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. 2001 Oct 15;359(Pt 2):273–284. doi: 10.1042/0264-6021:3590273

Promoter I of the ovine acetyl-CoA carboxylase-alpha gene: an E-box motif at -114 in the proximal promoter binds upstream stimulatory factor (USF)-1 and USF-2 and acts as an insulin-response sequence in differentiating adipocytes.

M T Travers 1, A J Vallance 1, H T Gourlay 1, C A Gill 1, I Klein 1, C B Bottema 1, M C Barber 1
PMCID: PMC1222145  PMID: 11583573

Abstract

Acetyl-CoA carboxylase-alpha (ACC-alpha) plays a central role in co-ordinating de novo fatty acid synthesis in animal tissues. We have characterized the regulatory region of the ovine ACC-alpha gene. Three promoters, PI, PII and PIII, are dispersed throughout 50 kb of genomic DNA. Expression from PI is limited to adipose tissue and liver. Sequence comparison of the proximal promoters of ovine and mouse PIs demonstrates high nucleotide identity and that they are characterized by a TATA box at -29, C/EBP (CCAAT enhancer-binding protein)-binding motifs and multiple E-box motifs. A 4.3 kb ovine PI-luciferase reporter construct is insulin-responsive when transfected into differentiated ovine adipocytes, whereas when this construct is transfected into ovine preadipocytes and HepG2 cells the construct is inactive and is not inducible by insulin. By contrast, transfection of a construct corresponding to 132 bp of the proximal promoter linked to a luciferase reporter is active and inducible by insulin in all three cell systems. Insulin signalling to the -132 bp construct in differentiated ovine adipocytes involves, in part, an E-box motif at -114. Upstream stimulatory factor (USF)-1 and USF-2, but not sterol regulatory element-binding protein 1 (SREBP-1), are major components of protein complexes that bind this E-box motif. Activation of the 4.3 kb PI construct in differentiated ovine adipocytes is associated with endogenous expression of PI transcripts throughout differentiation; PI transcripts are not detectable by RNase-protection assay in ovine preadipocytes, HepG2 cells or 3T3-F442A adipocytes. These data indicate the presence of repressor motifs in PI that are required to be de-repressed during adipocyte differentiation to allow induction of the promoter by insulin.

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