Insulin resistance is often associated with obesity and can precipitate type 2 diabetes. To date, most known approaches that improve insulin resistance must be preceded by the amelioration of obesity and hepatosteatosis. Here, we show that this provision is not mandatory; insulin resistance and hyperglycemia are improved by the modification of hepatic fatty acid composition, even in the presence of persistent obesity and hepatosteatosis. Mice deficient for Elovl6, the gene encoding the elongase that catalyzes the conversion of palmitate to stearate, were generated and shown to become obese and develop hepatosteatosis when fed a high-fat diet or mated to leptin-deficient ob/ob mice. However, they showed marked protection from hyperinsulinemia, hyperglycemia and hyperleptinemia. Amelioration of insulin resistance was associated with restoration of hepatic insulin receptor substrate-2 and suppression of hepatic protein kinase C epsilon activity resulting in restoration of Akt phosphorylation. Collectively, these data show that hepatic fatty acid composition is a new determinant for insulin sensitivity that acts independently of cellular energy balance and stress. Inhibition of this elongase could be a new therapeutic approach for ameliorating insulin resistance, diabetes and cardiovascular risks, even in the presence of a continuing state of obesity.

In an attempt to identify transcription factors which activate sterol-regulatory element-binding protein 1c (SREBP-1c) transcription, we screened an expression cDNA library from adipose tissue of SREBP-1 knockout mice using a reporter gene containing the 2.6-kb mouse SREBP-1 gene promoter. We cloned and identified the oxysterol receptors liver X receptor (LXR␣) and LXR␤ as strong activators of the mouse SREBP-1c promoter. Sterol-regulatory element (SRE)-binding proteins (SREBPs) are transcription factors which belong to the basic helix-loophelix leucine zipper family (3-5). In contrast to other members of this family, SREBPs are synthesized as precursor proteins which remain bound to the endoplasmic reticulum and the nuclear envelope in the presence of sufficient sterol concentrations. Upon sterol deprivation, the precursor protein undergoes a sequential two-step cleavage process to release the NH 2 -terminal portion (28). This mature SREBP then enters the nucleus and activates the transcription of genes involved in cholesterol and fatty acid synthesis by binding to SREs or to palindromic sequences called E boxes within their promoter regions (20, 39). Three forms of SREBP have been characterized: SREBP-1a and -1c (also known as ADD1) (14, 38, 43) and SREBP-2. It has been shown that all of the cultured cells analyzed to date express primarily SREBP-2 and the SREBP1a isoform, whereas most organs, including the liver and adipose tissue, express predominantly SREBP-2 and the SREBP1c isoform (36). SREBP-1a is a stronger activator than SREBP-1c due to its longer transactivation domain and has a wider range of target genes involved in both cholesterol and fatty acid synthesis (30, 31).Lipogenic enzymes, which are involved in energy storage through synthesis of fatty acids and triglycerides, are coordinately regulated at the transcriptional level during different metabolic states (9,11). Recent in vivo studies demonstrated that SREBP-1c plays a crucial role in the dietary regulation of most hepatic lipogenic genes, whereas SREBP-2 is actively involved in the transcription of cholesterogenic enzymes (13). These include studies of the effects of the absence or overexpression of SREBP-1 on hepatic lipogenic gene expression (30,31,33), as well as physiological changes of SREBP-1c protein in normal mice after dietary manipulation such as placement on high-carbohydrate diets, polyunsaturated fatty acid-enriched diets, and fasting-refeeding regimens (12,17,37,40,41). The similar coordinated changes in SREBP-1c and lipogenic gene expression upon fasting and refeeding were also observed in adipose tissue (18). In fat tissue, SREBP-1c (ADD1) appears to be involved in adipocyte differentiation and insulin resistance (19,35). Recent studies suggest that insulin or insulin-facilitated glucose uptake mediates lipogenesis through SREBP-1c induction (7,8,10,21,34). Previous reports on the regulation of SREBP-1c have all demonstrated the induction to be at the mRNA level. Up-regulation of hepatic SREBP-1 mRNA was observed in the livers...

Insulin receptor substrate 2 (IRS-2) is the main mediator of insulin signalling in the liver, controlling insulin sensitivity. Sterol regulatory element binding proteins (SREBPs) have been established as transcriptional regulators of lipid synthesis. Here, we show that SREBPs directly repress transcription of IRS-2 and inhibit hepatic insulin signalling. The IRS-2 promoter is activated by forkhead proteins through an insulin response element (IRE). Nuclear SREBPs effectively replace and interfere in the binding of these transactivators, resulting in inhibition of the downstream PI(3)K/Akt pathway, followed by decreased glycogen synthesis. These data suggest a molecular mechanism for the physiological switching from glycogen synthesis to lipogenesis and hepatic insulin resistance that is associated with hepatosteatosis.

