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1.  Limited Effect of Dietary Saturated Fat on Plasma Saturated Fat in the Context of a Low Carbohydrate Diet 
Lipids  2010;45(10):947-962.
We recently showed that a hypocaloric carbohydrate restricted diet (CRD) had two striking effects: (1) a reduction in plasma saturated fatty acids (SFA) despite higher intake than a low fat diet, and (2) a decrease in inflammation despite a significant increase in arachidonic acid (ARA). Here we extend these findings in 8 weight stable men who were fed two 6-week CRD (12%en carbohydrate) varying in quality of fat. One CRD emphasized SFA (CRD-SFA, 86 g/d SFA) and the other, unsaturated fat (CRD-UFA, 47 g SFA/d). All foods were provided to subjects. Both CRD decreased serum triacylglycerol (TAG) and insulin, and increased LDL-C particle size. The CRD-UFA significantly decreased plasma TAG SFA (27.48 ± 2.89 mol%) compared to baseline (31.06 ± 4.26 mol%). Plasma TAG SFA, however, remained unchanged in the CRD-SFA (33.14 ± 3.49 mol%) despite a doubling in SFA intake. Both CRD significantly reduced plasma palmitoleic acid (16:1n-7) indicating decreased de novo lipogenesis. CRD-SFA significantly increased plasma phospholipid ARA content, while CRD-UFA significantly increased EPA and DHA. Urine 8-iso PGF2α, a free radical-catalyzed product of ARA, was significantly lower than baseline following CRD-UFA (−32%). There was a significant inverse correlation between changes in urine 8-iso PGF2α and PL ARA on both CRD (r = −0.82 CRD-SFA; r = −0.62 CRD-UFA). These findings are consistent with the concept that dietary saturated fat is efficiently metabolized in the presence of low carbohydrate, and that a CRD results in better preservation of plasma ARA.
PMCID: PMC2974193  PMID: 20820932
Saturated fat; Palmitic acid; Palmitoleic acid; Plasma fatty acid composition; Ketogenic diet; Omega-3 eggs; Metabolic syndrome; Insulin sensitivity; Controlled human feeding study; EPA; DHA; LDL/HDL ratio
2.  Genetic parameters for milk fatty acids in Danish Holstein cattle based on SNP markers using a Bayesian approach 
BMC Genetics  2013;14:79.
For several years, in human nutrition there has been a focus on the proportion of unsaturated fatty acids (UFA) and saturated fatty acids (SFA) found in bovine milk. The positive health-related properties of UFA versus SFA have increased the demand for food products with a higher proportion of UFA. To be able to change the UFA and SFA content of the milk by breeding it is important to know whether there is a genetic component underlying the individual FA in the milk. We have estimated the heritability for individual FA in the milk of Danish Holstein. For this purpose we used information of SNP markers instead of the traditional pedigree relationships.
Estimates of heritability were moderate within the range of 0.10 for C18:1 trans-11 to 0.34 for C8:0 and C10:0, whereas the estimates for saturated fatty acids and unsaturated fatty acids were 0.14 and 0.18, respectively. Posterior standard deviations were in the range from 0.07 to 0.17. The correlation estimates showed a general pattern of two groups, one group mainly consisting of saturated fatty acids and one group mainly consisting of unsaturated fatty acids. The phenotypic correlation ranged from −0.95 (saturated fatty acids and unsaturated fatty acids) to 0.99 (unsaturated fatty acids and monounsaturated fatty acids) and the genomic correlation for fatty acids ranged from −0.29 to 0.91.
The heritability estimates obtained in this study are in general accordance with heritability estimates from studies using pedigree data and/or a genomic relationship matrix in the context of a REML approach. SFA and UFA expressed a strong negative phenotypic correlation and a weaker genetic correlation. This is in accordance with the theory that SFA is synthesized de novo, while UFA can be regulated independently from the regulation of SFA by the feeding regime.
PMCID: PMC3854798  PMID: 24024882
Genomic heritability; Genomic correlation; Bayesian mixed model; Milk fatty acids
3.  Distinct metabolic effects following short-term exposure of different high-fat diets in male and female mice 
Endocrine journal  2014;61(5):457-470.
Obesity-associated hepatic lipid accumulation and chronic low-grade inflammation lead to metabolic defects. Saturated fatty acids (SFA) are a risk factor for, whereas unsaturated fatty acids (UFA) are thought to be protective against, developing metabolic diseases. Sex differences exist in the regulation of metabolism. We tested the hypothesis that diets high in SFA, mono-UFA (MUFA), or poly-UFA (PUFA) had early, sex-distinct effects that differentially contribute to long-term metabolic disturbance such as fatty liver and insulin resistance. Metabolic changes including body and fat mass, circulating leptin and glucose levels, plasma lipid profile, hepatic lipid accumulation, expression levels of genes related to lipid metabolism and low-grade inflammation, and tissue insulin sensitivity were compared between male and female mice fed with a low-fat chow, or high-fat SFA, MUFA, or PUFA for a short period of four days. SFA and MUFA males increased adiposity associated with increased liver lipid accumulation and rapid activation of inflammation in adipose and muscle tissues, whereas PUFA males did not show lipid accumulation or tissue inflammation compared to chow males. All SFA and UFA males displayed tissue insulin resistance. In contrast, female high-fat diet groups had normal liver lipid content and maintained tissue insulin sensitivity without showing tissue inflammation. Therefore, sex differences existed during early phase of development of metabolic dysfunction. The beneficial effects of PUFA, but not MUFA, were corroborated in protection of obesity, hyperlipidemia, fatty liver, and low-grade inflammation. The benefit of MUFA and PUFA in maintaining tissue insulin sensitivity in males, however, was questioned.
PMCID: PMC4045093  PMID: 24646677
sex difference; de novo lipogenesis; β-oxidation; insulin sensitivity; low-grade inflammation
4.  Isolation and Characterization of Unsaturated Fatty Acid Auxotrophs of Streptococcus pneumoniae and Streptococcus mutans▿  
Journal of Bacteriology  2007;189(22):8139-8144.
