Modulation of the actomyosin interaction during fatigue of skeletal muscle

Cooke, Roger

doi:10.1002/mus.20891

Cited by 57 publications

(78 citation statements)

References 185 publications

(286 reference statements)

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“…Since our conditions of high P i and low pH are similar to those experienced during intense contractile activity (6), the results also have potential implications for the molecular basis of muscular fatigue. During fatigue, elevated levels of P i and H ϩ are thought to act additively to inhibit contraction, largely based on the effects on maximal isometric force (6). Since force is related to the number of attached cross bridges, our suggestion of a P iinduced detachment implies that the increase in velocity may come at the expense of force; thus our finding remains consistent with the notion that these ions can additively inhibit muscular force.…”

Section: Perspectives and Significancesupporting

confidence: 86%

Phosphate enhances myosin-powered actin filament velocity under acidic conditions in a motility assay

Debold

Turner

Stout

et al. 2011

American Journal of Physiology-Regulatory, Integrative and Comp

102

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Debold EP, Turner MA, Stout JC, Walcott S. Phosphate enhances myosin-powered actin filament velocity under acidic conditions in a motility assay. Am J Physiol Regul Integr Comp Physiol 300: R1401-R1408, 2011. First published February 23, 2011 doi:10.1152/ajpregu.00772.2010.-Elevated levels of inorganic phosphate (Pi) are believed to inhibit muscular force by reversing myosin's force-generating step. These same levels of Pi can also affect muscle velocity, but the molecular basis underlying these effects remains unclear. We directly examined the effect of Pi (30 mM) on skeletal muscle myosin's ability to translocate actin (Vactin) in an in vitro motility assay. Manipulation of the pH enabled us to probe rebinding of Pi to myosin's ADP-bound state, while changing the ATP concentration probed rebinding to the rigor state. Surprisingly, the addition of Pi significantly increased Vactin at both pH 6.8 and 6.5, causing a doubling of Vactin at pH 6.5. To probe the mechanisms underlying this increase in speed, we repeated these experiments while varying the ATP concentration. At pH 7.4, the effects of Pi were highly ATP dependent, with Pi slowing Vactin at low ATP (Ͻ500 M), but with a minor increase at 2 mM ATP. The Pi-induced slowing of Vactin, evident at low ATP (pH 7.4), was minimized at pH 6.8 and completely reversed at pH 6.5. These data were accurately fit with a simple detachment-limited kinetic model of motility that incorporated a Pi-induced prolongation of the rigor state, which accounted for the slowing of Vactin at low ATP, and a Pi-induced detachment from a strongly bound post-power-stroke state, which accounted for the increase in Vactin at high ATP. These findings suggest that Pi differentially affects myosin function: enhancing velocity, if it rebinds to the ADP-bound state, while slowing velocity, if it binds to the rigor state.fatigue; cross-bridge cycle; muscle contraction MUSCULAR FORCE AND MOTION are generated through the cyclical interaction of actin and myosin, in a process ultimately powered by the hydrolysis of ATP (Fig. 1). In the 1980s, several investigations established that elevated levels of organic phosphate (P i ) reduced muscular force, leading to the notion that P i release by myosin was closely associated with force generation (12) and the rotation of the lever arm (2). This hypothesis postulates that P i rebinds to myosin (M) in a state in which myosin is strongly bound to both actin (A) and ADP (AM.ADP) and, in one or more steps, reverses the forcegenerating step, leading to an increase in the population of the weakly bound myosin, in the M.ADP.P i state (32). While competing theories exist (3), models based on the P i -induced detachment can account for much of the effect on muscular force (26). However, the effects of P i on muscle's shortening velocity have been more difficult to explain using the same model.While the earliest experiments in muscle fibers suggested that P i had no effect on unloaded shortening velocity (V us ) (7), subsequent efforts revealed that P i could induc...

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Section: Perspectives and Significancesupporting

confidence: 86%

Phosphate enhances myosin-powered actin filament velocity under acidic conditions in a motility assay

Debold

Turner

Stout

et al. 2011

American Journal of Physiology-Regulatory, Integrative and Comp

102

View full text Add to dashboard Cite

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“…It is known that the inhibition in function involves, in part, mechanisms that directly affect the interaction of actin and myosin, and these include product inhibition due to the accumulation of metabolites. Although replication of these metabolite levels in vitro results in a decrease in shortening velocities, the observed inhibition does not account for the extent seen in vivo (Fitts 1994;Westerblad et al 1997Westerblad et al , 2002Myburgh 2004;Cooke 2007).…”

Section: Introductionmentioning

confidence: 83%

“…Numerous studies have investigated the mechanism of inhibition but the precise mechanism is unknown (Fitts 1994;Westerblad and Allen 2003;Cooke 2007). Fatigue affects every system involved in muscle mechanics.…”

Section: Introductionmentioning

confidence: 99%

Myosin regulatory light chain phosphorylation inhibits shortening velocities of skeletal muscle fibers in the presence of the myosin inhibitor blebbistatin

