Title: Molecular regulation of muscle development by Smyd1 Project Summary The goal of this project is to elucidate the molecular mechanisms by which Smyd1b functions to control muscle cell differentiation. Muscle cell differentiation is a complex process that depends on the coordinated gene expression and assembly of myofibrillar proteins into regular arrays of myofibrils that support muscle contraction. Defective myofibril assembly in skeletal and cardiac muscles leads to muscular dystrophy, muscle atrophy, and cardiomyopathy. The molecular regulation of myofibril assembly (myofibrillogenesis) is not well understood, but recent studies have demonstrated that Smyd1, a novel histone methyltransferase, plays a key role in muscle cell differentiation and myofibril assembly. Loss of Smyd1b (a Smyd1 orthologue) in zebrafish results in complete disruption of sarcomere organization and increased muscle protein degradation, causing paralysis and lack of cardiac muscle contraction. These phenotypes are hallmarks of myopathy and muscle atrophy. Intriguingly, Interestingly, Smyd1b translocates from the nucleus to the cytoplasm during muscle cell differentiation. Whereas it acts as a histone methyltransferase in the nucleus, Smyd1b binds to myosin and its chaperone, Hsp901, in the cytoplasm. We hypothesize that Smyd1b has dual regulatory functions. In the nucleus, Smyd1b regulates gene expression via histone methylation and interacting with other transcriptional regulators. In the cytoplasm, Smyd1b methylates muscle proteins at lysines and control protein folding, stability, and assembly into sarcomeres. To test this hypothesis, we will 1) identify Smyd1b target genes and define the molecular mechanisms by which Smyd1b functions in regulating gene expression; 2) determine the functional significance of nucleus-to-cytoplasm translocation to Smyd1b function in myofibrillogenesis; and 3) identify muscle proteins that are methylated at lysine residues by Smyd1b and determine the functional significance of lysine methylation on myofibrillogenesis. We will use the powerful zebrafish model system, which permits the use of genetic, biochemical, and proteomic approaches, to address these questions. A better understanding of the mechanisms by which Smyd1b functions in muscle cells will unravel the role of lysine methylation in muscle cell differentiation and myofibril assembly. Moreover, these studies may identify new diagnostic and therapeutic targets for treatment of muscular dystrophy and myopathy.