A number of important health-related conditions including, but not limited to, insulin deficiency, glucocorticoid excess, and nutrient insufficiency are associated with a dramatic loss of skeletal muscle mass due largely to development of an imbalance between rates of protein synthesis and degradation, particularly in those muscles composed of a high proportion of fast-twitch fibers. In order to better understand the causes of the imbalance for each of these conditions and to begin to develop therapeutic strategies for abrogating or reversing the loss of muscle mass, the project described herein is focused specifically on that aspect of protein synthesis in which ribosomal subunits bind to messenger RNA to commence deciphering the information contained therein, i.e. the event known as translation initiation. The overall goals of the project are to define for each condition the mechanisms responsible for producing alterations in function of various translation initiation factors (specific proteins or complexes of proteins that in eukaryotes are referred to as eIF's), to assess the contribution of each alteration in modulating the rate of overall translation initiation as well as that of specific mRNAs, and to elucidate the signaling pathways involved in mediating each of the functional changes.
The specific aims of the project are as follows: 1) to define unique and shared mechanisms and signaling pathways by which physiological concentrations of either insulin or insulin-like growth factor I (IGF-I) act to stimulate translation initiation; 2) to distinguish mechanisms and signaling pathways by which limitation of single essential amino acids modulates translation initiation from those underlying the unique means of translational control imposed by leucine in the context of relative normoaminoacidemia; 3) to define mechanisms and signaling pathways by which glucocorticoids act to down-regulate translation initiation; 4) to assess the physiological role of cAMP in modulating translation initiation; and 5) to explore how coordinated translational control is attained through the integration of positive (e.g. insulin, IGF-I, and amino acids) and adverse (e.g., glucocorticoids and cAMP) stimuli. The experimental plan for achieving the specific aims will employ both in vivo and in vitro models for producing variations in hormone and amino acid availability, assess function and specific-site phosphorylation status of various rate-controlling eIF's and their regulatory proteins, employ dominant interfering variations and gene disruption to validate the roles of specific proteins, and assess the relative contributions of various protein kinase signaling pathways. Overall, the studies described in this proposal should provide new insights into the biochemical and molecular mechanisms by which protein synthesis contributes to the net gain or loss of muscle mass under a variety of physiological and pathophysiological conditions.
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