Unsaturated fatty acid (UFA) biosynthesis is essential for the maintenance of membrane structure and function in many groups of anaerobic bacteria. Like Escherichia coli, the human pathogen Streptococcus pneumoniae produces straight-chain saturated fatty acids (SFA) and monounsaturated fatty acids. In E. coli UFA synthesis requires the action of two gene products, the essential isomerase/dehydratase encoded by fabA and an elongation condensing enzyme encoded by fabB. S. pneumoniae lacks both genes and instead employs a single enzyme with only an isomerase function encoded by the fabM gene. In this paper we report the construction and characterization of an S. pneumoniae 708 fabM mutant. This mutant failed to grow in complex medium, and the defect was overcome by addition of UFAs to the growth medium. S. pneumoniae fabM mutants did not produce detectable levels of monounsaturated fatty acids as determined by gas chromatography-mass spectrometry and thin-layer chromatography analysis of the radiolabeled phospholipids. We also demonstrate that a fabM null mutant of the cariogenic organism Streptococcus mutants is a UFA auxotroph, indicating that FabM is the only enzyme involved in the control of membrane fluidity in streptococci. Finally we report that the fabN gene of Enterococcus faecalis, coding for a dehydratase/isomerase, complements the growth of S. pneumoniae fabM mutants. Taken together, these results suggest that FabM is a potential target for chemotherapeutic agents against streptococci and that S. pneumoniae UFA auxotrophs could help identify novel genes encoding enzymes involved in UFA biosynthesis.
PMCID: PMC2168684  PMID: 17827283
5.  Membrane Synthesis, Specific Lipid Requirements, and Localized Lipid Composition Changes Associated with a Positive-Strand RNA Virus RNA Replication Protein 
Journal of Virology  2003;77(23):12819-12828.
Multifunctional RNA replication protein 1a of brome mosaic virus (BMV), a positive-strand RNA virus, localizes to the cytoplasmic face of endoplasmic reticulum (ER) membranes and induces ER lumenal spherules in which viral RNA synthesis occurs. We previously showed that BMV RNA replication in yeast is severely inhibited prior to negative-strand RNA synthesis by a single-amino-acid substitution in the ole1w allele of yeast Δ9 fatty acid (FA) desaturase, which converts saturated FAs (SFAs) to unsaturated FAs (UFAs). Here we further define the relationships between 1a, membrane lipid composition, and RNA synthesis. We show that 1a expression increases total membrane lipids in wild-type (wt) yeast by 25 to 33%, consistent with recent results indicating that the numerous 1a-induced spherules are enveloped by invaginations of the outer ER membrane. 1a did not alter total membrane lipid composition in wt or ole1w yeast, but the ole1w mutation selectively depleted 18-carbon, monounsaturated (18:1) FA chains and increased 16:0 SFA chains, reducing the UFA-to-SFA ratio from ∼2.5 to ∼1.5. Thus, ole1w inhibition of RNA replication was correlated with decreased levels of UFA, membrane fluidity, and plasticity. The ole1w mutation did not alter 1a-induced membrane synthesis, 1a localization to the perinuclear ER, or colocalization of BMV 2a polymerase, nor did it block spherule formation. Moreover, BMV RNA replication templates were still recovered from cell lysates in a 1a-induced, 1a- and membrane-associated, and nuclease-resistant but detergent-susceptible state consistent with spherules. However, unlike nearby ER membranes, the membranes surrounding spherules in ole1w cells were not distinctively stained with osmium tetroxide, which interacts specifically with UFA double bonds. Thus, in ole1w cells, spherule-associated membranes were locally depleted in UFAs. This localized UFA depletion helps to explain why BMV RNA replication is more sensitive than cell growth to reduced UFA levels. The results imply that 1a preferentially interacts with one or more types of membrane lipids.
PMCID: PMC262592  PMID: 14610203
6.  Effects on Coronary Heart Disease of Increasing Polyunsaturated Fat in Place of Saturated Fat: A Systematic Review and Meta-Analysis of Randomized Controlled Trials 
PLoS Medicine  2010;7(3):e1000252.
Dariush Mozaffarian and colleagues conduct a systematic review and meta-analysis to investigate the effect of consuming polyunsaturated fats in place of saturated fats for lowering the risk of coronary heart disease.
Reduced saturated fat (SFA) consumption is recommended to reduce coronary heart disease (CHD), but there is an absence of strong supporting evidence from randomized controlled trials (RCTs) of clinical CHD events and few guidelines focus on any specific replacement nutrient. Additionally, some public health groups recommend lowering or limiting polyunsaturated fat (PUFA) consumption, a major potential replacement for SFA.
Methods and Findings
We systematically investigated and quantified the effects of increased PUFA consumption, as a replacement for SFA, on CHD endpoints in RCTs. RCTs were identified by systematic searches of multiple online databases through June 2009, grey literature sources, hand-searching related articles and citations, and direct contacts with experts to identify potentially unpublished trials. Studies were included if they randomized participants to increased PUFA for at least 1 year without major concomitant interventions, had an appropriate control group, and reported incidence of CHD (myocardial infarction and/or cardiac death). Inclusions/exclusions were adjudicated and data were extracted independently and in duplicate by two investigators and included population characteristics, control and intervention diets, follow-up duration, types of events, risk ratios, and SEs. Pooled effects were calculated using inverse-variance-weighted random effects meta-analysis. From 346 identified abstracts, eight trials met inclusion criteria, totaling 13,614 participants with 1,042 CHD events. Average weighted PUFA consumption was 14.9% energy (range 8.0%–20.7%) in intervention groups versus 5.0% energy (range 4.0%–6.4%) in controls. The overall pooled risk reduction was 19% (RR = 0.81, 95% confidence interval [CI] 0.70–0.95, p = 0.008), corresponding to 10% reduced CHD risk (RR = 0.90, 95% CI = 0.83–0.97) for each 5% energy of increased PUFA, without evidence for statistical heterogeneity (Q-statistic p = 0.13; I2 = 37%). Meta-regression identified study duration as an independent determinant of risk reduction (p = 0.017), with studies of longer duration showing greater benefits.
These findings provide evidence that consuming PUFA in place of SFA reduces CHD events in RCTs. This suggests that rather than trying to lower PUFA consumption, a shift toward greater population PUFA consumption in place of SFA would significantly reduce rates of CHD.
Please see later in the article for the Editors' Summary
Editors' Summary
Coronary heart disease (CHD) is the leading cause of death among adults in developed countries. It is caused by disease of the coronary arteries, the blood vessels that supply the heart with oxygen and nutrients. With age, inflammatory deposits (atherosclerotic plaques) coat the walls of these arteries and restrict the heart's blood supply, causing angina (chest pains that are usually relieved by rest), shortness of breath, and, if these plaques rupture or break, heart attacks (myocardial infarctions), which can reduce the heart's function or even be fatal. The key risk factors for CHD are smoking, physical inactivity, and poor diet. Blood cholesterol levels are altered by consuming dietary fats. There are three main types of dietary fats—“saturated” fatty acids (SFA) and unsaturated fatty acids; the latter can be “mono” unsaturated (MUFA) or “poly” unsaturated (PUFA). Eating SFA-rich foods (for example, meat, butter, and cheese) increases the amount of LDL-C in the blood but also increases HDL-C (the “good” cholesterol) and decreases triglycerides. Eating foods that are rich in unsaturated fatty acids (for example, vegetable oils and fatty fish) decreases the amount of LDL-C and triglycerides in the blood and also raises HDL-C.
