scholarly journals Mga2p Processing by Hypoxia and Unsaturated Fatty Acids in Saccharomyces cerevisiae: Impact on LORE-Dependent Gene Expression

2002 ◽  
Vol 1 (3) ◽  
pp. 481-490 ◽  
Author(s):  
Yide Jiang ◽  
Michael J. Vasconcelles ◽  
Sharon Wretzel ◽  
Anne Light ◽  
Laura Gilooly ◽  
...  
Keyword(s):  
Saccharomyces Cerevisiae ◽  
Fatty Acids ◽  
Unsaturated Fatty Acids ◽  
Critical Role ◽  
Active Form ◽  
Dependent Manner ◽  
Low Oxygen ◽  
Mobility Shift ◽  
Hypoxic Induction ◽  
Hypoxic Conditions

ABSTRACT 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.

scholarly journals MGA2 Is Involved in the Low-Oxygen Response Element-Dependent Hypoxic Induction of Genes inSaccharomyces cerevisiae

2001 ◽  
Vol 21 (18) ◽  
pp. 6161-6169 ◽  
Author(s):  
Yide Jiang ◽  
Michael J. Vasconcelles ◽  
Sharon Wretzel ◽  
Anne Light ◽  
Charles E. Martin ◽  
...  
Keyword(s):  
Gene Expression ◽  
Unsaturated Fatty Acids ◽  
Low Oxygen ◽  
Mobility Shift ◽  
Response Element ◽  
Hypoxic Induction ◽  
Hypoxic Conditions ◽  
Treatment Conditions ◽  
Reporter Assays ◽  
Electrophoretic Mobility Shift Assays

ABSTRACT Eukaryotes have the ability to respond to changes in oxygen tension by alterations in gene expression. For example,OLE1 expression in Saccharomyces cerevisiae is upregulated under hypoxic conditions. Previous studies have suggested that the pathway regulating OLE1expression by unsaturated fatty acids may involve Mga2p and Spt23p, two structurally and functionally related proteins. To define the possible roles of each of these genes on hypoxia-inducedOLE1 expression, we examined OLE1expression under normoxia, hypoxia, and cobalt treatment conditions in Δmga2 or Δspt23 deletion strains. The results of OLE1promoter-lacZ reporter gene and Northern blot analyses showed that hypoxia- and cobalt-induced OLE1 expression was dramatically decreased in a Δmga2 strain but not in a Δspt23 strain. Further analyses using low-oxygen response element (LORE)-CYC1-lacZ fusion reporter assays and electrophoretic mobility shift assays (EMSAs) demonstrated that MGA2 significantly affects the LORE-dependent hypoxic induction pathway of gene expression. When MGA2 was supplied by a plasmid, the LORE-dependent hypoxia-inducible reporter expression was recovered, as was the hypoxia-inducible complex in EMSAs in the S. cerevisiae Δmga2 strain. Supershift analysis of EMSAs using crude extracts containing mycMga2p indicated that Mga2p is a component of the LORE-binding complex. Another LORE-dependent, hypoxia-inducible gene, ATF1, was similarly affected in the Δmga2 strain. These results indicate thatMGA2 is required for the LORE-dependent hypoxic gene induction in S. cerevisiae.

Yeast desaturases

2002 ◽  
Vol 30 (6) ◽  
pp. 1080-1082 ◽  
Author(s):  
C. E. Martin ◽  
C.-S. Oh ◽  
P. Kandasamy ◽  
R. Chellapa ◽  
M. Vemula
Keyword(s):  
Fatty Acids ◽  
Fatty Acid ◽  
Mrna Stability ◽  
Unsaturated Fatty Acids ◽  
Critical Role ◽  
Fatty Acid Desaturase ◽  
Proteolytic Processing ◽  
Low Oxygen ◽  
Response Element ◽  
Membrane Bound

The Saccharomyces OLE1 gene encodes the intrinsic membrane-bound Δ-9 fatty acid desaturase. OLE1 expression is regulated at the levels of transcription and mRNA stability by nutrient fatty acids and molecular oxygen. Its transcription is controlled through two distinct promoter elements, the fatty acid response element (FAR) region, and a downstream low-oxygen response element (LORE) that dramatically amplifies FAR-activated expression under hypoxic or cobalt-stimulated growth conditions. Transcription activation through both elements is repressed by unsaturated fatty acids. The half-life of the OLE1 mRNA is also dramatically reduced upon exposure to unsaturated fatty acids. OLE1 expression is governed by two homologous membrane-bound proteins, Spt23p and Mga2p, which activate OLE1 expression through N-terminal polypeptides that are released from the membrane through a ubiquitin-mediated mechanism that involves processing by the 23 S proteosome. Although proteolytic processing of Spt23p can be repressed by polyunsaturated fatty acids, Mga2p processing in normoxic cells appears to be regulated by a different mechanism. Mga2p is essential, however, for the induction of the high levels of expression that are triggered by hypoxia through the LORE promoter element. Surprisingly, Mga2p also plays a critical role in controlling OLE1 mRNA stability, suggesting that there may be a functional linkage between OLE1 transcription and the regulation of OLE1 mRNA stability.

