Daniela Botolin

Regulation of hepatic fatty acid elongase and desaturase expression in diabetes and obesity

Fatty acid elongases and desaturases play an important role in hepatic and whole body lipid composition. We examined the role that key transcription factors played in the control of hepatic elongase and desaturase expression. Studies with peroxisome proliferator-activated receptor α (PPARα)-deficient mice establish that PPARα was required for WY14643-mediated induction of fatty acid elongase-5 (Elovl-5), Elovl-6, and all three desaturases [Δ5 desaturase (Δ5D), Δ6D, and Δ9D]. Increased nuclear sterol-regulatory element binding protein-1 (SREBP-1) correlated with enhanced expression of Elovl-6, Δ5D, Δ6D, and Δ9D. Only Δ9D was also regulated independently by liver X receptor (LXR) agonist. Glucose induction of l-type pyruvate kinase, Δ9D, and Elovl-6 expression required the carbohydrate-regulatory element binding protein/MAX-like factor X (ChREBP/MLX) heterodimer. Suppression of Elovl-6 and Δ9D expression in livers of streptozotocin-induced diabetic rats and high fat-fed glucose-intolerant mice correlated with low levels of nuclear SREBP-1. In leptin-deficient obese mice (Lepob/ob), increased SREBP-1 and MLX nuclear content correlated with the induction of Elovl-5, Elovl-6, and Δ9D expression and the massive accumulation of monounsaturated fatty acids (18:1,n-7 and 18:1,n-9) in neutral lipids. Diabetes- and obesity-induced changes in hepatic lipid composition correlated with changes in elongase and desaturase expression. In conclusion, these studies establish a role for PPARα, LXR, SREBP-1, ChREBP, and MLX in the control of hepatic fatty acid elongase and desaturase expression and lipid composition.

The Role of Liver X Receptor-α in the Fatty Acid Regulation of Hepatic Gene Expression

Liver X receptors (LXR) α and β play an important role in regulating the expression of genes involved in hepatic bile and fatty acid synthesis, glucose metabolism, as well as sterol efflux. Studies with human embryonic kidney 293 cells indicate that unsaturated fatty acids interfere with oxysterols binding to LXR and antagonize oxysterol-induced LXRα activity. In this report, we evaluated the effects of unsaturated fatty acids on LXR-regulated hepatic gene expression. The LXR agonist, T1317, induced mRNAs encoding sterol regulatory element-binding protein 1c (SREBP-1c) and two SREBP-1c-regulated lipogenic genes, e.g. fatty-acid synthase and the S14 protein in primary hepatocytes. Treatment of hepatocytes with eicosapentaenoic acid (20:5n-3) suppressed these mRNAs in the absence and presence of T1317. The cis-regulatory elements targeted by T1317 were not required for fatty-acid suppression of FAS or S14 promoter activity. In contrast to SREBP-1-regulated lipogenic genes, 20:5n-3 had no effect on the T1317 induction of ABCG5 or ABCG8 in the rat hepatoma cell line, FTO-2B. These two genes require LXR but not SREBP-1c for their expression. Feeding rats a diet supplemented with fish oil suppressed hepatic SREBP-1c-regulated genes and induced PPARα-regulated genes but had no effect on the LXR-regulated transcripts, CYP7A1, ABCG5, or ABCG8. Transfection studies, using either full-length hLXRα or a chimera containing only the LXRα ligand binding domain, indicate that a wide array of unsaturated fatty acids had little effect on LXRα activity in primary hepatocytes or FTO-2B. These studies suggest that LXRα is not a target for unsaturated fatty acid regulation in primary rat hepatocytes or in liver. Thus, oxysterol/LXR-mediated regulation of transcripts involved in bile acid synthesis or sterol efflux appear insensitive to dietary unsaturated fatty acids. The unsaturated fatty acid suppression of SREBP-1 and its targeted lipogenic genes is independent of LXRα

Docosahexaenoic acid (DHA) and hepatic gene transcription.