Leptin-deficient ob/ob mice show many characteristics of obesity, including excess peripheral adiposity as well as severe hepatic steatosis, at least in part, due to increased hepatic lipogenesis. Polyunsaturated fatty acids (PUFAs) are not only ligands for peroxisome proliferator-activated receptor (PPAR) ␣ but are also negative regulators of hepatic lipogenesis, which is thought to be mediated by the repression of sterol regulatory element-binding protein ( S terol regulatory element-binding proteins (SREBPs) are members of the basic helix-loop-helix leucine zipper family of transcription factors that regulate fatty acid and cholesterol synthesis (reviewed in Brown and Goldstein 1 ). Unlike other members of the family, SREBPs are synthesized as precursors bound to the endoplasmic reticulum and nuclear envelope and are released from the membrane into the nucleus as mature proteins by cleavage processes. To date, 3 isoforms of SREBP, -1a, -1c, and -2, have been identified and characterized. The predominant SREBP-1 isoform in liver and adipose tissue is SREBP-1c. Whereas SREBP-2 plays a crucial role in regulation of cholesterol synthesis, SREBP-1c controls the transcription and expression of lipogenic enzymes such as fatty acid synthase (FAS) and stearoyl-CoA desaturase 1 (SCD1) (reviewed in Shimano 2 and Horton et al. 3 ). It is remarkable that SREBP-1c regulates not only the synthetic rate of triglycerides but also the amount of their storage in the liver. 4,5 Thus, SREBP-1 has been revealed to be a promising target for hepatic steatosis (fatty livers) from a therapeutic point of view.The leptin-deficient ob/ob mouse model of obesity exhibits severe obesity and obesity-related symptoms, including hepatic steatosis and insulin resistance (reviewed in Bray and York 6 ). The livers of ob/ob mice have an increase in triglyceride content, probably because of the increased lipogenesis paralleled by elevated messenger RNA (mRNA) expression and enzymatic activity of several lipogenic enzymes such as FAS and SCD1. 6,7 Recently, it has been reported that both SREBP-1c mRNA and its active nuclear protein are increased in ob/ob mouse livers. 8 Furthermore, we have demonstrated in a previous report 5 that the disruption of the SREBP-1 gene in ob/ob mice leads to marked amelioration of hepatic steatosis.Dietary polyunsaturated fatty acids (PUFAs) of the n-6 and n-3 families are well established as negative regulators of hepatic lipogenesis (reviewed in Clark and Jump 9 ). Recently, others and we have shown that the suppressive

Previous studies have demonstrated that polyunsaturated fatty acids (PUFAs) suppress sterol regulatory element-binding protein 1c (SREBP-1c) expression and, thus, lipogenesis. In the current study, the molecular mechanism for this suppressive effect was investigated with luciferase reporter gene assays using the SREBP-1c promoter in HEK293 cells. Consistent with previous data, the addition of PUFAs to the medium in the assays robustly inhibited the SREBP-1c promoter activity. Deletion and mutation of the two liver X receptor (LXR)-responsive elements (LXREs) in the SREBP-1c promoter region eliminated this suppressive effect, indicating that both LXREs are important PUFAsuppressive elements. The luciferase activities of both SREBP-1c promoter and LXRE enhancer constructs induced by co-expression of LXR␣ or -␤ were strongly suppressed by the addition of various PUFAs (arachidonic acid > eicosapentaenoic acid > docosahexaenoic acid > linoleic acid), whereas saturated or monounsaturated fatty acids had minimal effects. Gel shift mobility and ligand binding domain activation assays demonstrated that PUFA suppression of SREBP-1c expression is mediated through its competition with LXR ligand in the activation of the ligand binding domain of LXR, thereby inhibiting binding of LXR/retinoid X receptor heterodimer to the LXREs in the SREBP-1c promoter. These data suggest that PUFAs could be deeply involved in nutritional regulation of cellular fatty acid levels by inhibiting an LXR-SREBP-1c system crucial for lipogenesis. Sterol regulatory element (SRE)1 -binding proteins (SREBPs) are membrane-bound transcription factors that belong to the basic helix-loop-helix leucine zipper family (1-3). In the absence of sterols, by means of sterol-regulated cleavage, SREBP enters the nucleus and activates the transcription of genes involved in cholesterol and fatty acid synthesis by binding to an SRE or its related sequences including SRE-like sequences and E-boxes, within their promoter regions (4, 5). There are three forms of SREBP, SREBP-1a and -1c (also known as ADD1) and -2 (6 -8). Most organs, including the liver and adipose tissue, predominantly express SREBP-2 and the -1c isoform of SREBP-1 (9). Recent in vivo studies demonstrate that SREBP-1c plays a crucial role in the dietary regulation of most hepatic lipogenic genes, whereas SREBP-2 is actively involved in the transcription of cholesterogenic enzymes (10). These include studies of the effects of the absence or overexpression of SREBP-1 on hepatic lipogenic gene expression (10 -12) as well as physiological changes of SREBP-1c protein in normal mice refed after fasting (13-17). Polyunsaturated fatty acid (PUFA) administration has been well established as a negative regulator of hepatic lipogenesis as well as an activator of peroxisome proliferator-activated receptor (PPAR) ␣, which is crucial for lipid degradation. Consistent with the notion that SREBP-1c is a dominant regulator for lipogenesis, there are several reports demonstrating that administration of PUFA suppresses ...