Evidence for a novel pathway for the targeting of a Saccharomyces cerevisiae peroxisomal protein belonging to the isomerase/hydratase family

2000 ◽  
Vol 113 (3) ◽  
pp. 533-544
Author(s):  
I.V. Karpichev ◽  
G.M. Small
Keyword(s):  
Saccharomyces Cerevisiae ◽  
Fatty Acids ◽  
Unsaturated Fatty Acids ◽  
Beta Oxidation ◽  
Peroxisomal Protein ◽  
Intracellular Targeting ◽  
Targeting Signals ◽  
Peroxisome Targeting Signals

We, and others, have identified a novel Saccharomyces cerevisiae peroxisomal protein that belongs to the isomerase/hydratase family. The protein, named Dci1p, shares 50% identity with Eci1p, a delta(3)-cis-delta(2)-trans-enoyl-CoA isomerase that acts as an auxiliary enzyme in the beta-oxidation of unsaturated fatty acids. Both of these proteins are localized to peroxisomes, and both contain motifs at their amino- and carboxyl termini that resemble peroxisome targeting signals (PTS) 1 and 2. However, we demonstrate that the putative type 1 signaling motif is not required for the peroxisomal localization of either of these proteins. Furthermore, the correct targeting of Eci1p and Dci1p occurs in the absence of the receptors for the type 1 or type 2 peroxisome targeting pathway. Together, these data suggest a novel mechanism for the intracellular targeting of these peroxisomal proteins.

The effect of pantothenate on sulfate metabolism and sulfur isotope fractionation by Saccharomyces cerevisiae

1979 ◽  
Vol 25 (10) ◽  
pp. 1139-1144 ◽  
Author(s):  
R. G. L. McCready ◽  
G. A. Din ◽  
H. R. Krouse
Keyword(s):  
Saccharomyces Cerevisiae ◽  
Fatty Acids ◽  
Sulfate Reduction ◽  
Lipid Content ◽  
Isotope Fractionation ◽  
Sulfur Isotope ◽  
Unsaturated Fatty Acids ◽  
Cellular Lipid ◽  
Sulfur Isotope Fractionation ◽  
Sulfate Metabolism

Growth of Saccharomyces cerevisiae in minimal salts – glucose – SO42− medium with varying concentrations of pantothenate (0–1000 μg/L) produced changes in the cellular lipid content and in the ratio of saturated to unsaturated fatty acids. Substantial differences in SO42−diffusion were also observed with changes in pantothenate concentration. During sulfate reduction, the δ34S value of the evolved sulfide varied with the pantothenate concentration ranging from −31‰ in the absence of pantothenate to 0‰ at 400−1000 μg/L pantothenate. The isotope selectivity is related to the effect of pantothenate concentration on cellular metabolism.

scholarly journals Overlapping Repressor Binding Sites Result in Additive Regulation of Escherichia coli FadH by FadR and ArcA

2010 ◽  
Vol 192 (17) ◽  
pp. 4289-4299 ◽  
Author(s):  
Youjun Feng ◽  
John E. Cronan
Keyword(s):  
Escherichia Coli ◽  
Fatty Acids ◽  
Cyclic Amp ◽  
Unsaturated Fatty Acids ◽  
Genetic Regulation ◽  
Receptor Protein ◽  
Anaerobic Conditions ◽  
Mobility Shift ◽  
Long Chain

