Doctoral Dissertations

Date of Award

8-1989

Degree Type

Dissertation

Degree Name

Doctor of Philosophy

Major Professor

John W. Koontz

Committee Members

Jorge Churchich, Frank Kenney, W. D. Wicks

Abstract

Tyrosine aminotransferase (TAT) is a liver-specific enzyme subject to complex multi-hormonal regulation. TAT has been shown to be regulated at the transcriptional, post-transcriptional, and post-translational levels. In adult liver and in a variety of both human and rat hepatomas, TAT activity is induced by cAMP, glucocorticoids, and insulin. In most cases the increase in enzyme activity due to hormonal induction is preceded by an increase in the amount of hybridizable TAT mRNA. In adult rat liver and in some hepatomas this increase in mRNA has been shown to be a result of an increase in the transcriptional rate of the TAT gene. In all hepatic tissue( with the exception of certain fetal stages) tested, glucocorticoids and cAMP stimulate the induction of TAT. However, insulin regulation of TAT in rat hepatomas, and in the developing fetus is much more complex. In primary hepatocytes and in FT0-2B rat hepatomas, insulin alone has no effect of the transcription of the TAT gene, although in the FT0-2B cells insulin can inhibit the levels of transcription previously induced by cAMP. Insulin also appears to play an important role in developmental regulation of TAT. Insulin has been implicated in inhibiting the production of TAT in the developing fetus. However, at birth , when insulin levels drop, TAT is readily detectable and after several hours is expressed at the adult level. We have characterized a subclone (KRC-7) of the Reuber H-35 rat hepatoma with respect to hormonal regulation of TAT. In these cells insulin induces TAT activity while paradoxically inhibiting the rate of TAT transcription. In addition, insulin can dramatically inhibit the cAMP- or glucocorticoid-induced transcriptional rate. To further characterize this repression of TAT by insulin, we have analyzed the 5'-flanking region of the rat TAT gene for possible sequence elements which confer insulin responsiveness in the KRC-7 cells. Through the utilization of a transient transfectional analysis we have localized regions flanking the TAT gene which appear to be responsible for the insulin inhibition.

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