Search

Stearoyl-CoA desaturase (SCD)1 converts saturated fatty acids into monounsaturated fatty acids. Using muscle overexpression, we sought to determine the role of SCD1 expression in glucose and lipid metabolism and its effects on exercise capacity in mice. Wild-type C57Bl/6 (WT) and SCD1 muscle transgenic (SCD1-Tg) mice were generated, and expression of the SCD1 transgene was restricted to skeletal muscle. SCD1 overexpression was associated with increased triglyceride (TG) content. The fatty acid composition of the muscle revealed a significant increase in polyunsaturated fatty acid (PUFA) content of TG, including linoleate (18:2n6). Untrained SCD1-Tg mice also displayed significantly increased treadmill exercise capacity (WT = 6.6 ± 3 min, Tg = 71.9 ± 9.5 min; P = 0.0009). SCD1-Tg mice had decreased fasting plasma glucose, glucose transporter (GLUT)1 mRNA, fatty acid oxidation, mitochondrial content, and increased peroxisome proliferator-activated receptor (PPAR)δ and Pgc-1 protein expression in skeletal muscle. In vitro studies in C2C12 myocytes revealed that linoleate (18:2n6) and not oleate (18:1n9) caused a 3-fold increase in PPARδ and a 9-fold increase in CPT-1b with a subsequent increase in fat oxidation. The present model suggests that increasing delta-9 desaturase activity of muscle increases metabolic function, exercise capacity, and lipid oxidation likely through increased PUFA content, which increases PPARδ expression and activity. However, the mechanism of action that results in increased PUFA content of SCD1-Tg mice remains to be elucidated.

Stearoyl-CoA desaturase 1 (SCD1) deficiency partitions fatty acids away from lipid synthesis towards fatty acid oxidation in liver and skeletal muscle in part due to activation of AMP-activated protein kinase (AMPK) pathway. The mechanism of AMPK activation by SCD1 mutation is unknown, however since SCD1-/- animals have increased relative amounts of polyunsaturated fatty acids (PUFA), we hypothesized that the increased levels of PUFA might be responsible for the activation of AMPK in SCD1 deficient mice. Therefore, the present study was undertaken to analyze the effect of PUFA on AMPK in liver, skeletal muscle, and heart. We fed mice ad libitum for 14 days with diet supplemented with fish oil (5% fat). As expected, fish oil supplementation significantly increased n-3 PUFA content in each of the analyzed tissues. Hepatic mRNA levels of fatty acid synthase and acyl-CoA oxidase decreased by 92% and increased by 60%, respectively, consistent with known PUFA effects. However, after 14 days of PUFA feeding, we did not find any changes in AMPK phosphorylation and protein content in mouse liver, skeletal muscle, and heart. The data suggest that PUFA are not involved in AMPK activation in mouse tissues and that the increased activity of AMPK in SCD1-/- mice is probably PUFA-independent.

Apart from being an important macronutrient, dietary fat has recently gained much prominence for its role in regulating gene expression. Polyunsaturated fatty acids (PUFAs) affect gene expression through various mechanisms including, but not limited to, changes in membrane composition, intracellular calcium levels, and eicosanoid production. Furthermore, PUFAs and their various metabolites can act at the level of the nucleus, in conjunction with nuclear receptors and transcription factors, to affect the transcription of a variety of genes. Several of these transcription mediators have been identified and include the nuclear receptors peroxisome proliferator-activated receptor (PPAR), hepatocyte nuclear factor (HNF)-4alpha, and liver X receptor (LXR) and the transcription factors sterol-regulatory element binding protein (SREBP) and nuclear factor-kappaB (NFkappaB). Their interaction with PUFAs has been shown to be critical to the regulation of several key genes of lipid metabolism. Working out the mechanisms by which these interactions and consequent effects occur is proving to be complicated but is invaluable to our understanding of the role that dietary fat can play in disease management and prevention.

Polyunsaturated fatty acids (PUFAs), specifically the n-3 series, have been implicated in the prevention of various human diseases, including obesity, diabetes, coronary heart disease and stroke, and inflammatory and neurologic diseases. PUFAs function mainly by altering membrane lipid composition, cellular metabolism, signal transduction, and regulation of gene expression. PUFAs regulate the expression of genes in various tissues, including the liver, heart, adipose tissue, and brain. The role of transcription factors such as SREBP1c and nuclear receptors such as PPAR-alpha, HNF-4alpha, and LXRalpha in mediating the nuclear effects of PUFAs are addressed.