The type and quantity of dietary fat ingested contributes to the onset and progression of chronic diseases, like diabetes and atherosclerosis. The liver plays a central role in whole body lipid metabolism and responds rapidly to changes in dietary fat composition. Polyunsaturated fatty acids (PUFA) play a key role in membrane composition and function, metabolism and the control of gene expression. Certain PUFA, like the n-3 PUFA, enhance hepatic fatty acid oxidation and inhibit fatty acid synthesis and VLDL secretion, in part, by regulating gene expression. Our studies have established that key transcription factors, like PPARalpha, SREBP-1, ChREBP and MLX, are regulated by n-3 PUFA, which in turn control levels of proteins involved in lipid and carbohydrate metabolism. Of the n-3 PUFA, 22:6,n-3 has recently been established as a key controller of hepatic lipid synthesis. 22:6,n-3 controls the 26S proteasomal degradation of the nuclear form of SREBP-1. SREBP-1 is a major transcription factor that controls the expression of multiple genes involved fatty acid synthesis and desaturation. 22:6,n-3 suppresses nuclear SREBP-1, which in turn suppresses lipogenesis. This mechanism is achieved, in part, through control of the phosphorylation status of protein kinases. This review will examine both the general features of PUFA-regulated hepatic gene transcription and highlight the unique mechanisms by which 22:6,n-3 impacts gene expression. The outcome of this analysis will reveal that changes in hepatic 22:6,n-3 content has a major impact on hepatic lipid and carbohydrate metabolism. Moreover, the mechanisms involve 22:6,n-3 control of several well-known signaling pathways, such as Akt, Erk1/2, Gsk3beta and PKC (novel or atypical). 22:6,n-3 control of these same signaling pathways in non-hepatic tissues may help to explain the diverse actions of n-3 PUFA on such complex physiological processes as visual acuity and learning.

Docosahexaneoic acid (22:6,n-3) regulates rat hepatocyte SREBP-1 nuclear abundance by Erk- and 26S proteasome-dependent pathways

Insulin induces and dietary n-3 PUFAs suppress hepatic de novo lipogenesis by controlling sterol-regulatory element binding protein-1 nuclear abundance (nSREBP-1). Our goal was to define the mechanisms involved in this regulatory process. Insulin treatment of rat primary hepatocytes rapidly augments nSREBP-1 and mRNASREBP-1c while suppressing mRNAInsig-2 but not mRNAInsig-1. These events are preceded by rapid but transient increases in Akt and Erk phosphorylation. Removal of insulin from hepatocytes leads to a rapid decline in nSREBP-1 [half-time (T1/2) ∼ 10 h] that is abrogated by inhibitors of 26S proteasomal degradation. 22:6,n-3, the major n-3 PUFA accumulating in livers of fish oil-fed rats, suppresses hepatocyte levels of nSREBP-1, mRNASREBP-1c, and mRNAInsig-2 but modestly and transiently induces mRNAInsig-1. More importantly, 22:6,n-3 accelerates the disappearance of hepatocyte nSREBP-1 (T1/2 ∼ 4 h) through a 26S proteasome-dependent process. 22:6,n-3 has minimal effects on microsomal SREBP-1 and sterol-regulatory element binding protein cleavage-activating protein or nuclear SREBP-2. 22:6,n-3 transiently inhibits insulin-induced Akt phosphorylation but induces Erk phosphorylation. Inhibitors of Erk phosphorylation, but not overexpressed constitutively active Akt, rapidly attenuate 22:6,n-3 suppression of nSREBP-1. Thus, 22:6,n-3 suppresses hepatocyte nSREBP-1 through 26S proteasome- and Erk-dependent pathways. These studies reveal a novel mechanism for n-3 PUFA regulation of hepatocyte nSREBP-1 and lipid metabolism.

Selective proteolytic processing of rat hepatic sterol regulatory element binding protein-1 (SREBP-1) and SREBP-2 during postnatal development.

