The formation of skeletal muscle and its adaptation to the environment requires precise temporal and spatial regulation of a host of proteins, including the molecular motor protein, myosin. The precise adaptation of myosin heavy chain (MyHC) genes requires coordinate regulation, yet, little is known about its molecular biology. We propose to define the molecular aspects of fiber type specificity and the pathways that regulate these genes. In mammals, there are 6 characterized skeletal muscle MyHC genes. Although muscle fibers expressing each of them have unique contractile velocities, the enzymatic properties of the individual motors remain elusive. We will express the 6 human skeletal MyHC head domains in an inducible mammalian system and characterize their biochemical and biophysical properties. Despite the perception that the sarcomeric MyHC gene family had been defined, examination of the human genome revealed a novel striated MyHC that we propose to characterize. We have found that it is expressed in cardiac and skeletal muscle and that phylogenetically, it appears most closely related to the alpha and beta MyHC genes. We will compare the sequence features of the coding, regulatory regions and the intron/exon organization of this gene in mouse and human. We will also determine its expression in development and in the adult and test whether well-characterized muscle adaptations alter its pattern of expression. Until recently, there had been no diseases associated with mutations in skeletal MyHC. However, a mutation in the MyHC IIa gene has been reported which we propose to model in transgenic mice. We are also characterizing the IId gene of a childhood myopathy patient who appears to be null for its expression. An interesting feature of the MyHC gene family that may have relevance to Duchenne muscular dystrophy (DMD) is that the most abundant MyHC protein in rodents, IIb, is barely detectable in normal adults. However, we find its expression is induced in DMD. Because of the potential functional consequences of expression of this fast myosin motor, we will define the molecular basis for this species difference and its induction. Finally, we will extend our studies of an unusual cell type, the myofibroblast, which has properties of both muscle and nonmuscle cells, including expression of adult fast skeletal MyHCs, to understand the pathways that define these cells and distinguish them from skeletal muscle. ? ?
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