REGULATION OF GLUCOSE-6-PHOSPHATASE GENE EXPRESSION

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Type II, non-insulin dependent diabetes mellitus (NIDDM) is characterized by defects in insulin secretion, peripheral glucose utilization (PGU) and hepatic glucose production (HGP). The ability of insulin to stimulate PGU and repress HGP in patients with NIDDM is reduced as a consequence of insulin resistance. In addition, in Type I, insulin-dependent diabetes mellitus (IDDM), HGP can increase if circulating insulin levels are low or because poor glycemic control has led to the development of insulin resistance. Persistent hyperglycemia is the cause of many of the complications associated with diabetes. In both IDDM and NIDDM this increased HGP is a consequence of an increased rate of gluconeo-genesis. The final step of the gluconeogenic pathway is catalyzed by glucose-6-phosphatase (G6Pase). Recent data has shown that overexpression of the G6Pase catalytic subunit results in an increased rate of HGP. Thus, the suppression of G6Pase catalytic subunit gene expression may represent a potential strategy for reducing HGP in diabetic patients. The rational development of a pharmaceutical agent that suppresses G6Pase catalytic subunit gene expression will require a detailed knowledge of the cis-acting elements and trans-acting factors through which expression of the gene is regulated. Specific Aims 1 and 2 of this grant application propose to characterize the cis-acting elements and trans-acting factors that mediate the stimulatory effect of cAMP and the inhibitory effect of phorbol esters on G6Pase catalytic subunit gene transcription, respectively. This will be achieved using a fusion gene strategy in conjunction with the transfection of tissue culture cell lines as well as the generation of transgenic mice. From preliminary studies it is apparent that multiple cis-acting elements are required for the full effect of both agents. In addition, the cis-acting elements that mediate the stimulatory effect of cAMP in liver cells are distinct from those that mediate this effect in kidney cells. The inhibitory action of insulin on G6Pase catalytic subunit gene transcription requires two promoter regions designated A and B. Region A binds hepatocyte nuclear factor-1 (HNF-1) but does not directly mediate the action of insulin. Instead, HNF-1 enhances the action of insulin mediated through an unidentified transcription factor that binds Region B. The third Specific Aim of this application proposes to explore several aspects of the mechanism of insulin action on G6Pase gene transcription.