Sterol regulatory element-binding protein-1c (SREBP-1c) plays a major role in hepatic lipogenic gene expression. In adult animals, insulin and oxysterols induce SREBP-1c gene transcription, whereas polyunsaturated fatty acids suppress the nuclear content of SREBP-1c through pre-translational regulatory mechanisms. A decline in nuclear SREBP-1 is associated with suppression of hepatic lipogenesis. In contrast to adult rats, hepatic lipogenesis in preweaned neonatal rats is low. Ingestion of milk fat by the neonate may contribute to low hepatic lipogenesis. In this report, we tested the hypothesis that low lipogenic gene expression prior to weaning correlates with low mRNASREBP-1c, as well as low precursor and nuclear forms of SREBP-1. In contrast to expectations, levels of mRNASREBP-1c and the 125-kDa SREBP-1 precursor in livers of preweaned rats was comparable with adult levels. Despite high levels of SREBP-1 precursor, mature (65 kDa) SREBP-1 was not detected in rat liver nuclei prior to 18 days postpartum. Weaning rats at 21 days postpartum was accompanied by a rise in nuclear SREBP-1 levels as well as increased lipogenic gene expression. In contrast, SREBP-2 was present in rat liver nuclei, and its target gene,HMG-CoA reductase, was expressed above adult levels prior to weaning. These studies indicate that, prior to weaning, SREBP-2 but not SREBP-1 is proteolytically processed to the mature form. As such, SREBP-2-regulated genes are active. Failure of SREBP-1 to be processed to the mature form <18 days postpartum correlates with low hepatic lipogenic gene expression. This mechanism differs from the hormonal and fatty acid-mediated pre-translational control of SREBP-1c in adult liver.

Fatty acids and gene transcription

The type and quantity of dietary fat ingested contributes to the onset and progression of chronic diseases, such as diabetes and atherosclerosis. The liver plays a central role in whole-body lipid metabolism and responds rapidly to changes in dietary fat composition. In rodents, n-3 polyunsaturated fatty acids (PUFAs) enhance hepatic fatty acid oxidation and inhibit fatty acid synthesis and very low-density lipoprotein secretion, in part, by regulating key transcription factors, including peroxisome proliferator activated receptor-? (PPAR-?), sterol regulatory element binding protein-1 (SREBP-1), carbohydrate regulatory element binding protein (ChREBP) and Max-like factor X (MLX). These transcription factors control the expression of multiple genes involved in lipid synthesis and oxidation. Changes in PPAR-? target genes correlate well with changes in intracellular non-esterified fatty acids. Insulin stimulates hepatic de novo lipogenesis by rapidly inducing SR EBP-1 nuclear abundance (nSREBP-1). This mechanism is linked to insulin-induced protein kinase B (Akt) and glycogen synthase kinase (Gsk)-3? phosphorylation and inhibition of 26S proteasomal degradation of nSREBP-1. n-3 PUFAs, particularly 22:6 n-3, inhibit lipid synthesis by suppressing nSREBP-1. A major action of 22:6 n-3 is to stimulate the loss of nSREBP-1 through 26S proteasomal and extracellular regulated kinase (Erk)-dependent pathways. 22:6 n-3 is the only n-3 PUFA accumulating in livers of rodents or humans ingesting essential fatty acid-sufficient or n-3 PUFA-enriched diets. As such, 22:6 n-3 is a major feedback regulator of hepatic lipid synthesis. Finally, insulin-stimulated glucose metabolism augments de novo lipogenesis by elevating nuclear levels of ChREBP, a key regulator of glycolytic and lipogenic genes. ChREBP binding to promoters requires MLX. n-3 PUFAs repress expression of the glycolytic gene, L-pyruvate kinase and lipogenic genes by suppressing MLX nu c lear abundance. In summary, n-3 PUFAs control the activity or abundance of several hepatic transcription factors that impact hepatic carbohydrate and lipid metabolism. Recent studies have identified Erk, Gsk-3? and MLX as novel targets of fatty acid-regulated gene expression. Keywords: gene transcription; hepatic fatty acid metabolism