Why Was This Study Done?
Because of the connection between eating SFA and high blood LDL-C levels, reduced SFA consumption is recommended as a way to avoid CHD. However, the evidence from individual randomized controlled trials that have studied CHD events (such as heart attacks and CHD-related deaths) have been mixed and could not support this recommendation. Furthermore, dietary recommendations to reduce SFA have generally not specified any replacement, i.e., whether SFA should be replaced with carbohydrate, protein, or unsaturated fats. Because of their beneficial effects on blood LDL-C and HDL-C levels, PUFA could be one important replacement for SFA, but, surprisingly, some experts argue that eating PUFA could actually increase CHD risk. Consequently, some guidelines recommend that PUFA consumption should be limited or even reduced. In this systematic review (a study that uses predefined criteria to identify all the research on a specific topic) and meta-analysis (a statistical method for combining the results of several studies) of randomized controlled trials, the researchers assess the impact of increased PUFA consumption as replacement for SFA on CHD events.
What Did the Researchers Do and Find?
The researchers' search of the published literature, “grey” literature (doctoral dissertations, technical reports, and other documents not printed in books and journals), and contacts with relevant experts identified eight trials in which participants were randomized to increase their PUFA intake for at least a year and in which CHD events were reported. 1,042 CHD events were recorded among the 13,614 participants enrolled in these trials. In their meta-analysis, the researchers found that on average the consumption of PUFA accounted for 14.9% of total energy intake in the intervention groups compared with only 5% of total energy intake in the control groups. Participants in the intervention groups had a 19% reduced risk of CHD events compared to participants in the control groups. Put another way, each 5% increase in the proportion of energy obtained from PUFA reduced the risk of CHD events by 10%. Finally, the researchers found that the benefits associated with PUFA consumption increased with longer duration of the trials.
What Do These Findings Mean?
These findings suggest that the replacement of some dietary SFA with PUFA reduces CHD events. Because the trials included in this study looked only at replacing SFA with PUFA, it is not possible from this evidence alone to distinguish between the benefits of reducing SFA and the benefits of increasing PUFA. Furthermore, the small number of trials identified in this study all had design faults, so the risk reductions reported here may be inaccurate. However, other lines of evidence (for example, observational studies that have examined associations between the fat intake of populations and their risk of CHD) also suggest that consumption of PUFA in place of SFA reduces CHD risk. Thus, in the light of these findings, future recommendations to reduce SFA in the diet should stress the importance of replacing SFA with PUFA rather than with other forms of energy, and the current advice to limit PUFA intake should be revised.
Additional Information
Please access these Web sites via the online version of this summary at
The American Heart Association provides information about all aspects of coronary heart disease for patients, caregivers, and professionals, including advice on dietary fats (in several languages)
The UK National Health Service Choices Web site provides information about coronary heart disease
Eatwell, a resource provided by the UK Food Standards Agency, gives advice on all aspects of healthy eating, including fat consumption
MedlinePlus provides links to further resources on coronary heart disease and on cholesterol (in English and Spanish)
PMCID: PMC2843598  PMID: 20351774
7.  Ethanol Tolerance in the Yeast Saccharomyces cerevisiae Is Dependent on Cellular Oleic Acid Content 
In this investigation, we examined the effects of different unsaturated fatty acid compositions of Saccharomyces cerevisiae on the growth-inhibiting effects of ethanol. The unsaturated fatty acid (UFA) composition of S. cerevisiae is relatively simple, consisting almost exclusively of the mono-UFAs palmitoleic acid (Δ9Z-C16:1) and oleic acid (Δ9Z-C18:1), with the former predominating. Both UFAs are formed in S. cerevisiae by the oxygen- and NADH-dependent desaturation of palmitic acid (C16:0) and stearic acid (C18:0), respectively, catalyzed by a single integral membrane desaturase encoded by the OLE1 gene. We systematically altered the UFA composition of yeast cells in a uniform genetic background (i) by genetic complementation of a desaturase-deficient ole1 knockout strain with cDNA expression constructs encoding insect desaturases with distinct regioselectivities (i.e., Δ9 and Δ11) and substrate chain-length preferences (i.e., C16:0 and C18:0); and, (ii) by supplementation of the same strain with synthetic mono-UFAs. Both experimental approaches demonstrated that oleic acid is the most efficacious UFA in overcoming the toxic effects of ethanol in growing yeast cells. Furthermore, the only other UFA tested that conferred a nominal degree of ethanol tolerance is cis-vaccenic acid (Δ11Z-C18:1), whereas neither Δ11Z-C16:1 nor palmitoleic acid (Δ9Z-C16:1) conferred any ethanol tolerance. We also showed that the most ethanol-tolerant transformant, which expresses the insect desaturase TniNPVE, produces twice as much oleic acid as palmitoleic acid in the absence of ethanol and undergoes a fourfold increase in the ratio of oleic acid to palmitoleic acid in response to exposure to 5% ethanol. These findings are consistent with the hypothesis that ethanol tolerance in yeast results from incorporation of oleic acid into lipid membranes, effecting a compensatory decrease in membrane fluidity that counteracts the fluidizing effects of ethanol.
PMCID: PMC150070  PMID: 12620835
8.  Non-covalent interaction of phospholipase A2 (PLA2) and kaouthiotoxin (KTX) from venom of Naja kaouthia exhibits marked synergism to potentiate their cytotoxicity on target cells 
Present study shows that non-covalent interaction of kaouthiotoxin (KTX) with their respective pohospholipase A2 (PLA2) from the venom of N. kaouthia displayed marked synergism to exert cytotoxicity without altering the biochemical properties of PLA2. For example, although NK-PLA2 or KTX alone did not induce appreciable hemolysis of washed human erythrocytes; however, the hemolytic potency of NK-PLA2: KTX complex was significantly higher. Identically, selective lysis of virus infected Sf9 and normal Tn insect cells was further enhanced by the cognate NK-PLA2: KTX complex as compared to individual components of the complex. Gas-chromatographic analysis of fatty acids released from intact erythrocytes by cytotoxic action of individual NK-PLA2 and NK-PLA2: KTX complex demonstrated that ratio between saturated fatty acids (SFA) and unsaturated FA (UFA) was increasing with time of hydrolysis of RBC either in the case of NK-PLA2 or NK-PLA2-KTX complex suggesting NK-PLA2-KTX complex apparently displayed the more preference for glycerophospholipids with SFAs on the sn-2 position. Therefore, it may be suggested that KTX first destabilize the target cell membrane followed by higher enzymatic activity of PLA2 on dislocated and disorganized phospholipid bilayers resulting in a significantly higher (p < 0.05) membrane damage by NK-PLA2-KTX complex compared to individual components of the complex.
