Date of Graduation

1984

Document Type

Thesis

Abstract

The objective of my research project was to examine how insulin and the glucocorticoids, alone, and in combination, regulate hepatic glucose-6-phosphate dehydrogenase (G6PD) at the level of functional mRNA coding for G6PD. The model system I utilized for these studies was primary cultures of adult rat liver parenchymal cells. These cells were maintained in a serum-free and chemically defined medium so that direct effects of added hormones on the hepatocytes could be ascertained. Dexamethasone (Dex), a synthetic glucocorticoid, when added alone did not have an effect on G6PD activity. Insulin caused about a 1.6-fold increase in enzyme activity, and when Dex and insulin were present together, Dex acted in a "permissive" manner to amplify the insulin stimulation of G6PD (2.5-fold increase). These effects on enzyme activity were paralleled by increases in the relative rate of G6PD synthesis. Upon measurement of the levels of functional mRNA coding for G6PD it was observed that Dex by itself stimulated G6PD mRNA levels 4-fold while insulin treatment resulted in a 2-fold increase. When Dex and insulin were added together G6PD mRNA levels were increased in an additive fashion (an approximate 7-fold stimulation). The results of my studies suggest that glucocorticoid exerts its regulation on G6PD at the pretranslational level by modulating levels of mRNA coding for G6PD, however, this mRNA is not expressed unless insulin has also been added. Thus insulin would appear to regulate G6PD at the pretranslational level (as evidenced by insulin treatment alone) and possibly at the translational or posttranscriptional level (as evidenced by combined Dex and insulin treatment). More definitive conclusions as to the exact nature of the pretranslational regulation being exerted by Dex and insulin, i.e., whether this represents authentic regulation of gene transcription as opposed to regulation of posttranscriptional events such as mRNA processing, transport of mRNA to the cytoplasm, or stabilization of cytoplasmic mRNA must await the availability of a cDNA probe specific for rat liver glucose-6-phosphate dehydrogenase.

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