Functional antagonism between inhibitor of DNA binding (Id) and adipocyte determination and differentiation factor 1/sterol regulatory element-binding protein-1c (ADD1/SREBP-1c) trans-factors for the regulation of fatty acidsynthase promoter in adipocytes

We show that Id (inhibitor of DNA binding) 2 and Id3, dominant negative mem bers of the helix-loop-helix (HLH) family, interact with the adipocyte dete rmination and differentiation factor 1 (ADD1)/sterol regulatory element-bin ding protein (SREBP) 1c, a transcription factor of the basic HLH-leucine zi pper family that controls the expression of several key genes of adipose me tabolism. Gel mobility-shift assays performed with in vitvo-translated ADD1 , Id2 or Id3 proteins and a fatty acid synthase (FAS) promoter oligonucleot ide showed evidence for a marked inhibition of the formation of DNA-ADD1 co mplexes by Id2 or Id3 proteins. Co-immunoprecipitation studies using in vit ro-translated proteins demonstrated further the physical interaction of Id and ADD1/SREBP-1c proteins in the absence of DNA. Using the FAS gene as a m odel of an ADD1-regulated promoter in transiently transfected isolated rat adipocytes or mature 3T3-L1 adipocytes, a potent inhibition of the activity of the FAS-chloramphenicol acetyltransferase reporter gene was observed by overexpression of Id2 or Id3. Reciprocally, co-transfection of Id3 antisen se and ADD1 expression vectors in preadipocytes potentiated the ADD1/SREBP- 1c effect on the FAS promoter activity. Finally, in the non adipogenic NIH- 3T3 cell line, most of the ADD1-mediated trans-activation of the FAS promot er was counteracted by co-transfection of Id2 or Id3 expression vectors. Pr evious studies have indicated Id gene expression to be down-regulated durin g adipogenesis [Moldes, Lasnier, Five, Pairault and Djian (1997) Mol. Cell. Biol. 17, 1796-1804]. We here demonstrated that there was a dramatic rise of Id2 and Id3 mRNA levels when 3T3-L1 adipocytes or isolated rat fat cells were exposed to lipolytic and anti-lipogenic agents, forskolin and isoprot erenol. Taken together, our data show that Id products are functionally inv olved in modulating ADD1/SREBP-1c transcriptional activity, and thus lipoge nesis in adipocytes.