PMCID: PMC3086192  PMID: 21544180
Cytotoxicity; cobra venom; kaouthiotoxins; Naja kaouthia; phospholipase A2; protein-protein interaction
9.  Dietary unsaturated fatty acids affect the mammary gland integrity and health in lactating dairy cows 
BMC Proceedings  2011;5(Suppl 4):S35.
Information about the effects of unsaturated fatty acids (UFA) supplementation on the health and integrity of the mammary gland in lactating dairy cows is lacking. Therefore, the aim of this study was to determine the effects of unprotected dietary UFA on the global expression pattern of genes in the mammary gland tissue of grazing dairy cows, and to translate this information into relevant biological knowledge.
Twenty-eight Holstein-Friesian dairy cows were randomly assigned to 4 different concentrated UFA-sources for 23 days after which all cows were switched to a non-UFA-supplemented concentrate for an additional 28 days. On the last day of both periods, mammary gland biopsies were taken to study genome-wide differences in gene expression on Bovine Genome Arrays.
Supplementation with UFA reduced the concentration of short chain fatty acids (FA), C16 FA and saturated FA in the milk, whereas that of trans-FA increased. One major finding was that canonical pathways associated with remodelling and immune functions of the mammary gland were predominantly down-regulated during UFA supplementation and negatively correlated with the concentration of milk trans-FA.
Supplementing grazing dairy cows with unprotected dietary UFA can affect the remodelling and immune functions of the mammary gland with potential consequences for its integrity and health, as well as milk quality.
PMCID: PMC3108231  PMID: 21645316
10.  Coordinated Defects in Hepatic Long Chain Fatty Acid Metabolism and Triglyceride Accumulation Contribute to Insulin Resistance in Non-Human Primates 
PLoS ONE  2011;6(11):e27617.
Non-Alcoholic fatty liver disease (NAFLD) is characterized by accumulation of triglycerides (TG) in hepatocytes, which may also trigger cirrhosis. The mechanisms of NAFLD are not fully understood, but insulin resistance has been proposed as a key determinant.
To determine the TG content and long chain fatty acyl CoA composition profile in liver from obese non-diabetic insulin resistant (IR) and lean insulin sensitive (IS) baboons in relation with hepatic and peripheral insulin sensitivity.
Twenty baboons with varying grades of adiposity were studied. Hepatic (liver) and peripheral (mainly muscle) insulin sensitivity was measured with a euglycemic clamp and QUICKI. Liver biopsies were performed at baseline for TG content and LCFA profile by mass spectrometry, and histological analysis. Findings were correlated with clinical and biochemical markers of adiposity and insulin resistance.
Obese IR baboons had elevated liver TG content compared to IS. Furthermore, the concentration of unsaturated (LC-UFA) was greater than saturated (LC-SFA) fatty acyl CoA in the liver. Interestingly, LC-FA UFA and SFA correlated with waist, BMI, insulin, NEFA, TG, QUICKI, but not M/I. Histological findings of NAFLD ranging from focal to diffuse hepatic steatosis were found in obese IR baboons.
Liver TG content is closely related with both hepatic and peripheral IR, whereas liver LC-UFA and LC-SFA are closely related only with hepatic IR in non-human primates. Mechanisms leading to the accumulation of TG, LC-UFA and an altered UFA: LC-SFA ratio may play an important role in the pathophysiology of fatty liver disease in humans.
PMCID: PMC3220682  PMID: 22125617
11.  Effects of ensiling processes and antioxidants on fatty acid concentrations and compositions in corn silages 
Corn silage is the main dietary component used for ruminant breeding in China and is an important dietary source of fatty acids for these animals. However, little is known regarding effective means to protect the fatty acid (FA) contents in silages. In this study, we examined the changes in FA contents and compositions during corn ensiling and screened several antioxidants for their inhibition of lipid oxidation during corn ensiling.
We conducted two different experiments. In Experiment 1, corn was ensiled in 30 polyethylene bottles (bottle volume: 1 L, silage density: 600 g/dm3) and three bottles were opened at 0.5 d, 1 d, 1.5 d, 2 d, 2.5 d, 3 d, 5 d, 7 d, 14 d, and 28 d after ensiling. In Experiment 2, corn was treated with various antioxidants: (1) No additives (CK); (2) BHA (Butylated hydroxyanisole); (3) TBHQ (Tertiary butyl hydroquinone); (4) TPP (Tea polyphenols); and (5) VE (Vitamin E). These treatments were applied at 50 mg/kg and 100 mg/kg of fresh weight with each treatment replicated 3 times.
During ensiling in Experiment 1, saturated fatty acids (SFA; C16:0 and C18:0) and malondialdehyde (MDA) contents tended to increase, whereas unsaturated fatty acids (UFA; C18:1, C18:2 and C18:3) tended to decrease. However, these changes were only significant on the first 2 days of ensiling. In Experiment 2, all of the antioxidants tested affected the total FA contents and those of unsaturated fatty acids (C18:1, C18:2 and C18:3) and MDA. The effects of TBHQ and TPP were greater than those of the other antioxidants.
The reduced total FA contents in corn silages were due to unsaturated fatty acids’ oxidation during the early stages of ensiling. Adding an antioxidant could prevent fatty acids’ oxidation in corn silages.
PMCID: PMC3866975  PMID: 24304647
Antioxidant; Fatty acid; Silage
12.  Nutritional Profile of Phytococktail from Trans-Himalayan Plants 
PLoS ONE  2013;8(12):e83008.
