Carbohydrate Regulation of Hepatic Gene Expression

doi:10.1074/jbc.272.11.7525

Carbohydrate Regulation of Hepatic Gene Expression

EVIDENCE AGAINST A ROLE FOR THE UPSTREAM STIMULATORY FACTOR*

From the Department of Biochemistry and Institute of Human Genetics, Medical School, University of Minnesota, Minneapolis, Minnesota 55455

^¶ To whom correspondence should be addressed:
Dept. of Biochemistry, 4-225 Millard Hall, 435 Delaware St. S.E., Minneapolis, MN 55455
. Tel.: 612-625-3662; Fax: 612-625-2163.

↵^§ Present address: Graduate Institute of Medical Technology, College of Medicine, National Taiwan University, Taipei 10016, Taiwan, Republic of China.

Abstract

Hepatic expression of the genes encoding L-type pyruvate kinase (L-PK) and S₁₄ is induced in rats upon feeding them a high carbohydrate, low fat diet. A carbohydrate response element (ChoRE) containing two CACGTG-type E boxes has been mapped in the 5′-flanking region of both of these genes. The nature of the ChoRE suggests that a member of the basic/helix-loop-helix/leucine zipper family of proteins may be responsible for mediating the response to carbohydrate. Indeed, the upstream stimulatory factor (USF), a ubiquitous basic/helix-loop-helix/leucine zipper protein, is present in hepatic nuclear extracts and binds to the ChoREs of L-PK and S₁₄ in vitro We have conducted experiments to determine whether USF is involved in the carbohydrate-mediated regulation of L-PK and S₁₄. For this purpose, dominant negative forms of USF that are capable of heterodimerizing with endogenous USF but not of binding to DNA were expressed in primary hepatocytes. Expression of these forms did not block either S₁₄ or L-PK induction by glucose. In addition, we have constructed mutant ChoREs that retain their carbohydrate responsiveness but have lost the ability to bind USF. Together, these data suggest that USF is not the carbohydrate-responsive factor that stimulates S₁₄ and L-PK expression and that a distinct hepatic factor is likely to be responsible for the transcriptional response.

Footnotes

↵^‡ Supported in part by NIH Training Grant 5T32-GM07323.
↵* This work was supported in part by National Institutes of Health (NIH) Grant DK26919. The costs of publication of this article were defrayed in part by the payment of page charges. The article must therefore be hereby marked “advertisement” in accordance with 18 U.S.C. Section 1734 solely to indicate this fact.
↵¹ The abbreviations used are:

L-PK

liver-type pyruvate kinase

ChoRE

carbohydrate response element

b/HLH/LZ

basic/helix-loop-helix/leucine zipper

USF

upstream stimulatory factor

EMSA

electrophoretic mobility shift assay

CAT

chloramphenicol acetyltransferase

PCR

polymerase chain reaction

SREBP

sterol response element-binding protein.
↵² H.-M. Shih and H. C. Towle, unpublished results.
↵³ E. N. Kaytor and H. C. Towle, unpublished results.
- Received September 25, 1996.
- Revision received December 3, 1996.