Myosin Phosphates and Myocardin: Regulatory Pathways Modulating Smooth Muscle Contractility and Differentiation

Neppl. Jr., Ronald Lee, Department of Molecular Physiology and Biological Physics, University of Virginia
Somlyo, Avril, Department of Molecular Physiology and Biological Physics, University of Virginia

The cardiovascular system is the organ system responsible for the transport of nutrients, gases and wastes to and from other cells in the body. During development, the heart and circulatory systems are the first organ systems to form. The development and persistence of these organ systems is regulated by the transcriptional programs which maintain the proper gene expression profiles throughout the life of the organism, and likely involve feedback mechanisms in which physiological (dys)function can modulate gene expression. In order to better understand these two disparate processes, we examined the effects of eicosanoid and cyclic nucleotide signaling cascades on the regulation of cerebrovascular tone, as well as the necessity of myocardin for the normal development of cardiac and vascular smooth muscles. Smooth muscle (SM) contractility is controlled via a variety of upstream signaling cascades that ultimately modulate the activities of myosin light chain kinase (MLCK) and myosin light chain phosphatase (MLCP) thereby controlling myosin regulatory light chain (RLC 20 ) phosphorylation and contractile force generation. Normal physiologic vascular tone is a function of the vascular smooth muscle contractile state. As such, we investigated the effects of eicosanoid and cyclic nucleotide signaling on cerebrovascular SM contractility. We find that pretreatment with cGMP, the downstream effector of the endothelium derived relaxing factor nitric oxide, significantly inhibits eicosanoid induced contractile force and RhoA activation, without altering the phosphorylation state of the MLCP targeting subunit normally observed following eicosanoid stimulation alone. We ii conclude that in intact cerebrovascular SM, cyclic nucleotide induced vasorelaxation is not through the conventional mechanism observed in other smooth muscles. The potent serum response factor coactivator myocardin, and its related cofactors, has been shown to be both necessary and sufficient for smooth muscle differentiation in conventional mouse knockouts. However, despite its high expression in cardiomyocytes during embryogenesis, there is limited evidence demonstrating the requirement of myocardin for proper cardiac development. Using a chimeric mouse model, we demonstrate that although myocardin -/- cells fail to populate the ventricle as compared to myocardin +/- controls, those that do ultimately differentiate into cardiomyocytes exhibit a normal ultrastructural organization. In addition, we demonstrate that although myocardin is not required for the development of a functional contractile SMC phenotype, it is required for the development of multiple visceral smooth muscle tissues.

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PHD (Doctor of Philosophy)
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