Phosphorylation of the regulatory light chains of myosin affects Ca²⁺sensitivity of skeletal muscle contraction

Abstract

The role of phosphorylation of the myosin regulatory light chains (RLC) is well established in smooth muscle contraction, but in striated (skeletal and cardiac) muscle its role is still controversial. We have studied the effects of RLC phosphorylation in reconstituted myosin and in skinned skeletal muscle fibers where Ca2+ sensitivity and the kinetics of steady-state force development were measured. Skeletal muscle myosin reconstituted with phosphorylated RLC produced a much higher Ca2+ sensitivity of thin filament-regulated ATPase activity than nonphosphorylated RLC (change in -log of the Ca2+ concentration producing half-maximal activation = approximately 0.25). The same was true for the Ca2+ sensitivity of force in skinned skeletal muscle fibers, which increased on reconstitution of the fibers with the phosphorylated RLC. In addition, we have shown that the level of endogenous RLC phosphorylation is a crucial determinant of the Ca2+ sensitivity of force development. Studies of the effects of RLC phosphorylation on the kinetics of force activation with the caged Ca2+, DM-nitrophen, showed a slight increase in the rates of force development with low statistical significance. However, an increase from 69 to 84% of the initial steady-state force was observed when nonphosphorylated RLC-reconstituted fibers were subsequently phosphorylated with exogenous myosin light chain kinase. In conclusion, our results suggest that, although Ca2+ binding to the troponin-tropomyosin complex is the primary regulator of skeletal muscle contraction, RLC play an important modulatory role in this process.

Phosphorylation of Myosin Regulatory Light Chain by Myosin Light Chain Kinase, and Muscle Contraction

“…However, enhancement of contractility by phosphorylation of MLC is obvious under extreme experimental conditions, such as the comparison of completely unphosphorylated MLC by phosphatase and sufficiently phosphorylated MLC by skMLCK. 9 These data suggest that the basic function of MLCK as enhancing contraction is still preserved in skMLCK.…”

Familial Hypertrophic Cardiomyopathy-linked Alterations in Ca2+ Binding of Human Cardiac Myosin Regulatory Light Chain Affect Cardiac Muscle Contraction

“…The affinity of this site for Ca 2ϩ in the presence of 2 mM Mg 2ϩ was shown to be ϳ2-fold lower (30). It is not known whether the site inactivated for Ca 2ϩ does or does not bind Mg 2ϩ .…”

Diversity of human skeletal muscle in health and disease: Contribution of proteomics

“…Differences were also observed in myosin light chain isoform distribution with changing abundance of slow and fast isoforms. Concentrations of fast isoforms of myosin regulatory light chains (MLC) and, in particular, of the regulatory phosphorylatable isoforms known to modulate the rate of force development and of twitch maximal force at different initial tension levels [81][82][83], both of which are relevant to muscle function, were decreased. In vitro phosphorylation of MLC is known to increase calcium sensitivity thereby controlling the rate of force development and maximal force [84].…”