Dietary fatty acids specifically modulate the onset and progression of various diseases, including cancer, atherogenesis, hyperlipidaemia, insulin resistances and hypertension, as well as blood coagulability and fibrinolytic defects; their effects depend on their chain length and degree of saturation. Hepatocyte nuclear factor-4alpha (HNF-4alpha) is an orphan transcription factor of the superfamily of nuclear receptors and controls the expression of genes that govern the pathogenesis and course of some of these diseases. Here we show that long-chain fatty acids directly modulate the transcriptional activity of HNF-4alpha by binding as their acyl-CoA thioesters to the ligand-binding domain of HNF-4alpha. This binding may shift the oligomeric-dimeric equilibrium of HNF-4alpha or may modulate the affinity of HNF-4alpha for its cognate promoter element, resulting in either activation or inhibition of HNF-4alpha transcriptional activity as a function of chain length and the degree of saturation of the fatty acyl-CoA ligands. In addition to their roles as substrates to yield energy, as an energy store, or as constituents of membrane phospholipids, dietary fatty acids may affect the course of a disease by modulating the expression of HNF-4alpha-controlled genes.

The hypolipidemic effect exerted by beta,beta'-tetramethyl-hexadecanedioic acid (Medica 16) is accounted for by enhanced catabolism of plasma triglyceride-rich lipoproteins due to a decrease in plasma apolipoprotein C-III (Frenkel, B., Mayorek, N., Hertz, R., and Bar-Tana, J. (1988) J. Biol. Chem. 263, 8491-8497; Frenkel, B., Bishara-Shieban, J., and Bar-Tana, J. (1994) Biochem. J. 298, 409-414). Decrease in apolipoprotein C-III exerted by peroxisome proliferators/hypolipidemic amphipathic carboxylates (e.g. Medica 16, fibrate drugs) is shown here to result from suppression of apolipoprotein C-III gene expression. Transcriptional suppression of apolipoprotein C-III is due to transcriptional suppression of hepatic nuclear factor (HNF)-4 as well as displacement of HNF-4 from the apolipoprotein C-III promoter. HNF-4 displacement exerted by peroxisome proliferators/hypolipidemic amphipathic carboxylates is mediated by the peroxisome proliferators activated receptor (PPAR). Transcriptional suppression of HNF-4-enhanced genes (e.g. apolipoprotein C-III) along with transcriptional activation of peroxisomal and other genes by hypolipidemic drugs may account for their broad spectrum pharmacological effect.

Hepatic nuclear factor-4␣ (HNF-4␣ Hepatic nuclear factor 4 (HNF-4)1 is a member of the superfamily of nuclear receptors which includes steroid hormone receptors and nonsteroid ligand dependent transcription factors, e.g. thyroid hormone receptor, retinoid X receptor, retinoid acid receptor, and peroxisome proliferator-activated receptors (PPARs) (reviewed in Ref. 1). HNF-4␣ isoforms (␣ 1 -␣ 3 ) have been cloned and characterized and are expressed in mammals in liver, kidney, intestine, and pancreas (reviewed in Ref.2). Unlike retinoid X receptor ␣, with which it has 40% amino acid sequence homology, HNF-4 does not form heterodimers with any other nuclear receptor, but binds to direct repeat-1 DNA sequences as homodimer (3). Direct repeat-1 motifs are promiscuous binding sites for HNF-4, PPAR, retinoid acid receptor, retinoid X receptor, chicken ovalbumin upstream-promoter transcription factor, and chicken ovalbumin upstream-promoter transcription factor homo-or heterodimers (4).HNF-4␣ responsive genes encode transcription factors (HNF-1␣, PXR), proteins involved in fatty acid, lipoprotein, and lipid metabolism (apoA-I, apoA-II, apoB, apoC-II, apoC-III, L p (a), microsomal triglyceride transfer protein, mitochondrial fatty acyl-CoA dehydrogenases, and fatty acid-binding protein), carbohydrate metabolism (insulin, glut2, glucose-6-phosphatase, phosphoenolpyruvate carboxykinase, pyruvate kinase, aldolase B, and glyceraldehyde-3-phosphate dehydrogenase), amino acid and protein metabolism (ornithine transcarbamylase, tyrosine aminotransferase, phenylalanine hydroxylase, and antitrypsin ␣ 1 ), P450 enzymes (steroid 15␣-hydroxylase, fatty acyl -hydroxylase, cholesterol-7␣-hydroxylase, and drug metabolizing P450 enzymes (cyp3␣ 4 -6)), hematopoiesis (erythropoietin and transferrin), blood coagulation (factors VII, IX, and X and fibrinogen), and others (e.g. cellular retinolbinding protein and transthyretin) (reviewed in Refs. 2 and 5-14). Since HNF-4␣ activates the transcription of some nuclear receptors (HNF-1␣) and may further directly interact with other transcription factors (HNF-1␣), the above list of HNF-4␣ responsive genes may include some which are transcriptionally affected by HNF-4␣ indirectly.The first demonstration of putative HNF-4 ligands showed that long chain fatty acyl-CoA (LCFA-CoAs) thioesters (15), as well as CoA-thioesters of hypolipidemic peroxisome prolifera-

