scholarly journals The direct molecular effects of fatigue and myosin regulatory light chain phosphorylation on the actomyosin contractile apparatus

2010 ◽  
Vol 298 (4) ◽  
pp. R989-R996 ◽  
Author(s):  
Michael J. Greenberg ◽  
Tanya R. Mealy ◽  
Michelle Jones ◽  
Danuta Szczesna-Cordary ◽  
Jeffrey R. Moore
Keyword(s):  
Skeletal Muscle ◽  
Light Chain ◽  
Molecular Species ◽  
Regulatory Light Chain ◽  
Myosin Regulatory Light Chain ◽  
Sliding Velocity ◽  
In Vitro Motility Assay ◽  
Individual Molecular Species ◽  
Molecular Effects

Skeletal muscle, during periods of exertion, experiences several different fatigue-based changes in contractility, including reductions in force, velocity, power output, and energy usage. The fatigue-induced changes in contractility stem from many different factors, including alterations in the levels of metabolites, oxidative damage, and phosphorylation of the myosin regulatory light chain (RLC). Here, we measured the direct molecular effects of fatigue-like conditions on actomyosin's unloaded sliding velocity 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 and whether the effects of each individual molecular species are additive. We found that the primary causes of the reduction in unloaded sliding velocity are increased [ADP] and lowered pH and that the combined effects of the molecular species are nonadditive. Furthermore, since an increase in RLC phosphorylation is often associated with fatigue, we examined the differential effects of myosin RLC phosphorylation and fatigue on actin filament velocity. We found that phosphorylation of the RLC causes a 22% depression in sliding velocity. On the other hand, RLC phosphorylation ameliorates the slowing of velocity under fatigue-like conditions. We also found that phosphorylation of the myosin RLC increases actomyosin affinity for ADP, suggesting a kinetic role for RLC phosphorylation. Furthermore, we showed that ADP binding to skeletal muscle is load dependent, consistent with the existence of a load-dependent isomerization of the ADP bound state.

scholarly journals The molecular effects of skeletal muscle myosin regulatory light chain phosphorylation

2009 ◽  
Vol 297 (2) ◽  
pp. R265-R274 ◽  
Author(s):  
Michael J. Greenberg ◽  
Tanya R. Mealy ◽  
James D. Watt ◽  
Michelle Jones ◽  
Danuta Szczesna-Cordary ◽  
...  
Keyword(s):  
Skeletal Muscle ◽  
Light Chain ◽  
Duty Cycle ◽  
Isometric Force ◽  
Thick Filament ◽  
Regulatory Light Chain ◽  
Myosin Regulatory Light Chain ◽  
Skeletal Muscle Myosin ◽  
Molecular Effects ◽  
Muscle Myosin

Phosphorylation of the myosin regulatory light chain (RLC) in skeletal muscle has been proposed to act as a molecular memory of recent activation by increasing the rate of force development, ATPase activity, and isometric force at submaximal activation in fibers. It has been proposed that these effects stem from phosphorylation-induced movement of myosin heads away from the thick filament backbone. In this study, we examined the molecular effects of skeletal muscle myosin RLC phosphorylation using in vitro motility assays. We showed that, independently of the thick filament backbone, the velocity of skeletal muscle myosin is decreased upon phosphorylation due to an increase in the myosin duty cycle. Furthermore, we did not observe a phosphorylation-dependent shift in calcium sensitivity in the absence of the myosin thick filament. These data suggest that phosphorylation-induced movement of myosin heads away from the thick filament backbone explains only part of the observed phosphorylation-induced changes in myosin mechanics. Last, we showed that the duty cycle of skeletal muscle myosin is strain dependent, consistent with the notion that strain slows the rate of ADP release in striated muscle.

scholarly journals Orientation of the N-Terminal Lobe of the Myosin Regulatory Light Chain in Skeletal Muscle Fibers

2012 ◽  
Vol 102 (6) ◽  
pp. 1418-1426 ◽  
Author(s):  
Daniela Romano ◽  
Birgit D. Brandmeier ◽  
Yin-Biao Sun ◽  
David R. Trentham ◽  
Malcolm Irving
Keyword(s):  
Skeletal Muscle ◽  
Light Chain ◽  
Muscle Fibers ◽  
Regulatory Light Chain ◽  
Myosin Regulatory Light Chain ◽  
Skeletal Muscle Fibers

scholarly journals Fluorescence Polarization Transients from Rhodamine Isomers on the Myosin Regulatory Light Chain in Skeletal Muscle Fibers

1998 ◽  
Vol 74 (6) ◽  
pp. 3093-3110 ◽  
Author(s):  
Seth C. Hopkins ◽  
Cibele Sabido-David ◽  
John E.T. Corrie ◽  
Malcolm Irving ◽  
Yale E. Goldman
Keyword(s):  
Skeletal Muscle ◽  
Light Chain ◽  
Fluorescence Polarization ◽  
Muscle Fibers ◽  
Regulatory Light Chain ◽  
Myosin Regulatory Light Chain ◽  
Skeletal Muscle Fibers

scholarly journals Cardiac myosin regulatory light chain kinase modulates cardiac contractility by phosphorylating both myosin regulatory light chain and troponin I

2020 ◽  
Vol 295 (14) ◽  
pp. 4398-4410 ◽  
Author(s):  
Ivanka R. Sevrieva ◽  
Birgit Brandmeier ◽  
Saraswathi Ponnam ◽  
Mathias Gautel ◽  
Malcolm Irving ◽  
...  
Keyword(s):  
Light Chain ◽  
Troponin I ◽  
Regulatory Light Chain ◽  
Myosin Regulatory Light Chain ◽  
Cardiac Myosin ◽  
Sarcomeric Proteins ◽  
Ventricular Trabeculae

Heart muscle contractility and performance are controlled by posttranslational modifications of sarcomeric proteins. Although myosin regulatory light chain (RLC) phosphorylation has been studied extensively in vitro and in vivo, the precise role of cardiac myosin light chain kinase (cMLCK), the primary kinase acting upon RLC, in the regulation of cardiomyocyte contractility remains poorly understood. In this study, using recombinantly expressed and purified proteins, various analytical methods, in vitro and in situ kinase assays, and mechanical measurements in isolated ventricular trabeculae, we demonstrate that human cMLCK is not a dedicated kinase for RLC but can phosphorylate other sarcomeric proteins with well-characterized regulatory functions. We show that cMLCK specifically monophosphorylates Ser23 of human cardiac troponin I (cTnI) in isolation and in the trimeric troponin complex in vitro and in situ in the native environment of the muscle myofilament lattice. Moreover, we observed that human cMLCK phosphorylates rodent cTnI to a much smaller extent in vitro and in situ, suggesting species-specific adaptation of cMLCK. Although cMLCK treatment of ventricular trabeculae exchanged with rat or human troponin increased their cross-bridge kinetics, the increase in sensitivity of myofilaments to calcium was significantly blunted by human TnI, suggesting that human cTnI phosphorylation by cMLCK modifies the functional consequences of RLC phosphorylation. We propose that cMLCK-mediated phosphorylation of TnI is functionally significant and represents a critical signaling pathway that coordinates the regulatory states of thick and thin filaments in both physiological and potentially pathophysiological conditions of the heart.

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