We estimated the nutritive value, vitamin content, amino acid composition, fatty acid content, and mineral profile of a phytococktail comprising sea buckthorn (Hippophae rhamnoides), apricot (Prunus armeniaca), and roseroot (Rhodiola imbricata) from trans-Himalaya. The free vitamin forms in the phytococktail were determined by rapid resolution liquid chromatography/tandem mass spectrometry (RRLC-MS/MS). Vitamin E and B-complex vitamins were detected as the principle vitamins. Reversed-phase high performance liquid chromatography (RP-HPLC) with pre-column derivatization was used for identification and quantification of amino acids. Eight essential and eleven non-essential amino acids were quantified, and the content ranged between 76.33 and 9485.67 µg/g. Among the essential amino acids, L-methionine, L-phenylalanine, L-lysine, L-leucine, and L-histidine were found to be the dominant contributors. We also quantified the fatty acids in the phytococktail by using gas chromatography coupled with a flame ionization detector (GC-FID) with fatty acid methyl esters (FAMEs) derivatization. The analysis revealed the presence of 4 major fatty acids contributing to the total lipid content. Palmitic acid was found to be the rich source of saturated fatty acid (SFA) and constituted ∼31% of the total lipid content. Among the unsaturated fatty acids (UFAs), palmitoleic acid (43.47%), oleic acid (20.89%), and linoleic acid (4.31%) were prominent. The mineral profiling was carried out by inductively coupled plasma optical emission spectrometer (ICP-OES), and it was found to contain a number of important dietary mineral elements. The harsh climatic conditions, difficult terrain, and logistic constraints at high altitude regions of Indian trans-Himalayan cold desert lead to the scarcity of fresh fruits and vegetables. Therefore, the source of multiple vitamins, essential amino acids, fatty acids, and dietary minerals from the phytococktail would provide great health benefit in the stressful environment and could be used as a high value nutritional supplement.
PMCID: PMC3871620  PMID: 24376624
13.  The Stearoyl-Coenzyme A Desaturase 1 Is Essential for Virulence and Membrane Stress in Candida parapsilosis through Unsaturated Fatty Acid Production ▿  
Infection and Immunity  2010;79(1):136-145.
Unsaturated fatty acids (UFA) are essential components of cells. In Saccharomyces cerevisiae, stearoyl-coenzyme A (CoA) desaturase 1 (OLE1) affects cell viability through the regulation of oleic (18:1) or palmitoleic (16:1) acid production. In this study, we used a targeted gene deletion approach to determine the impact of OLE1 on the emerging human pathogenic fungus Candida parapsilosis. We found that the deletion of OLE1 resulted in an auxotrophic yeast strain (designated OLE1 KO) that required unsaturated fatty acids for growth but not saturated fatty acids. Additionally, the production of UFA by OLE1 KO yeast cells was markedly reduced, suggesting that Ole1 is essential for UFA production. In contrast to wild-type C. parapsilosis, which produced pseudohyphal growth on UFA-supplemented medium agar, pseudohyphal formation in the OLE1 KO cells was severely impaired, suggesting that Ole1 regulates morphology. Furthermore, the OLE1 KO cells were hypersensitive to various stress-inducing factors, such as salts, SDS, and H2O2, especially at the physiological temperature. The results indicate that OLE1 is essential for the stress response, perhaps through the production of UFA for cell membrane biosynthesis. The OLE1 KO cells also were hypersensitive to human and fetal bovine serum, suggesting that targeting Ole1 could suppress the dissemination of yeast cells in the bloodstream. Murine-like macrophage J774.16 more efficiently killed the OLE1 KO yeasts, and significantly larger amounts of nitric oxide were detected in cocultures of macrophages and OLE1 KO cells than with wild-type or heterozygous strains. Moreover, the disruption of OLE1 significantly reduced fungal virulence in systemic murine infection. Taken together, these results demonstrate that Ole1 regulates the pathobiology of C. parapsilosis via UFA and that the OLE1 pathway is a promising antifungal target.
PMCID: PMC3019882  PMID: 20974817
14.  Identification of a conserved protein involved in anaerobic unsaturated fatty acid synthesis in Neiserria gonorrhoeae: implications for facultative and obligate anaerobes that lack FabA 
Molecular microbiology  2011;82(2):489-501.
Transcriptome analysis of the facultative anaerobe, Neisseria gonorrhoeae, revealed that many genes of unknown function were induced under anaerobic conditions. Mutation of one such gene, NGO1024, encoding a protein belonging to the 2-nitropropane dioxygenase-like superfamiliy of proteins, was found to result in an inability of gonococci to grow anaerobically. Anaerobic growth of an NG1024 mutant was restored upon supplementation with unsaturated fatty acids (UFA), but not with the saturated fatty acid palmitate. Gonococcal fatty acid profiles confirmed that NGO1024 was involved in UFA synthesis anaerobically, but not aerobically, demonstrating that gonococci contain two distinct pathways for the production of UFAs, with a yet unidentified aerobic mechanism, and an anaerobic mechanism involving NGO1024. Expression of genes involved in classical anaerobic UFA synthesis, fabA, fabM, and fabB, was toxic in gonococci and unable to complement a NGO1024 mutation, suggesting that the chemistry involved in gonococcal anaerobic UFA synthesis is distinct from that of the classical pathway. NGO1024 homologs, which we suggest naming UfaA, form a distinct lineage within the 2-nitropropane dioxygenase-like superfamily, and are found in many facultative and obligate anaerobes that produce UFAs but lack fabA, suggesting that UfaA is part of a widespread pathway involved in UFA synthesis.
PMCID: PMC3192263  PMID: 21895795
UFA; ufaA; nitropropane dioxygenase; anaerobiosis; COG2070
15.  The Effect of Slaughter Season on the Fatty Acid Profile in Four Types of Fat Deposits in Crossbred Beef Bulls 
The objective of this study was to determine the effect of slaughter season on the fatty acid profile in four types of fat deposits in crossbred (Polish Holstein Friesian Black-and-White×Limousine) beef bulls. The percentage share of fatty acids was determined by gas chromatography and were divided into the following categories of fatty acids: saturated (SFAs), unsaturated (UFAs), monounsaturated (MUFAs), polyunsaturated (PUFAs), desirable hypocholesterolemic (DFAs) and undesirable hypercholesterolemic (OFAs), n-3 and n-6. Perinephric fat was characterized by the highest SFA concentrations (59.89%), and subcutaneous fat had the highest MUFA content (50.63%). Intramuscular fat was marked by a high percentage share of PUFAs and the highest PUFA/SFA ratio. The slaughter season had a significant effect on the levels of C18:3, C20:4 (p≤0.01) and conjugated linoleic acid (p≤0.05). There was an interaction between the slaughter season and fat type for the content of C20:4 (p≤0.01) and C20:5 (p≤0.05). The results of this study show that beef from cattle slaughtered in the summer season has a higher nutritional value and more health-promoting properties.
