NUTRITIONAL AND HORMONAL REGULATION OF HEPATIC GENES

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Mammalian organisms regulate the expression of their genetic information in response to nutritional and metabolic signals; however, little is currently known regarding the molecular mechanisms involved in these processes. The long range goal of this work is to understand one system of metabolic regulation. This system involves the induction of enzymes catalyzing lipogenesis in the liver in response to increased carbohydrate metabolism. We have focused on two genes - the L-type pyruvate kinase (PK) and S14 genes - that are transcriptionally activated by increased glucose metabolism in the hepatocyte. Past work has defined specific regulatory regions found in the 5'-flanking region of these genes that are necessary and sufficient for control. These regions share a common arrangement including two elements: a carbohydrate response element (ChoRE) that by itself is capable of responding to increased glucose metabolism and an accessory factor site that enhances the response. The ChoRE contains a unique arrangement of two E box motifs (CACGTG) that are spaced by 5 base pairs. We hypothesize that this element serves as the binding site for a carbohydrate-responsive factor or complex. Understanding the nature of this transcriptional factor is the first objective of this work. This will be addressed by: (1) determining whether the factor USF, which binds to the sequence CACGTG, is a component of the carbohydrate-responsive complex; (2) isolating and cloning the factor binding to the ChoRE from rat liver; and (3) identifying other components of the carbohydrate-responsive complex involved in gene regulation using a protein interaction screen. With this information, we will pursue studies to explore the mechanism of activation of the factor in hepatocytes undergoing elevated glycolysis. In particular, we will explore the role of phosphorylation in this process. Finally, we will characterize the nature of a novel accessory factor that binds to both the PK and S14 genes and determine whether this factor is involved in receiving signals from other physiological regulators of these genes. These studies should help in deciphering the signaling process by which the hepatocyte can sense increased glycolysis and respond by changing its transcriptional program and serve as a model for transcriptional regulation by nutrients in higher eucaryotes.