Estrogen, Myosin Phosphorylation and Muscle Thermogenesis

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The long term goals of my research program are to further our understanding of how the posttranslational modification of myosin by phosphorylation modulates contractile function of skeletal muscle.  In mammalian striated muscle the transfer of a phosphate moiety from adenosine triphosphate (i.e. ATP) to a serine residue on the myosin regulatory light chain (RLC) is catalyzed by skeletal myosin light chain kinase (skMLCK). This reaction is regulated by a calcium and calmodulin signalling cascade that is activated during  contraction.  In turn, skMLCK catalyzed phosphorylation of the myosin RLC increases the calcium sensitivity of the contractile proteins, thus increasing submaximal force and producing what is known as "potentiation".   Strong associations between contraction-induced RLC phosphorylation and potentiation have been demonstrated in a variety of rodent fast twitch skeletal muscle models.  By contrast, although rodent slow twitch muscle has a limited enzymatic ability to phosphorylate the RLC they display no potentiation; accordingly, the presence of basal RLC phosphorylation in the slow twitch muscle phenotype may serve a different purpose that may or may not be shared with the fast muscle phenotype.  Because the same calcium signals that regulate muscle contraction also activate skMLCK activity it is possible that working skeletal muscle most often operates in the potentiated state produced by myosin RLC phosphorylation. Interesting in this regard is the possibility that skMLCK activity and thus RLC phosphorylation may be sensitive to hormonal influences, a signalling pathway linking system and muscle physiology. Accordingly, the goals of this application are to examine how hormones such as estrogen may influence myosin RLC phosphorylation (and thus potentiation) of mouse fast twitch skeletal muscle (as suggested by Lai et al. 2016) and if this influence involves skMLCK signalling. Another way we will study the system physiology-muscle physiology link is by examining how myosin RLC phosphorylation modulates muscle metabolic rate and heat production, i.e. thermogenesis.  The putative ability of myosin RLC phosphorylation to regulate muscle thermogenesis (McNamara et al. 2015) may influence both resting and shivering thermogenesis.  A key feature of all of the experiments is the use of skMLCK "knockout" mice from our colony to provide a robust negative control condition that is simply not feasible with wildtype mice. Thus, we will be able to critically examine how estrogen effects muscle function in the presence and/or absence of myosin RLC phosphorylation. We will also be able to assess the influence of environmental interventions such as diet and cold on resting and shivering thermogenesis, respectively, in mice with and without myosin RLC phosphorylation in their skeletal muscles. These studies will thus examine pathways regulating myosin RLC phosphorylation and how, in turn, this molecular mechanism may regulate muscle metabolism.