PMCID: PMC4093157  PMID: 25049787
Beef; Fatty Acids; Gas Chromatography; Slaughter Season; Fat Deposit
16.  The inhibitory effect of cerulenin to yeasts is fungicidal 
Fatty acid biosynthesis plays a significant role in the growth and survival of diverse organisms. In yeasts, the de novo fatty acid synthesis (FAS) pathway produces and regulates essential fatty acid species such as saturated (SFA) and unsaturated (UFA) fatty acids that are required for generation and maintenance of cell membranes. Inhibition of enzymes in this pathway, such as fatty acid synthase and fatty acid desaturase, impede yeast cell growth unless appropriate exogenous fatty acids are provided.1,2 Although, the fatty acid biosynthesis pathway is essential to yeast cells, exploration of this pathway for combating fungal infections has been largely neglected. We and others have shown that deletion of a fatty acid synthase dramatically attenuates the virulence of the yeast Candida parapsilosis 2 and Candida albicans.1 Significantly, our data has revealed that inhibition of FAS enzymes results in the hypersensitivity of the yeast to serum, indicating that targeting this pathway is potentially an ideal way to combat systemic yeast infections.2 We demonstrated that using the minimal inhibitory concentration of cerulenin, a fatty acid synthase inhibitor, we could kill the wild type yeast cells in serum.2 Thus, the inhibitory effect of cerulenin (ie. blockade of the FAS pathway) on the yeast cells is fungicidal.
PMCID: PMC3306321  PMID: 22448300
17.  Palmitoleic acid prevents palmitic acid-induced macrophage activation and consequent p38 MAPK-mediated skeletal muscle insulin resistance 
Molecular and Cellular Endocrinology  2014;393(1-2):129-142.
•Palmitate-treated macrophage-conditioned medium causes myotube insulin resistance.•This involves activation of myotube p38 mitogen activated protein kinase.•Conditioned medium effects are mediated by tumour necrosis factor-α.•These effects are prevented by addition of palmitoleate.•Palmitoleate treatment of macrophages is insulin sensitising for myotubes.
Obesity and saturated fatty acid (SFA) treatment are both associated with skeletal muscle insulin resistance (IR) and increased macrophage infiltration. However, the relative effects of SFA and unsaturated fatty acid (UFA)-activated macrophages on muscle are unknown. Here, macrophages were treated with palmitic acid, palmitoleic acid or both and the effects of the conditioned medium (CM) on C2C12 myotubes investigated. CM from palmitic acid-treated J774s (palm-mac-CM) impaired insulin signalling and insulin-stimulated glycogen synthesis, reduced Inhibitor κBα and increased phosphorylation of p38 mitogen-activated protein kinase (MAPK) and c-Jun N-terminal kinase in myotubes. p38 MAPK inhibition or siRNA partially ameliorated these defects, as did addition of tumour necrosis factor-α blocking antibody to the CM. Macrophages incubated with both FAs generated CM that did not induce IR, while palmitoleic acid-mac-CM alone was insulin sensitising. Thus UFAs may improve muscle insulin sensitivity and counteract SFA-mediated IR through an effect on macrophage activation.
PMCID: PMC4148479  PMID: 24973767
ABAF, anti-bacterial, anti-fungal; ANOVA, analysis of variance; AS160, Akt substrate of 160 kDa; BSA, bovine serum albumin; CM, conditioned medium; CXCL2, Chemokine (C-X-C motif) ligand 2; DMEM, Dulbecco’s modified Eagle's medium; DMSO, dimethylsulphoxide; ERK, extracellular signal-related kinase; FA, fatty acid; FBS, foetal bovine serum; GLUT, glucose transporter; GSK, glycogen synthase kinase; IKK, inhibitor κ kinase; IκBα, inhibitor κBα; IL, interleukin; iNOS, inducible nitric oxide synthase; IR, insulin resistance; IRS1, insulin receptor substrate-1; JNK, C-jun n-terminal kinase; LPS, lipopolysaccharide; mac, macrophage; MAPK, mitogen-activated protein kinase; MCP1, monocyte chemoattractant protein; NFκB, nuclear factor-κB; PI3K, phosphoinositol 3-kinase; palm, palmitate; PBS, phosphate-buffered saline; PKC, protein kinase C; PMA, phorbol myristate acetate; RIPA, radioimmunoprecipitation; SDS-PAGE, sodium dodecyl sulphate, polyacrylamide gel electrophoresis; SFA, saturated fatty acid; siRNA, small interfering RNA; T2D, type 2 diabetes; TLR, Toll-like Receptor; TNFα, tumour necrosis factor-α; UFA, unsaturated fatty acid; Fatty acid; Tumour necrosis factor-α; p38 Mitogen-activated protein kinase; Insulin resistance; Skeletal muscle; Macrophage
18.  Production of free monounsaturated fatty acids by metabolically engineered Escherichia coli 
Monounsaturated fatty acids (MUFAs) are the best components for biodiesel when considering the low temperature fluidity and oxidative stability. However, biodiesel derived from vegetable oils or microbial lipids always consists of significant amounts of polyunsaturated and saturated fatty acids (SFAs) alkyl esters, which hampers its practical applications. Therefore, the fatty acid composition should be modified to increase MUFA contents as well as enhancing oil and lipid production.
The model microorganism Escherichia coli was engineered to produce free MUFAs. The fatty acyl-ACP thioesterase (AtFatA) and fatty acid desaturase (SSI2) from Arabidopsis thaliana were heterologously expressed in E. coli BL21 star(DE3) to specifically release free unsaturated fatty acids (UFAs) and convert SFAs to UFAs. In addition, the endogenous fadD gene (encoding acyl-CoA synthetase) was disrupted to block fatty acid catabolism while the native acetyl-CoA carboxylase (ACCase) was overexpressed to increase the malonyl coenzyme A (malonyl-CoA) pool and boost fatty acid biosynthesis. The finally engineered strain BL21ΔfadD/pE-AtFatAssi2&pA-acc produced 82.6 mg/L free fatty acids (FFAs) under shake-flask conditions and FFAs yield on glucose reached about 3.3% of the theoretical yield. Two types of MUFAs, palmitoleate (16:1Δ9) and cis-vaccenate (18:1Δ11) made up more than 75% of the FFA profiles. Fed-batch fermentation of this strain further enhanced FFAs production to a titer of 1.27 g/L without affecting fatty acid compositions.
This study demonstrated the possibility to regulate fatty acid composition by using metabolic engineering approaches. FFAs produced by the recombinant E. coli strain consisted of high-level MUFAs and biodiesel manufactured from these fatty acids would be more suitable for current diesel engines.
