The major insulin-responsive tissue primarily responsible for the maintenance of normal glucose homeostasis is skeletal muscle. This tissue expresses a specific glucose transporter isoform termed GLUT4 which is substantially decreased in insulin-deficient diabetes. Further, several studies have documented that marked insulin resistance is associated with several forms of diabetes and may be the initiating event in the development of NIDDM. In addition, diabetic patients have an increased risk of coronary morbidity associated with acute myocardial infarction. Patient studies have suggested that the decrease in glucose uptake in the diabetic heart may be a key feature for lack of tissue viability during ischemic episodes. Since cardiac muscle GLUT4 mRNA and protein levels are also decreased in diabetes, the inability to increase glucose uptake during ischemia may be directly related to the decreased expression of GLUT4 mRNA. Based upon the central role of muscle GLUT4 expression in the pathophysiology associated with diabetes, we have proposed a series of studies to address the muscle-specific and hormonal/metabolic-dependent regulation of this gene. The basic molecular events involved in glucose transporter gene regulation are clearly central issues which are important for both our understanding of muscle-specific gene regulation as well as in the control of glucose homeostasis, metabolism and energy production. To accomplish these goals we plan to examine the hormonal/metabolic regulation of muscle GLUT4 transcription and to identify the cis-DNA elements responsible for the control of GLUT4 expression. In addition, we propose to identify and characterize muscle-specific DNA binding factors which mediate the normal physiological regulation of this gene. In these studies we will use nuclear run-on analysis to determine the transcription rate of the endogenous GLUT4 gene. Transient transfection assays of reporter constructs using direct injection of DNA into hindquarter muscle will be used to identify cis-DNA elements responsible for the tissue-specific and hormonal/metabolic dependent regulation of GLUT4 expression. In a complimentary approach, several reporter constructs will be integrated into transgenic mice and assayed for the normal program of GLUT4 expression. In this manner, we hope to develop an understanding of the complex regulation of muscle GLUT4 expression and a molecular basis for peripheral tissue resistance to insulin action in diabetes.
