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.

Agency
National Institute of Health (NIH)
Institute
National Institute of Diabetes and Digestive and Kidney Diseases (NIDDK)
Type
Research Project (R01)
Project #
5R01DK026919-20
Application #
6124772
Study Section
Endocrinology Study Section (END)
Program Officer
Laughlin, Maren R
Project Start
1980-04-01
Project End
2000-11-30
Budget Start
1999-12-01
Budget End
2000-11-30
Support Year
20
Fiscal Year
2000
Total Cost
$209,569
Indirect Cost
Name
University of Minnesota Twin Cities
Department
Biochemistry
Type
Schools of Medicine
DUNS #
168559177
City
Minneapolis
State
MN
Country
United States
Zip Code
55455
Aipoalani, Derrick L; O'Callaghan, Brennon L; Mashek, Douglas G et al. (2010) Overlapping roles of the glucose-responsive genes, S14 and S14R, in hepatic lipogenesis. Endocrinology 151:2071-7
Davies, Michael N; O'Callaghan, Brennon L; Towle, Howard C (2010) Activation and repression of glucose-stimulated ChREBP requires the concerted action of multiple domains within the MondoA conserved region. Am J Physiol Endocrinol Metab 299:E665-74
Davies, Michael N; O'Callaghan, Brennon L; Towle, Howard C (2008) Glucose activates ChREBP by increasing its rate of nuclear entry and relieving repression of its transcriptional activity. J Biol Chem 283:24029-38
Tsatsos, Nikolas G; Augustin, Lance B; Anderson, Grant W et al. (2008) Hepatic expression of the SPOT 14 (S14) paralog S14-related (Mid1 interacting protein) is regulated by dietary carbohydrate. Endocrinology 149:5155-61
Tsatsos, Nikolas G; Davies, Michael N; O'Callaghan, Brennon L et al. (2008) Identification and function of phosphorylation in the glucose-regulated transcription factor ChREBP. Biochem J 411:261-70
Ma, Lin; Sham, Yuk Y; Walters, Kylie J et al. (2007) A critical role for the loop region of the basic helix-loop-helix/leucine zipper protein Mlx in DNA binding and glucose-regulated transcription. Nucleic Acids Res 35:35-44
Tsatsos, Nikolas G; Towle, Howard C (2006) Glucose activation of ChREBP in hepatocytes occurs via a two-step mechanism. Biochem Biophys Res Commun 340:449-56
Ma, Lin; Tsatsos, Nikolas G; Towle, Howard C (2005) Direct role of ChREBP.Mlx in regulating hepatic glucose-responsive genes. J Biol Chem 280:12019-27
Towle, Howard C (2005) Glucose as a regulator of eukaryotic gene transcription. Trends Endocrinol Metab 16:489-94
Stoeckman, Angela K; Ma, Lin; Towle, Howard C (2004) Mlx is the functional heteromeric partner of the carbohydrate response element-binding protein in glucose regulation of lipogenic enzyme genes. J Biol Chem 279:15662-9

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