DIFFERENCES IN BINDING OF HEPATIC NUCLEAR PROTEINS FROM LEAN AND OBESE RATS TO THE 5'-UPSTREAM REGION OF TYROSINE AMINOTRANSFERASE

The glucocorticoid effects on liver tyrosine aminotransferase mRNA lev els have been studied in young, lean, and obese Zucker (fa/fa) rats an d 5'-upstream regions of the tyrosine aminotransferase (TAT) gene have been used in gel retardation studies to investigate nuclear protein b inding, Hepatic TAT mRNA levels were increased in obese fa/fa rats but were normalized seven days after adrenalectomy. Corticosterone replac ement to adrenalectomized rats restored the increased levels of TAT mR NA in the obese animals. A 60-bp fragment of upstream TAT DNA (-2463 t o -2403) was identified which showed higher levels of band shifting af ter incubation with hepatic nuclear proteins of obese rats compared wi th the proteins from lean animals. This differential level of gel reta rdation was substantially reduced by alkaline phosphatase treatment of nuclear proteins. Gel retardation was reduced when nuclear proteins m ere prepared from adrenalectomized obese rats, and increased with nucl ear proteins from adrenalectomized rats replaced with corticosterone. DNA affinity chromatography and gel electrophoresis identified three p roteins of approximately 58, 62, and 65 kDa in the DNA-protein complex . Increased amounts of these three proteins were purified from nuclei of obese rats. HNF3(alpha) antibodies induced hypershift of the gel re tardation pattern implicating HNF3, as one of the proteins that binds to the 60 bp DNA fragment, The data support the hypothesis that decrea sed phosphorylation of nuclear proteins in obese rats is glucocorticoi d-dependent and may contribute to the altered transcriptional activity of glucocorticoid-responsive genes.