Doctoral Dissertations
Date of Award
12-1994
Degree Type
Dissertation
Degree Name
Doctor of Philosophy
Major
Biochemistry and Cellular and Molecular Biology
Major Professor
John W. Koontz
Committee Members
David Brian, Elizabeth Howell, Wesley Wicks
Abstract
Insulin inhibits glucocorticoid-induced expression of the liver-specific gene tyrosine aminotransferase (TAT). In order to identify sequences that mediate this insulin effect, KRC-7 rat hepatoma cells were transiently transfected with constructs containing mutant TAT 5'-flanking sequences. Results of these studies demonstrated that the glucocorticoid responsive unit at -2.5 kb and the TATA region of TAT gene were sufficient to confer at least part of the insulin inhibitory effect on glucocorticoid response. However, the TAT glucocorticoid responsive unit alone was incapable of mediating any insulin response in the context of a heterologous promoter. Thus, insulin appears to inhibit glucocorticoid-induced TAT expression by interfering with the interaction between the glucocorticoid responsive unit-bound factors and the TAT transcriptional apparatus. The interaction between glucocorticoids and insulin in regulating gene expression was also studied using a series of chimeric genes constructed to contain synthetic glucocorticoid responsive element(s) 5'- of the herpes simplex virus thymidine kinase promoter and the chloramphenicol acetyltransferase reporter gene. The magnitude of induction by glucocorticoids was dependent on the number of glucocorticoid responsive elements. Insulin alone had little effect on expression of these genes but together with dexamethasone acted in a synergistic manner. This synergy diminished as the number of glucocorticoid responsive elements in the promoter increases. The synergy is independent of promoter sequences other than the glucocorticoid responsive elements and a functional TATA box. Three different approaches demonstrate that the effect of insulin is not directly on the glucocorticoid signal transduction pathway. Insulin does not change the dose-response relationship for dexamethasone. The effect of insulin is independent of the intracellular concentration of glucocorticoid receptor. The effect is independent of any specific domain of the glucocorticoid receptor. The target of insulin action is likely to be part of the normal host cell transcriptional apparatus or an adaptor molecule.
Recommended Citation
Pan, Li, "Regulation of glucocorticoid-induced gene expression by insulin. " PhD diss., University of Tennessee, 1994.
https://trace.tennessee.edu/utk_graddiss/10560