Stearoyl-CoA desaturase (SCD) catalyzes the rate-limiting step in the cellular synthesis of monounsaturated fatty acids, mainly oleate (18:1) and palmitoleate (16:1), which are the major monounsaturated fatty acids of membrane phospholipids, cholesteryl esters, waxes, and triglycerides. The mouse expresses three well-characterized SCD genes (SCD1, 2, and 3). SCD1 is the main isoform expressed in the liver of mice. Previous in vivo studies have shown that the transcriptional repression by n-3 and n-6 polyunsaturated fatty acids (PUFAs) and the induction by cholesterol of the SCD1 gene are dependent on the maturation of the sterol regulatory element-binding protein-1c (SREBP-1c). We studied the regulation of SREBP-1, SCD1, and other SREBP-1 target genes when a high cholesterol diet is combined with PUFA as n-6 PUFA rich soybean oil (SO), or n-3 PUFA rich fish oil (FO). While the PUFA/cholesterol (PUFA/CH) diets repressed the maturation of the SREBP-1, the SCD1 mRNA levels, and protein and enzyme activity were induced. Compared with PUFA diets, hepatic cholesterol ester and triglyceride were enriched with 16:1 and 18:1 monounsaturated fatty acids in mice fed PUFA/CH diets. Total plasma cholesterol levels were not altered but plasma triglycerides were reduced in SO/CH-fed mice compared with SO-fed mice. The mRNA for SREBP-1 was increased by the PUFA/CH diet but the mRNA levels of SREBP-1 target genes such as fatty acid synthase and LDL receptor were decreased, indicating that the main control of PUFA-mediated suppression of SREBP-1 target genes is the maturation of SREBP-1. This study demonstrates that cholesterol overrides the PUFA-mediated repression of the SCD1 gene and regulates SCD1 gene expression through a mechanism independent of SREBP-1 maturation.

Cellular cholesterol and fatty acid metabolism in mammals is controlled by a family of transcription factors called sterol regulatory element-binding protein isoforms, three of which (SREBP-1a, 1c, and 2) are well characterized. These proteins, which are synthesized as precursors, are inserted into the endoplasmic reticulum (ER) membrane with both the amino and carboxylic acid domains facing the cytosolic face of the membrane. In sterol-deficient cells, proteolytic cleavage of SREBPs occurs, thereby releasing their N-terminal mature and active forms and enabling them to enter the nucleus, where they bind to the sterol regulatory response element (SRE) and/or E-box sequences and activate genes involved in cholesterol, triglyceride, and fatty acid biosynthesis. Of the three SREBP isoforms, SREBP-1c gene expression is induced by cholesterol and repressed by polyunsaturated fatty acids (PUFA). We have examined the changes in SREBP-1c mRNA and protein levels as well as the mRNA levels of several SREBP-1c target genes when a high-cholesterol diet is combined with diets rich in PUFA of the n-6 series. Our studies show that PUFA oppose the cholesterol-mediated SREBP-1 maturation without affecting the cholesterol-mediated increase of SREBP-1c mRNA and precursor protein. The decrease in SREBP-1 mature protein paralleled the decrease in mRNAs for genes of fatty acid and cholesterol biosynthesis, such as HMG-CoA synthase and fatty acid synthase, but interestingly gene expression of stearoyl-CoA desaturase 1 (SCD1) was instead induced. These studies suggest that the main point of control of PUFA-mediated suppression of lipogenic gene expression is the inhibition of SREBP-1 maturation. The studies also reveal that the induction of SCD1 gene expression by cholesterol occurs through a mechanism independent of SREBP-1 maturation.

Stearoyl-CoA desaturase (SCD) is a microsomal enzyme required for the biosynthesis of oleate (C18:1) and palmitoleate (C16:1) which are the major monounsaturated fatty acids of membrane phospholipids, triglycerides and cholesterol esters. Previously the full-length human skin cDNA was sequenced and the exon and intron structure of the single functional SCD gene determined. Here we report on the cloning and characterization of the promoter region of the human SCD gene. The human promoter structure is very similar to that of the mouse SCD1 isoform and contains conserved regulatory sequences for the binding of several transcription factors including the sterol regulatory element binding protein (SREBP), CCAAT enhancer binding protein-alpha (C/EBPalpha) and nuclear factor-1 (NF-1) that have been shown to transactivate the transcription of the mouse SCD1 gene. Polyunsaturated fatty acids and cholesterol decreased the SCD promoter-luciferase activity when transiently transfected into HepG2 cells. The decrease in promoter activity correlated with decreases in endogenous SCD mRNA and protein levels. Cotransfection experiment in HepG2 cells showed transactivation of the SCD promoter-luciferase activity by an expression vector containing SREBP-1a and 1c. Our studies indicate that the transcription of the human SCD gene is repressed by polyunsaturated fatty acids and cholesterol and that SREBP plays a role in the transcriptional activation of this gene., (Copyright 2001 Academic Press.)