Acyl-CoA-binding protein (ACBP) 1 is a ubiquitous intracellular lipid-binding protein whose physiological function remains to be determined (reviewed in Refs. 1-3). Until recently, ACBP was thought to be primarily cytosolic and unique among the soluble intracellular lipid proteins in exhibiting very high affinity (K d values of 0.5-10 nM) and exclusive specificity for long chain fatty acyl-CoAs (LCFA-CoAs) (1, 4 -7). Based on the fact that ACBP exclusively binds LCFA-CoAs, it may participate in several aspects of LCFA-CoA metabolism.First, ACBP may be involved in directly presenting LCFACoAs as substrates for lipid metabolic enzymes. A variety of studies in vitro suggest that ACBP extracts LCFA-CoAs from membranes (6) to increase the soluble LCFA-CoA pool available for intracellular transport (reviewed in Ref.2). The cytosolic ACBP⅐LCFA-CoA complexes then interact with and present LCFA-CoA to acyltransferase enzymes involved in phospholipid synthesis in the endoplasmic reticulum (8, 9), lysophosphatidic acid synthesis in mitochondria (10), cholesteryl ester synthesis in the endoplasmic reticulum (11), and oxidation in mitochondria (10).Second, ACBP may control the level of unbound LCFA-CoA available for interaction with regulatory sites on metabolic enzymes (e.g. acetyl-CoA carboxylase) and intracellular signaling proteins (e.g. protein kinase C) in the cytosol (reviewed in Refs. 12 and 13).Third, recent data demonstrating the presence of significant amounts of ACBP in the nuclei of transfected cells overexpressing ACBP suggest that ACBP may also be involved in direct or ligand (LCFA-CoA)-dependent regulation of nuclear proteins that activate transcription of genes involved in lipid and glucose metabolism (14,15). Several members of the nuclear receptor superfamily including the hepatocyte nuclear receptor 4␣ (HNF-4␣) (16 -18), thyroid hormone receptor (TR) (19), and peroxisome proliferator-activated receptor-␣ and -␦ (PPAR-␣ and -␦) (20, 21) interact with LCFA-CoAs. The relative order of affinities of these nuclear receptors for LCFA-CoAs is HNF-4␣ (K d of 1.5-4 nM) Ͼ Ͼ TR (K d of 120 nM) Ͼ Ͼ PPAR␣ (displaces Wy14643). On this basis, it appears that only HNF-4␣ binds LCFA-CoAs with affinities in the physiological range of LCFACoA levels in the nucleus, Ͻ10 nM (17, 18). HNF-4␣ is a nuclear receptor with major roles in hepatocyte differentiation during liver development and in regulating the transcription of nu-

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