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Molecular and Cellular Biology, August 2003, p. 5896-5907, Vol. 23, No. 16
0270-7306/03/$08.00+0     DOI: 10.1128/MCB.23.16.5896-5907.2003
Copyright © 2003, American Society for Microbiology. All Rights Reserved.

Occupancy and Function of the -150 Sterol Regulatory Element and -65 E-Box in Nutritional Regulation of the Fatty Acid Synthase Gene in Living Animals

Maria-Jesus Latasa,1 Michael J. Griffin,1 Yang Soo Moon,1,{dagger} Chulho Kang,2 and Hei Sook Sul1*

Department of Nutritional Sciences and Toxicology,1 Department of Molecular and Cellular Biology, University of California, Berkeley, California 947202

Received 23 December 2002/ Returned for modification 3 March 2003/ Accepted 20 May 2003

Upstream regulatory factor (USF) and sterol regulatory element binding protein (SREBP) play key roles in the transcriptional regulation of the fatty acid synthase (FAS) gene by feeding and insulin. Due to the dual binding specificity of SREBP, as well as the presence of multiple consensus sites for these transcription factors in the FAS promoter, their physiologically relevant functional binding sites have been controversial. Here, in order to determine the occupancy of the putative USF and SREBP binding sites, we examined their protein-DNA interactions in living animals by using formaldehyde cross-linking and immunoprecipitation of chromatin and tested the function of these elements by employing mice transgenic for a reporter gene driven by various 5' deletions as well as site-specific mutations of the FAS promoter. We show that the -332 and -65 E-boxes are bound by USF in both fasted and refed mice, while the -150 SRE is bound by SREBP-1 only in refed mice. We also found that mutation of either the -150 SRE or the -65 E-box abolishes the feeding-induced activation of the FAS promoter in transgenic mice. Furthermore, in vivo occupancy of the FAS promoter by SREBP in the fed state can be prevented by mutation not only of the -150 SRE but, unexpectedly, of the -65 E-box as well. We conclude that the FAS promoter is activated during refeeding via the induced binding of SREBP to the -150 SRE and that USF binding to the -65 E-box is also required for SREBP binding and activation of the FAS promoter.

* Corresponding author. Mailing address: Department of Nutritional Sciences and Toxicology, University of California, Berkeley, CA 94720. Phone: (510) 642-3978. Fax: (510) 642-0535. E-mail: hsul{at}nature.berkeley.edu.

{dagger} Present address: Dept. of Animal Sciences & Biotechnology, Jinju National University, Jinju 660-758, Korea.

Molecular and Cellular Biology, August 2003, p. 5896-5907, Vol. 23, No. 16
0022-538X/03/$08.00+0     DOI: 10.1128/MCB.23.16.5896-5907.2003
Copyright © 2003, American Society for Microbiology. All Rights Reserved.



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