The Molecular Effects of Skeletal Muscle Fatigue on Myosin Mechanics and Kinetics

Abstract

Skeletal muscle, during periods of exertion, experiences several different fatigue based changes in contraction including reductions in force, velocity, power output, and energy usage. The physiological bases of fatigue induced changes in contractility stem from many different factors including changes in neuronal activity, excitation-contraction coupling, and actomyosin contraction. The direct changes in the actomyosin contractile complex have previously been shown to be the result of alterations in the levels of metabolites, oxidative damage, and an increase in myosin regulatory light chain phosphorylation. Here, we measured the direct molecular effects of fatigue-like conditions on actomyosin velocity and force generation using the in vitro motility assay. We examined how changes in ATP, ADP, Pi, and pH affect the ability of the myosin to translocate actin. We show that fatigue-induced increases in metabolites decrease unloaded shortening velocity, with ADP and pH contributing the most to the observed changes in velocity. We also examined whether myosin regulatory light chain phosphorylation alters the sensitivity of the myosin to fatigue-like conditions. We show that under fatigue-like conditions, phosphorylation of the myosin regulatory light chian enhances force production and reduces actin sliding velocity, similar to the effects of phosphorylation under fatigue-like conditions observed in both muscle fiber and in vitro motility studies. Furthermore, we found that force production by dephosphorylated myosin is very sensitive to fatigue-like conditions whereas force production by phosphorylated myosin is rather fatigue insensitive. These results suggest that phosphorylation of the myosin regulatory light chain in skeletal muscle may serve as a protective mechanism against fatigue. Supported by NIH-HL077280, AHA-0435434T (J.R.M.), AHA-0815704D (to M.J.G.), and NIH-HL071778 (D.S-C.).