PMCID: PMC4021618  PMID: 24716602
Free monounsaturated fatty acids; Thioesterase; Fatty acid desaturase; acyl-CoA synthetase; acetyl-CoA carboxylase
19.  A Bacillus subtilis Gene Induced by Cold Shock Encodes a Membrane Phospholipid Desaturase 
Journal of Bacteriology  1998;180(8):2194-2200.
Bacillus subtilis grown at 37°C synthesizes saturated fatty acids with only traces of unsaturated fatty acids (UFAs). However, when cultures growing at 37°C are transferred to 20°C, UFA synthesis is induced. We report the identification and characterization of the gene encoding the fatty acid desaturase of B. subtilis. This gene, called des, was isolated by complementation of Escherichia coli strains with mutations in either of two different genes of UFA synthesis. The des gene encodes a polypeptide of 352 amino acid residues containing the three conserved histidine cluster motifs and two putative membrane-spanning domains characteristic of the membrane-bound desaturases of plants and cyanobacteria. Expression of the des gene in E. coli resulted in desaturation of palmitic acid moieties of the membrane phospholipids to give the novel mono-UFA cis-5-hexadecenoic acid, indicating that the B. subtilis des gene product is a Δ5 acyl-lipid desaturase. The des gene was disrupted, and the resulting null mutant strains were unable to synthesize UFAs upon a shift to low growth temperatures. The des null mutant strain grew as well as its congenic parent at 20 or 37°C but showed severely reduced survival during stationary phase. Analysis of operon fusions in which the des promoter directed the synthesis of a lacZ reporter gene showed that des expression is repressed at 37°C, but a shift of cultures from 37 to 20°C resulted in a 10- to 15-fold increase in transcription. This is the first report of a membrane phospholipid desaturase in a nonphotosynthetic organism and the first direct evidence for cold induction of a desaturase.
PMCID: PMC107148  PMID: 9555904
20.  Evolved differences in larval social behavior mediated by novel pheromones 
eLife  null;3:e04205.
Pheromones, chemical signals that convey social information, mediate many insect social behaviors, including navigation and aggregation. Several studies have suggested that behavior during the immature larval stages of Drosophila development is influenced by pheromones, but none of these compounds or the pheromone-receptor neurons that sense them have been identified. Here we report a larval pheromone-signaling pathway. We found that larvae produce two novel long-chain fatty acids that are attractive to other larvae. We identified a single larval chemosensory neuron that detects these molecules. Two members of the pickpocket family of DEG/ENaC channel subunits (ppk23 and ppk29) are required to respond to these pheromones. This pheromone system is evolving quickly, since the larval exudates of D. simulans, the sister species of D. melanogaster, are not attractive to other larvae. Our results define a new pheromone signaling system in Drosophila that shares characteristics with pheromone systems in a wide diversity of insects.
eLife digest
The release of chemical signals called pheromones is a common tactic used by animals in many social situations, such as to attract potential mates or to follow trails left by other members of their colony. Larvae of the fruit fly Drosophila melanogaster—a species commonly studied in the laboratory—gather together when sharing a food source and then cooperate in a way that may increase how efficiently they feed. It has been proposed that pheromones coordinate this behavior, but no larval pheromones had been identified.
Mast et al. noticed that Drosophila larvae crawling on a surface tended to occupy areas where other larvae had crawled before. This suggested that larvae had left attractive chemicals on the surface. Mast et al. identified two such substances by analyzing the chemicals left on the surface and then by testing the response of larvae to each compound.
Ultimately, Mast et al. found that a single sensory neuron in the larva is responsible for detecting these attractive chemical signals. Furthermore, two genes called pickpocket23 and pickpocket29 control this response. These genes were previously known for their roles in detecting sex pheromones, and they are members of a diverse family of calcium channel subunits that are involved in detecting multiple ‘sensory modalities’ such as touch and taste. When either pickpocket23 or pickpocket29 are inactivated, larvae ignore the social cues left by their neighbors.
Mast et al. also looked for an evolutionary role for these pheromones. Larvae of a closely related fly species called Drosophila simulans produce a subtly different blend of compounds to D. melanogaster, and this blend is not attractive to any of the species tested. While Drosophila simulans larvae were not attracted to the cues left by their own species, they were attracted to the pheromones produced by Drosophila melanogaster, indicating that they retain the sensory mechanisms to detect and respond to these pheromones. These results suggest that larvae experience a rapidly evolving, complex, pheromone-rich environment that may help them tailor their behavior to survive.
PMCID: PMC4270068  PMID: 25497433
Drosophila sechellia; Drosophila simulans; pheromone; pheromone receptor; evolution; social behavior; D. melanogaster
21.  Transient and Permanent Experience with Fatty Acids Changes Drosophila melanogaster Preference and Fitness 
PLoS ONE  2014;9(3):e92352.
Food and host-preference relies on genetic adaptation and sensory experience. In vertebrates, experience with food-related cues during early development can change adult preference. This is also true in holometabolous insects, which undergo a drastic nervous system remodelling during their complete metamorphosis, but remains uncertain in Drosophila melanogaster. We have conditioned D. melanogaster with oleic (C18∶1) and stearic (C18∶0) acids, two common dietary fatty acids, respectively preferred by larvae and adult. Wild-type individuals exposed either during a transient period of development–from embryo to adult–or more permanently–during one to ten generation cycles–were affected by such conditioning. In particular, the oviposition preference of females exposed to each fatty acid during larval development was affected without cross-effect indicating the specificity of each substance. Permanent exposure to each fatty acid also drastically changed oviposition preference as well as major fitness traits (development duration, sex-ratio, fecundity, adult lethality). This suggests that D. melanogaster ability to adapt to new food sources is determined by its genetic and sensory plasticity both of which may explain the success of this generalist-diet species.
PMCID: PMC3965419  PMID: 24667657
22.  Role for fadR in unsaturated fatty acid biosynthesis in Escherichia coli. 
Journal of Bacteriology  1983;154(2):554-560.
Escherichia coli K-12 mutants constitutive for the synthesis of the enzymes of fatty acid degradation (fad) synthesize significantly less unsaturated fatty acid (UFA) than do wild-type (fadR+) strains. The constitutive fadR mutants synthesize less UFA than do fadR+) strains both in vivo and in vitro. The inability of fadR strains to synthesize UFAs at rates comparable to those of fadR+ strains is phenotypically asymptomatic unless the fadR strain also carries a lesion in fabA, the structural gene for beta-hydroxydecanoyl-thioester dehydrase. Unlike fadR+ fabA(Ts) mutants, fadR fabA(Ts) strains synthesize insufficient UFA to support their growth even at low temperatures and, therefore, must be supplemented with UFA at both low and high temperatures. The low levels of UFA in fadR strains are not due to the constitutive level of fatty acid-degrading enzymes in these strains. These results suggest that a functional fadR gene is required for the maximal expression of UFA biosynthesis in E. coli.
