5′ Sequences Direct Developmental Expression and Hormone Responsiveness of Tyrosine Aminotransferase in Primary Cultures of Fetal Rat Hepatocytes

Abstract

Tyrosine aminotransferase (TyrAT) is one of several gluconeogenic enzymes which appear postnatally in humans and rodents in response to increased glucocorticoid and glucagon levels and decreased insulin. Primary cultured fetal rat hepatocytes older than day 15 of gestation (>E15) transcribe the TyrAT gene in response to the synergistic effect of dexamethasone and Nh,2'-O-dibutyryl-adenosine 3',5'-monophosphate (Bt,cAMP), whereas less mature hepatocytes ( Biochem. 199,. Therefore, we consider >El5 hepatocytes, and not <El5 hepatocytes, to be determined. This study reports that 11.1 kb of sequences upstream of the TyrAT transcription start site, which include a CAMP-responsive element (CRE) and a glucocorticoid-responsive element (GRE), are required for correct developmental regulation of gene expression in determined fetal hepatocytes. In contrast, the TyrAT CRE alone does not have this capability. Dexamethasone augments basal and Bt,cAMP-stimulated activity of the TyrAT CRE alone, suggesting that synergism may be due to interaction between the glucocorticoid and CAMP-signaling pathways. However, Bt,cAMP does not further increase dexamethasone-induced activity of the 11.1 kb 5' sequences when the TyrAT CRE is removed, thus excluding interaction of Bt,cAMP with the glucocorticoid pathway. Finally, insulin inhibition of dexamethasone-induced gene transcription is shown to be conferred by TyrAT 5' sequences. This study shows that cellular components, other than those which mediate hormonal regulation of genes, are required for determination of hepatocytes with respect to TyrAT. Since this phenomenon is observed with transient transfections, it is unlikely to involve higher-order chromatin structure.Keywords : tyrosine aminotransferase ; hepatocyte ; fetal development ; hormone ; gene regulationThe appearance of tyrosine arninotransferase (TyrAT) in the liver of human and rodent neonates coincides with cessation of the maternal glucose supply and the subsequent de novo synthesis of glucose by the new-born. Glucagon and glucocorticoid fluxes and decreased levels of circulating insulin in the newborn are associated with these metabolic changes.Fetal liver is capable of synthesising TyrAT under conditions which mimic the nutritional or hormonal environment at birth. In rats, the enzyme can be induced by premature delivery [ l ] or by in utero injection of glucagon or Nh,2'-O-dibutyryl-adenosine 3',5'-monophosphate (Bt,cAMP) [2]. Glucocorticoid analogues and glucagon act synergistically in fetal liver to induce TyrAT catalytic activity and steady-state mRNA levels [3]. Thus, glucagon and glucocorticoids play a fundamental role in the appearance of active TyrAT enzyme. Studies in adult rat liver demon-