Fatty Acid Synthase, S14, PPARa and Gene Expression
Donald B. Jump, Michelle Mater, Annette Thelen and David Pan.
Physiology Department, Michigan State University, East Lansing, MI, USA
Our studies have focused on the dietary fatty acid regulation of genes involved in hepatic fatty acid synthesis and fatty acid oxidation. Highly unsaturated long chain fatty acids, like 20:5(n-3), activate PPARa and induce the expression of genes encoding acyl CoA oxidase and cytochrome P450 4A while inhibiting the expression of several lipogenic genes. Studies with PPARa-null mice showed that while n-3 PUFA induction of AOX and CYP4A2 mRNA required PPARa, PUFA suppression of lipogenic gene expression was independent of PPARa. This suggested that PUFA controlled a second pathway affecting fatty acid synthesis. The sterol response element binding protein (SREBP1) has been implicated in the hormonal/nutrient control of lipogenic gene expression. Accordingly, we assessed the role SREBP1 plays in the PUFA control of 3 hepatic lipogenic genes: fatty acid synthase (FAS), L-pyruvate kinase (LPK) and the S14 protein (S14). Feeding rats diets supplemented with n-3 or n-6 PUFA or treatment of primary rat hepatocytes with n-3 or n-6 PUFA coordinately suppressed the mRNAs encoding SREBP1, FAS, S14 and LPK. Western blot analysis of hepatic nuclear extracts showed a decline in SREBP1 following PUFA treatment of primary hepatocytes. Like other lipogenic genes, PUFA suppression of mRNASREBP1 did not require PPARa. To assess the role SREBP1c played in lipogenic gene transcription, primary hepatocytes were co-transfected with an expression vector encoding a nuclear form of SREBP1 (nSREBP1c) along with S14CAT, FASCAT or LPKCAT. Titration studies indicate a differential sensitivity of the three promoters to nSREBP1c over expression with S14CAT and FASCAT being more sensitive than LPKCAT. Promoter deletion and block mutation studies localized multiple functional SREs within the S14 promoter. Moreover, gel shift studies showed that SREBP1c interacts with these elements. While over expression of nSREBP1c significantly induced S14CAT activity, it did not completely abrogate PUFA suppression of S14CAT activity. In conclusion, SREBP1c functionally interacts with the S14, FAS and LPK promoters to stimulate gene transcription. PUFA down regulates SREBP1 gene expression through a PPARa-independent mechanism. However, PUFA regulation of SREBP1 gene expression or its nuclear content may not account fully for the PUFA-mediated suppression of hepatic lipogenic gene expression. (Supported by NIH DK43220, USDA 98-35200-6064 and the Michigan Agricultural Experiment Station).