PMCID: PMC217500  PMID: 6341354
23.  Cyclopropane ring formation in membrane lipids of bacteria. 
It has been known for several decades that cyclopropane fatty acids (CFAs) occur in the phospholipids of many species of bacteria. CFAs are formed by the addition of a methylene group, derived from the methyl group of S-adenosylmethionine, across the carbon-carbon double bond of unsaturated fatty acids (UFAs). The C1 transfer does not involve free fatty acids or intermediates of phospholipid biosynthesis but, rather, mature phospholipid molecules already incorporated into membrane bilayers. Furthermore, CFAs are typically produced at the onset of the stationary phase in bacterial cultures. CFA formation can thus be considered a conditional, postsynthetic modification of bacterial membrane lipid bilayers. This modification is noteworthy in several respects. It is catalyzed by a soluble enzyme, although one of the substrates, the UFA double bond, is normally sequestered deep within the hydrophobic interior of the phospholipid bilayer. The enzyme, CFA synthase, discriminates between phospholipid vesicles containing only saturated fatty acids and those containing UFAs; it exhibits no affinity for vesicles of the former composition. These and other properties imply that topologically novel protein-lipid interactions occur in the biosynthesis of CFAs. The timing and extent of the UFA-to-CFA conversion in batch cultures and the widespread distribution of CFA synthesis among bacteria would seem to suggest an important physiological role for this phenomenon, yet its rationale remains unclear despite experimental tests of a variety of hypotheses. Manipulation of the CFA synthase of Escherichia coli by genetic methods has nevertheless provided valuable insight into the physiology of CFA formation. It has identified the CFA synthase gene as one of several rpoS-regulated genes of E. coli and has provided for the construction of strains in which proposed cellular functions of CFAs can be properly evaluated. Cloning and manipulation of the CFA synthase structural gene have also enabled this novel but extremely unstable enzyme to be purified and analyzed in molecular terms and have led to the identification of mechanistically related enzymes in clinically important bacterial pathogens.
PMCID: PMC232619  PMID: 9409147
24.  Assessment of erythrocyte phospholipid fatty acid composition as a biomarker for dietary MUFA, PUFA or saturated fatty acid intake in a controlled cross-over intervention trial 
Dietary intervention trials rely on self-reported measures of intake for assessment of energy and macronutrient composition. Dietary fat intake is of particular interest due to strong associations with pathophysiology. In epidemiological trials phospholipid fatty acid composition may reflect composition of habitual diet, although strong correlations have been identified only for essential polyunsaturated fatty acids (PUFAs). Preliminary evidence shows that saturated fatty acids (SFA) C15:0 and C17:0 may be acceptable biomarkers. This study measured changes in erythrocyte membrane fatty acids during a period of strictly controlled fat feeding to investigate their use as a short-term marker of compliance, particularly for intake of SFAs.
This was a randomised cross-over trial in which diet was provided and strictly controlled. 20 healthy, male subjects were given a 40 energy % (en%) fat diet, high in saturated (high-SFA, 20 en%) or unsaturated (high-USFA, 24 en%) fatty acids for 2 periods of 3 weeks. Subjects were residential during intervention with all food and beverages provided. Dietary composition was verified by direct chemical analysis. Blood samples were collected on days 1,7,14, 21 and analysed for red blood cell (RBC) membrane fatty acid composition. Pearson correlation showed RBC fatty acid composition to mimic dietary composition by 3 weeks, but the relationships were weak. Of the SFAs only RBC C16:0 decreased in response to decreased dietary content on high-USFA treatment (ANOVA, diet, P < 0.05). Of the USFAs, higher levels of C18:1 MUFA, C20:4 and C22:6 long chain PUFA on high-USFA diet lead to higher C18:1, C20:4 and C22:6 within RBCs (ANOVA, time*diet, P < 0.05). Pearson's correlation was significant between dietary and RBC fatty acids during the 21d dietary manipulation for C18:1, and C20:5, C22:6 only (P < 0.05).
RBC membrane fatty acids cannot reliably be used as an independent measure of compliance for dietary SFA intake in short-term studies. The MUFA oleic acid and PUFAs EPA and DHA may be more useful as markers of compliance during short term intervention trials.
PMCID: PMC1334191  PMID: 16329761
erythrocyte phospholipids; fatty acids; biomarkers; residential intervention
25.  Mga2p Processing by Hypoxia and Unsaturated Fatty Acids in Saccharomyces cerevisiae: Impact on LORE-Dependent Gene Expression 
Eukaryotic Cell  2002;1(3):481-490.
In Saccharomyces cerevisiae, OLE1 encodes a Δ9 fatty acid desaturase, an enzyme that plays a critical role in maintaining the correct ratio of saturated to monounsaturated fatty acids in the cell membrane. Previous studies have demonstrated that (i) OLE1 expression is repressed by unsaturated fatty acids (UFAs) and induced by low oxygen tension, (ii) a component of this regulation is mediated through the same low oxygen response element (LORE) in the OLE1 promoter, and (iii) Mga2p is involved in LORE-dependent hypoxic induction of OLE1. We now report that LORE-CYC1 basal promoter-lacZ fusion reporter assays demonstrate that UFAs repress the reporter expression under hypoxic conditions in a dose-dependent manner via LORE. Electrophoretic mobility shift assays show that UFAs repress the hypoxia-induced complex formation with LORE. Studies with a construct encoding a truncated form of Mga2p support the hypothesis that both hypoxia and UFA signals affect the processing of Mga2p and the UFA repression of OLE1 hypoxic induction is mediated through Mga2p. Data from Western blot assays provide evidence that under normoxic conditions, Mga2p processing produces approximately equimolar levels of the membrane-bound and processed forms and is unaffected by UFAs. Hypoxic induction of OLE1, however, is associated with increased processing of the protein, resulting in an approximately fivefold increase in the soluble active form that is counteracted by exposure of the cells to unsaturated fatty acids. Data from this study suggest that the Mga2p-LORE interaction plays an important role in OLE1 expression under both normoxic and hypoxic conditions.
PMCID: PMC118009  PMID: 12455996

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