The functional capacity of skeletal muscle is important for the performance of daily activities for all individuals: from the demanding training regimens of elite athletes, through routine activities of daily living for the healthy individual, to those of patients with the peripheral limitations of neurological and musculoskeletal diseases, as well as to those with coronary artery disease and congestive heart failure who are limited by central hemodynamic factors. After weeks of moderate to intense exercise training, the functional capacity of an individual to perform given tasks can be remarkably improved through a transformation of skeletal muscle that is mediated by alterations in the abundance of skeletal muscle proteins essential for excitation, contraction and energy metabolism. These changes in protein abundance are the consequence of exercise-induced changes in the transcriptional activity of nuclear and mitochondrial genes, implying that gene transcription is a target for signaling pathways that link contractile activity to changes in the phenotype of skeletal muscle. The overall goal is to address the question of how the skeletal myocyte senses a need to alter its molecular makeup to meet the external demands of tonic exercise, and responds to those demands chronically by altering gene expression for whole sets of genes in a programmed, orderly, but often divergent, manner. Early work has focused on the aldolase A gene, which is activated during myocyte differentiation, but whose activity is down-regulated in response to neural signals induced during tonic exercise. The protein products of this gene is among those that determine fiber phenotype and functional properties. We propose to extend these studies to address the following specific aims: 1) To clone the promoter of the rabbit skeletal muscle- specific aldolase A gene, followed by a detailed analysis, using transfection techniques, of the structure and function of the human aldolase A promoter in response to muscle-specific signals and signals for myocyte differentiation. 2) To determine the cis-elements that mediate transcriptional repression of this gene in response to increased contractile work in skeletal muscle by using direct DNA microinjection techniques and exercise conditioning. 3) To determine candidate proteins (third messengers or transcriptional effectors) that proximally regulate the aldolase A gene in response to increased contractile activity by analysis of cDNA subtraction libraries. 4) To further define the signaling pathways involved in transducing signals from the membrane to the nucleus in response to electrical depolarization, membrane deformation, calcium release and contraction. This will involve further study of the role of the adenylyl cyclase and cAMP in mediating regulation of the aldolase A gene during the phenotypic transformation of skeletal muscles during chronic exercise conditioning. Understanding the signaling pathways mediating the process of muscular adaption to increased contractile work may lead to better approaches for the treatment of patients with cardiovascular, musculoskeletal and neurological diseases.