Stewart

Franks-Skiba

Cooke

2009

J Muscle Res Cell Motil

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Phosphorylation of skeletal myosin regulatory light chain (RLC) occurs in fatigue and may play a role in the inhibition of shortening velocities observed in vivo. Forces and shortening velocities were measured in permeabilized rabbit psoas fibers with either phosphorylated or dephosphorylated RLCs and in the presence or absence of the myosin inhibitor blebbistatin. Addition of 20 lM blebbistatin decreased tensions by *80% in fibers, independent of phosphorylation. In blebbistatin maximal shortening velocities (V max ) at 30°C, were decreased by 45% (3.2 ± 0.34 vs. 5.8 ± 0.18 lengths/s) in phosphorylated fibers but were not inhibited in dephosphorylated fibers (6.0 ± 0.30 vs. 5.4 ± 0.30). In the presence of 20 lM blebbistatin, K m for V max as a function of [ATP] was lower for phosphorylated fibers than for dephosphorylated fibers (50 ± 20 vs. 330 ± 84 lM) indicating that the apparent binding of ATP is stronger in these fibers. Phosphorylation of RLC in situ during fiber preparation or by addition of myosin light chain kinase yielded similar data. RLC phosphorylation inhibited velocity in blebbistatin at both 30 and 10°C, unlike previous reports where RLC phosphorylation only affected shortening velocities at higher temperatures.

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“…It has also been shown that both phosphorylation of the RLC (7,8) and the presence of bound cation (9) play important roles in the structure and function of myosin, reinforcing the notion that there is a high degree of interdomain communication between these sites.…”

mentioning

confidence: 82%

Cardiomyopathy-linked myosin regulatory light chain mutations disrupt myosin strain-dependent biochemistry

Greenberg

Kazmierczak

Szczesna‐Cordary

et al. 2010

Proc. Natl. Acad. Sci. U.S.A.

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Familial hypertrophic cardiomyopathy (FHC) is caused by mutations in sarcomeric proteins including the myosin regulatory light chain (RLC). Two such FHC mutations, R58Q and N47K, located near the cationic binding site of the RLC, have been identified from population studies. To examine the molecular basis for the observed phenotypes, we exchanged endogenous RLC from native porcine cardiac myosin with recombinant human ventricular wild type (WT) or FHC mutant RLC and examined the ability of the reconstituted myosin to propel actin filament sliding using the in vitro motility assay. We find that, whereas the mutant myosins are indistinguishable from the controls (WT or native myosin) under unloaded conditions, both R58Q-and N47K-exchanged myosins show reductions in force and power output compared with WT or native myosin. We also show that the changes in loaded kinetics are a result of mutation-induced loss of myosin strain sensitivity of ADP affinity. We propose that the R58Q and N47K mutations alter the mechanical properties of the myosin neck region, leading to altered loaddependent kinetics that may explain the observed mutant-induced FHC phenotypes.in vitro motility | load-dependent kinetics | familial hypertrophic cardiomyopathy | R58Q | N47K F amilial hypertrophic cardiomyopathy (FHC), the leading cause of sudden cardiac death in people under 30 (1), is characterized by several changes in cardiac structure including myofibrillar disarray and thickening of the left ventricle, papillary muscles, and/or septum. FHC is a disease of the sarcomere, resulting from mutations of cardiac proteins (reviewed in refs. 1 and 2) including the regulatory (RLC) and essential (ELC) light chains of myosin.Each myosin molecule is a hexamer composed of two myosin heavy chains (MHC), two RLCs, and two ELCs (3). The α-helical neck region of the MHC has been proposed to act as a lever arm, amplifying small conformational changes that originate at the myosin catalytic site into large movements needed to produce contractile force. The α-helical neck, supported by the two light chains, has been proposed to function as a compliant element (4-6), with the light chains imparting stiffness to the lever arm. Therefore, disruption of the light chain binding to the MHC could conceivably impair force generation as well as the transmission of external loads to the myosin active site, leading to a loss of myosin strain sensitivity.The RLC is a calmodulin homology protein that contains an EF-hand Ca 2+ -Mg 2+ binding site and also a highly conserved phosphorylatable serine, both sites located at the N terminus of the RLC. It has also been shown that both phosphorylation of the RLC (7, 8) and the presence of bound cation (9) play important roles in the structure and function of myosin, reinforcing the notion that there is a high degree of interdomain communication between these sites.Given the functional importance of the RLC-containing neck domain of myosin, it is not surprising that mutations of the RLC can cause FHC. Population studies have i...

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Modulation of the actomyosin interaction during fatigue of skeletal muscle

Cited by 57 publications

References 185 publications

Phosphate enhances myosin-powered actin filament velocity under acidic conditions in a motility assay

Phosphate enhances myosin-powered actin filament velocity under acidic conditions in a motility assay

Myosin regulatory light chain phosphorylation inhibits shortening velocities of skeletal muscle fibers in the presence of the myosin inhibitor blebbistatin

Cardiomyopathy-linked myosin regulatory light chain mutations disrupt myosin strain-dependent biochemistry

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