ABSTRACT Escherichia coli fadH encodes a 2,4-dienoyl reductase that plays an auxiliary role in β-oxidation of certain unsaturated fatty acids. In the 2 decades since its discovery, FadH biochemistry has been studied extensively. However, the genetic regulation of FadH has been explored only partially. Here we report mapping of the fadH promoter and document its complex regulation by three independent regulators, the fatty acid degradation FadR repressor, the oxygen-responsive ArcA-ArcB two-component system, and the cyclic AMP receptor protein-cyclic AMP (CRP-cAMP) complex. Electrophoretic mobility shift assays demonstrated that FadR binds to the fadH promoter region and that this binding can be specifically reversed by long-chain acyl-coenzyme A (CoA) thioesters. In vivo data combining transcriptional lacZ fusion and real-time quantitative PCR (qPCR) analyses indicated that fadH is strongly repressed by FadR, in agreement with induction of fadH by long-chain fatty acids. Inactivation of arcA increased fadH transcription by >3-fold under anaerobic conditions. Moreover, fadH expression was increased 8- to 10-fold under anaerobic conditions upon deletion of both the fadR and the arcA gene, indicating that anaerobic expression is additively repressed by FadR and ArcA-ArcB. Unlike fadM, a newly reported member of the E. coli fad regulon that encodes another auxiliary β-oxidation enzyme, fadH was activated by the CRP-cAMP complex in a manner similar to those of the prototypical fad genes. In the absence of the CRP-cAMP complex, repression of fadH expression by both FadR and ArcA-ArcB was very weak, suggesting a possible interplay with other DNA binding proteins.

Unsaturated fatty acids and cyclooxygenase inhibitors: effects on pial arterioles

1982 ◽  
Vol 242 (4) ◽  
pp. H629-H632
Author(s):  
W. I. Rosenblum
Keyword(s):  
Fatty Acids ◽  
Unsaturated Fatty Acids ◽  
Eicosatrienoic Acid ◽  
Inhibitory Effect ◽  
Cyclooxygenase Inhibitors ◽  
Dependent Manner ◽  
Double Bonds ◽  
Pial Arterioles ◽  
Dose Dependent

Cerebral surface arterioles of the mouse were constricted in a dose-dependent manner by three different unsaturated fatty acids each with one of its double bonds in the n-6 position: arachidonate, linoleic, and 11,14,17-eicosatrienoic acid (ETA) in doses of 10-200 micrograms/ml. The constriction was transient, and its magnitude was significantly reduced by pretreatment of the mice with intraperitoneal injections of indomethacin (5 mg/kg), aspirin (100 mg/kg), or sodium 2-amino-3-(4 chlorobenzyl)-phenylacetate (AHR-6293, 100 mg/kg). The inhibitory effect of these cyclooxygenase inhibitors suggests that this enzyme is involved in the response to these fatty acids and is in keeping with suggestions in the literature stating that such unsaturated fatty acids may interact with cyclooxygenase even when they cannot form prostaglandin (PG) endoperoxides, The PG endoperoxide formed by arachidonate or the analogous hydroperoxy compounds formed by linoleic or 11,14,17 ETA, may then alter cerebrovascular tone by production of reactive, O2-containing species. Alternate explanations for the data are also proposed.

Coexpression with β1-subunit modifies the kinetics and fatty acid block of hH1α Na+channels

2000 ◽  
Vol 279 (1) ◽  
pp. H35-H46 ◽  
Author(s):  
Yong-Fu Xiao ◽  
Sterling N. Wright ◽  
Ging Kuo Wang ◽  
James P. Morgan ◽  
Alexander Leaf
Keyword(s):  
Fatty Acids ◽  
Fatty Acid ◽  
Unsaturated Fatty Acids ◽  
Monounsaturated Fatty Acids ◽  
Dependent Manner ◽  
Concentration Dependent Manner ◽  
Α Subunit ◽  
Dependent Inactivation ◽  
Voltage Dependent ◽  
Steady State Inactivation

Voltage-gated cardiac Na+ channels are composed of α- and β1-subunits. In this study β1-subunit was cotransfected with the α-subunit of the human cardiac Na+ channel (hH1α) in human embryonic kidney (HEK293t) cells. The effects of this coexpression on the kinetics and fatty acid-induced suppression of Na+currents were assessed. Current density was significantly greater in HEK293t cells coexpressing α- and β1-subunits ( I Na,αβ) than in HEK293t cells expressing α-subunit alone ( I Na,α). Compared with I Na,α, the voltage-dependent inactivation and activation of I Na,αβ were significantly shifted in the depolarizing direction. In addition, coexpression with β1-subunit prolonged the duration of recovery from inactivation. Eicosapentaenoic acid [EPA, C20:5(n–3)] significantly reduced I Na,αβ in a concentration-dependent manner and at 5 μM shifted the midpoint voltage of the steady-state inactivation by −22 ± 1 mV. EPA also significantly accelerated channel transition from the resting state to the inactivated state and prolonged the recovery time from inactivation. Docosahexaenoic acid [C22:6(n–3)], α-linolenic acid [C18:3(n–3)], and conjugated linoleic acid [C18:2(n–6)] at 5 μM significantly inhibited both I Na,αβ and I Na,α.In contrast, saturated and monounsaturated fatty acids had no effects on I Na,αβ. This finding differs from the results for I Na,α, which was significantly inhibited by both saturated and unsaturated fatty acids. Our data demonstrate that functional association of β1-subunit with hH1α modifies the kinetics and fatty acid block of the Na+ channel.