Polyunsaturated fatty acids (PUFAs), specifically the n-3 and n-6 series, play a key role in the progression or prevention of human diseases such as obesity, diabetes, cancer, neurological and heart disease, mainly by affecting cellular membrane lipid composition, metabolism, signal-transduction pathways, and by direct control of gene expression. PUFAs show regulation of gene expression in several tissues, including brain, liver, heart, and adipose. Most recently, research has focused on identifying the mechanisms by which PUFAs regulate lipogenic gene expression. Research to date indicates that PUFA-mediated regulation of the genetic expression and proteolytic maturation of a group of transcription factors termed sterol regulatory element binding proteins (SREBPs) accounts for the suppression of hepatic lipogenic gene expression. However, our recent studies on the transcriptional regulation of the stearoyl-coenzyme A (CoA) desaturase gene, encoding a key enzyme in the cellular synthesis of monounsaturated fatty acids from saturated fatty acids indicates that PUFA can suppress gene transcription by a mechanism independent of SREBP maturation.

For the past three decades, polyunsaturated fatty acids (PUFA) have been recognized as important energy sources and membrane components. PUFA also play key roles in many cellular events, such as gene regulation. Most recently, research has focused on identifying the mechanisms by which PUFA modulate gene transcription, mRNA stability and cellular differentiation. It is the purpose of this review to examine the effects of PUFA on gene expression in lipogenic as well as other tissues. Because the (n-3) and (n-6) series of PUFA are intimately involved in gene regulation, they will be the focus of review. The effects of other fatty acid families on gene expression are reviewed elsewhere.

Polyunsaturated fatty acids (PUFA) repress stearoyl-CoA desaturase gene 1 (SCD1) expression in liver and adipose tissues. We used HepG2 cells to localize genetic regulatory elements for PUFA in the SCD1 5'-flanking region. A chimeric reporter gene construct containing the 4.3 kb SCD1 putative promoter was transiently transfected into HepG2 cells, which were then treated with various fatty acids. We observed greater than 60% repression of transcription with 18:3n - 3 and 75% repression with 20:4n - 6 and 20:5n - 3. No significant change was seen with 18:0. Using smaller SCD1 chimeric constructs, we localized the genetic regulatory region to a 237 bp sequence within the SCD1 proximal promoter. DNA mobility shift analysis with HepG2 and mouse liver nuclear extracts demonstrated specific binding of nuclear proteins to this region. Mobility shift analysis with nuclear extract from 3T3-L1 adipocytes showed a similar pattern of protein binding. Competitive DNA mobility shift analysis identified a 60 bp region containing sites that specifically bind and compete for nuclear proteins. This region conferred responsiveness to PUFA when placed in a heterologous promoter. A homologous region in the stearoyl-CoA desaturase gene 2 (SCD2) promoter also mediated PUFA-specific repression in transfection experiments. These data suggest that a common transcriptional mechanism may exist in liver and adipose tissues for inhibition of lipogenesis by PUFA.

The effects of arachidonic acid (20:4, n-6) and other fatty acids on the expression of stearoyl-CoA desaturase gene 1 were investigated in fully differentiated 3T3-L1 adipocytes. Treatment of 3T3-L1 adipocytes with arachidonic acid resulted in a decrease in stearoyl-CoA desaturase (Scd) enzyme activity and scd1 mRNA. Arachidonic acid did not alter the transcription of the scd1 gene, whereas the half-life of the scd1 mRNA was reduced from 25.1 to 8.5 h. Blocking the conversion of arachidonic acid to eicosanoids by pretreatment of the cells with cyclooxygenase, lipoxygenase, or cytochrome P-450 epoxygenase inhibitors did not reverse the inhibition caused by arachidonic acid, indicating that eicosanoid synthesis is not necessary for the repression of scd1 mRNA expression. Treatment of adipocytes with linoleic (18:2, n-6) and linolenic (18:3, n-3) acids also resulted in inhibition of scd1 mRNA accumulation. By contrast, oleic acid (18:1, n-9) and stearic acid (18:0) had no effect on scd1 mRNA levels. Taken together, these results suggest that polyunsaturated fatty acids repress the expression of the scd1 gene in mature adipocytes by reducing the stability of scd1 mRNA.

Insulin and polyunsaturated fatty acids (PUFAs) regulate the expression of SCD1 gene in mouse liver. Accordingly, we examined the insulin and PUFA regulation of SCD1 gene expression in H2.35 cells. The levels of SCD1 mRNA in H2.35 cells increased at the restrictive temperature of 39 degrees C, when the glucose-containing medium was supplemented with insulin. The insulin-stimulated expression of SCD1 mRNA was significantly blunted when the induction medium was supplemented with linolenic acid (18:2n-3) and arachidonic acid (20:4n-6). Stearic acid (18:0n-9) and oleic acid (18:1n-9) were without dramatic effects. The effect of insulin and PUFAs on a transfected SCD1 fusion gene (SCD1.CAT4.3) was also examined in H2.35 cells. Whereas insulin stimulated SCD1.CAT4.3 expression, arachidonic acid significantly decreased SCD1.CAT4.3 activity. These studies suggest that insulin and PUFAs regulate SCD1 gene transcription via regulatory DNA sequences flanking the 5' end of the gene.