actin-myosin bundles are crucial to the cAMP-driven activation of NKCC1 and subsequent apical Cl- efflux.
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PMID:Myosin regulation of NKCC1: effects on cAMP-mediated Cl- secretion in intestinal epithelia. 1048 31

There are numerous causes for slow or delayed wound healing. Because slowly healing wounds are often inflamed, we quantitated the inflammatory chemokine, interleukin-8, produced by slowly healing human burn wounds and compared this to interleukin-8 from healed wounds and normal intact skin. Interleukin-8 levels were increased significantly in unhealed wounds (19.7 ng/ml) compared to healed wounds (7.7 ng/ml) or normal skin (5.7 ng/ml). Interleukin-8 in these ranges was added to adult human keratinocytes and fibroblasts. Interleukin-8 significantly decreased keratinocyte replication but had no effect on fibroblast replication. The rate or final degree of fibroblast populated collagen lattice contraction was inhibited at interleukin-8 concentrations between 10 and 30 ng/ml, but not altered at concentrations below 10 ng/ml and above 100 ng/ml. The concurrent application of indomethacin at 10 microg/ml reversed this interleukin-8 induced inhibition. Interleukin-8 inhibited myosin ATPase activity, apparently by reducing the phosphorylation of nonmuscle myosin light chain. We conclude that elevated levels of interleukin-8 may be found during delayed healing, and these elevated interleukin-8 levels may directly contribute to retarded wound closure.
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PMID:Interleukin-8 levels and activity in delayed-healing human thermal wounds. 1088 12

Myosin light chain kinase (MLCK) is a regulatory protein for smooth muscle contraction, which acts by phosphorylating 20-kDa myosin light chain (MLC20) to activate the myosin ATPase activity. Although this mode of action is well-established, there are numerous reports of smooth muscle contraction that is not associated with MLC20 phosphorylation. The kinase activity for the phosphorylation is localized at the central part of MLCK, which is also furnished with actin-binding activity at its N terminal and myosin-binding activity at its C terminal. This article overviews as to how such multifunctional properties of MLCK modify the actin-myosin interaction and presents our observations that the phosphorylation is not obligatory in induction of smooth muscle contraction.
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PMID:Myosin light chain kinase as a multifunctional regulatory protein of smooth muscle contraction. 1175

Seasonal cooling can modify the thermal preferenda of ectothermic vertebrates and elicit a variety of physiological responses ranging from winter dormancy to an acclimation response that partially compensates for the effects of low temperature on activity. Partial compensation of activity levels is particularly common in aquatic species for which seasonal temperature changes provide a stable cue for initiating the response. Thermal plasticity of locomotory performance has evolved independently on numerous occasions, and there is considerable phylogenetic diversity with respect to the mechanisms at the physiological and molecular levels. In teleosts, neuromuscular variables that can be modified include the duration of motor nerve stimulation, muscle activation and relaxation times, maximum force and unloaded shortening velocity (V(max)), although not all are modified in every species. Thermal plasticity in V(max) has been associated with changes in myosin ATPase activity and myosin heavy chain (MyHC) composition and/or with a change in the ratio of myosin light chain isoforms. In common carp (Cyprinus carpio), there are continuous changes in phenotype with acclimation temperature at lower levels of organisation, such as MyHC composition and V(max), but a distinct threshold for an effect in terms of locomotory performance. Thus, there is no simple relationship between whole-animal performance and muscle phenotype. The nature and magnitude of temperature acclimation responses also vary during ontogeny. For example, common carp acquire the ability to modify MyHC composition with changes in acclimation temperature during the juvenile stage. In contrast, the thermal plasticity of swimming performance observed in tadpoles of the frog Limnodynastes peronii is lost in the terrestrial adult stage. Although it is often assumed that the adjustments in locomotory performance associated with temperature acclimation enhance fitness, this has rarely been tested experimentally. Truly integrative studies of temperature acclimation are scarce, and few studies have considered both sensory and motor function in evaluating behavioural responses. Developmental plasticity is a special case of a temperature acclimation response that can lead to temporary or permanent changes in morphology and/or physiological characteristics that affect locomotory performance.
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PMID:Thermal plasticity of skeletal muscle phenotype in ectothermic vertebrates and its significance for locomotory behaviour. 1211 Jun 64

The adapter protein paxillin localizes to the focal adhesions of adherent cells and has been implicated in the regulation of cytoskeletal organization and cell motility. Paxillin undergoes tyrosine phosphorylation in response to the contractile stimulation of tracheal smooth muscle. We therefore hypothesized that paxillin may be involved in regulating smooth muscle contraction. Tracheal smooth muscle strips were treated with paxillin antisense oligonucleotides to inhibit the expression of paxillin protein selectively. Paxillin antisense or sense was introduced into muscle strips by reversible permeabilization and strips were incubated with antisense or sense for 3 days. Paxillin antisense selectively depressed paxillin expression, but it did not affect the expression of vinculin, focal adhesion kinase, myosin light chain kinase, myosin heavy chain or myosin light chain. Tension development in response to stimulation with ACh or KCl was markedly depressed in paxillin-depleted muscle strips. Active force and paxillin protein expression were restored by incubation of antisense-treated strips in the absence of oligonucleotides. The depletion of paxillin did not inhibit the increase in intracellular free Ca2+, myosin light chain phosphorylation or myosin ATPase activity in response to contractile stimulation. The concentration of G-actin was significantly lower in unstimulated paxillin-depleted smooth muscle tissues than in normal tissues. While stimulation with acetylcholine caused a decrease in G-actin in normal muscle strips, it caused little change in the G-actin concentration in paxillin-depleted muscle strips, suggesting that paxillin is necessary for normal actin dynamics in smooth muscle. We conclude that paxillin is required for active tension development in smooth muscle, but that it does not regulate increases in intracellular Ca2+, myosin light chain phosphorylation or myosin ATPase activity during contractile stimulation. Paxillin may be important in regulating actin filament dynamics and organization during smooth muscle contraction.
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PMID:The focal adhesion protein paxillin regulates contraction in canine tracheal smooth muscle. 1212 48


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