scholarly journals Characterization and Biological Activities of Seed Oil Extracted from Berberis dasystachya Maxim. by the Supercritical Carbon Dioxide Extraction Method

Molecules ◽  
2020 ◽  
Vol 25 (8) ◽  
pp. 1836
Author(s):  
Lijuan Han ◽  
Qingqing Han ◽  
Yongjing Yang ◽  
Honglun Wang ◽  
ShuLin Wang ◽  
...  
Keyword(s):  
Carbon Dioxide ◽  
Fatty Acids ◽  
Gas Chromatography ◽  
Supercritical Carbon Dioxide ◽  
Extraction Method ◽  
Unsaturated Fatty Acids ◽  
Biological Activities ◽  
Maximum Yield ◽  
Dependent Manner ◽  
Supercritical Carbon

Characterization of the structure and pharmacological activity of Berberis dasystachya Maxim., a traditional Tibetan medicinal and edible fruit, has not yet been reported. In this study, central composite design (CCD) combined with response surface methodology (RSM) was applied to optimize the extraction conditions of B. dasystachya oil (BDSO) using the supercritical carbon dioxide (SC-CO2) extraction method, and the results were compared with those obtained by the petroleum ether extraction (PEE) method. The chemical characteristics of BDSO were analyzed, and its antioxidant activity and in vitro cellular viability were studied by DPPH, ABTS, reducing power assay, and MTT assay. The results showed that the maximum yield of 12.54 ± 0.56 g/100 g was obtained at the optimal extraction conditions, which were: pressure, 25.00 MPa; temperature 59.03 °C; and CO2 flow rate, 2.25 SL/min. The Gas chromatography (GC) analysis results showed that BDSO extracted by the SC-CO2 method had higher contents of unsaturated fatty acids (85.62%) and polyunsaturated fatty acids (57.90%) than that extracted by the PEE method. The gas chromatography used in conjunction with ion mobility spectrometry (GC–IMS) results showed that the main volatile compounds in BDSO were aldehydes and esters. BDSO also exhibited antioxidant ability in a dose-dependent manner. Moreover, normal and cancer cells incubated with BDSO had survival rates of more than 85%, which indicates that BDSO is not cytotoxic. Based on these results, the BDSO extracted by the SC-CO2 method could potentially be used in other applications, e.g., those that involve using berries of B. dasystachya.

scholarly journals Expressing a cytosolic pyruvate dehydrogenase complex to increase free fatty acid production in Saccharomyces cerevisiae

2020 ◽  
Vol 19 (1) ◽  
Author(s):  
Yiming Zhang ◽  
Mo Su ◽  
Ning Qin ◽  
Jens Nielsen ◽  
Zihe Liu
Keyword(s):  
Saccharomyces Cerevisiae ◽  
Fatty Acids ◽  
Fatty Acid ◽  
Free Fatty Acid ◽  
Pyruvate Dehydrogenase ◽  
Unsaturated Fatty Acids ◽  
Pyruvate Dehydrogenase Complex ◽  
Cell Factory ◽  
Acetyl Coa ◽  
Fatty Acid Production

Abstract Background Saccharomyces cerevisiae is being exploited as a cell factory to produce fatty acids and their derivatives as biofuels. Previous studies found that both precursor supply and fatty acid metabolism deregulation are essential for enhanced fatty acid synthesis. A bacterial pyruvate dehydrogenase (PDH) complex expressed in the yeast cytosol was reported to enable production of cytosolic acetyl-CoA with lower energy cost and no toxic intermediate. Results Overexpression of the PDH complex significantly increased cell growth, ethanol consumption and reduced glycerol accumulation. Furthermore, to optimize the redox imbalance in production of fatty acids from glucose, two endogenous NAD+-dependent glycerol-3-phosphate dehydrogenases were deleted, and a heterologous NADP+-dependent glyceraldehyde-3-phosphate dehydrogenase was introduced. The best fatty acid producing strain PDH7 with engineering of precursor and co-factor metabolism could produce 840.5 mg/L free fatty acids (FFAs) in shake flask, which was 83.2% higher than the control strain YJZ08. Profile analysis of free fatty acid suggested the cytosolic PDH complex mainly resulted in the increases of unsaturated fatty acids (C16:1 and C18:1). Conclusions We demonstrated that cytosolic PDH pathway enabled more efficient acetyl-CoA provision with the lower ATP cost, and improved FFA production. Together with engineering of the redox factor rebalance, the cytosolic PDH pathway could achieve high level of FFA production at similar levels of other best acetyl-CoA producing pathways.

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