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1.
Sheng Wu Gong Cheng Xue Bao ; 36(10): 2171-2180, 2020 Oct 25.
Artigo em Chinês | MEDLINE | ID: mdl-33169581

RESUMO

In most insects, polyunsaturated fatty acids (PUFAs) are mainly polyunsaturated fatty acids with a carbon-chain length less than 18 carbon atoms, hardly any long-chain polyunsaturated fatty acids such as C20 and C22 that are more valuable and bioactive. This study, by using Drosophila melanogaster (Fruit fly) as a model organism, optimized the Δ6-fatty acid elongase enzyme Elovl5 gene from mice and transferred it to fruit flies for expression. Vectors containing Elovl5 gene were successfully injected into drosophila embryo through the microscopic injection. There were enhanced green fluorescent proteins expressed in the whole developmental stage of Drosophila be means of fluorescence microscope. At the same time, expression of Elovl5 gene significantly contributed to the transformation of fruit flies C18-polyunsaturated fatty acids in the body towards the biosynthesis of longer-chain polyunsaturated fatty acids. The transgenic fruit fly model rich in long-chain polyunsaturated fatty acids such as C20 and C22 were obtained, providing a basis for further research on biosynthesis of polyunsaturated fatty acids in fruit flies.


Assuntos
Acetiltransferases , Drosophila melanogaster , Elongases de Ácidos Graxos , Ácidos Graxos , Acetiltransferases/genética , Animais , Drosophila melanogaster/genética , Elongases de Ácidos Graxos/genética , Elongases de Ácidos Graxos/metabolismo , Ácidos Graxos/biossíntese , Ácidos Graxos/genética , Técnicas de Transferência de Genes , Camundongos
2.
J Ind Microbiol Biotechnol ; 47(4-5): 403-412, 2020 May.
Artigo em Inglês | MEDLINE | ID: mdl-32372295

RESUMO

Past research has sought to improve the production of cyclopropane fatty acids by the oleaginous yeast Yarrowia lipolytica by heterologously expressing the E. coli fatty acid synthase gene and improving cultivation processes. Cyclopropane fatty acids display properties that hold promise for biofuel applications. The E. coli fatty acid synthase gene was introduced into several genetic backgrounds of the yeast Y. lipolytica to optimize lipid synthesis; the mean cyclopropane fatty acid productivity was 43 mg L-1 h-1 on glucose, and the production rate reached its maximum (3.06 g L-1) after 72 h of cultivation in a bioreactor. The best strain (JMY6851) overexpressed simultaneously the E. coli cyclopropane fatty acid synthase gene under a hybrid promoter (hp8d) and Y. lipolytica LRO1 gene. In fed-batch process using crude glycerol as carbon source, JMY6851 strain displayed high lipid accumulation (78% of dry cell weight) and high biomass production (56 g L-1). After 165 h of cultivation, cyclopropane fatty acids represented 22% of the lipids produced; cyclopropane fatty acid productivity (103.3 mg L-1 h-1) was maximal at 72.5 h of cultivation.


Assuntos
Ácidos Graxos/biossíntese , Fermentação , Yarrowia/metabolismo , Biomassa , Reatores Biológicos , Ciclopropanos , Escherichia coli/genética , Escherichia coli/metabolismo , Glucose , Yarrowia/genética
3.
PLoS One ; 15(5): e0233485, 2020.
Artigo em Inglês | MEDLINE | ID: mdl-32470050

RESUMO

Antimicrobial resistance is a growing global health and economic concern. Current antimicrobial agents are becoming less effective against common bacterial infections. We previously identified pyrrolocins A and C, which showed activity against a variety of Gram-positive bacteria. Structurally similar compounds, known as pyrrolidinediones (e.g., TA-289, equisetin), also display antibacterial activity. However, the mechanism of action of these compounds against bacteria was undetermined. Here, we show that pyrrolocin C and equisetin inhibit bacterial acetyl-CoA carboxylase (ACC), the first step in fatty acid synthesis. We used transcriptomic data, metabolomic analysis, fatty acid rescue and acetate incorporation experiments to show that a major mechanism of action of the pyrrolidinediones is inhibition of fatty acid biosynthesis, identifying ACC as the probable molecular target. This hypothesis was further supported using purified proteins, demonstrating that biotin carboxylase is the inhibited component of ACC. There are few known antibiotics that target this pathway and, therefore, we believe that these compounds may provide the basis for alternatives to current antimicrobial therapy.


Assuntos
Acetil-CoA Carboxilase/antagonistas & inibidores , Proteínas de Bactérias/antagonistas & inibidores , Bactérias Gram-Positivas/efeitos dos fármacos , Bactérias Gram-Positivas/metabolismo , Pirrolidinonas/farmacologia , Tetra-Hidronaftalenos/farmacologia , Acetil-CoA Carboxilase/química , Antibacterianos/farmacologia , Proteínas de Bactérias/química , Domínio Catalítico/efeitos dos fármacos , Metabolismo Energético/efeitos dos fármacos , Ácidos Graxos/biossíntese , Perfilação da Expressão Gênica , Bactérias Gram-Positivas/crescimento & desenvolvimento , Humanos , Metabolômica , Mycobacterium tuberculosis/efeitos dos fármacos , Mycobacterium tuberculosis/metabolismo , Staphylococcus aureus/efeitos dos fármacos , Staphylococcus aureus/crescimento & desenvolvimento , Staphylococcus aureus/metabolismo
4.
Biochem Biophys Res Commun ; 526(4): 871-879, 2020 06 11.
Artigo em Inglês | MEDLINE | ID: mdl-32279995

RESUMO

LAG1 longevity assurance homolog 2 (LASS2), a highly conserved transmembrane protein, has been reported to be associated with nonalcoholic fatty liver disease (NAFLD). However, the effect of LASS2 on energy homeostasis and its mechanism remains unknown. In this study, we found lower expression levels of LASS2 in the livers of mice with liver steatosis induced by a high-fat diet (HFD) and free fatty acids (FFAs)-treated hepatocytes. In FFAs-treated Hepa1-6 cells and mouse primary hepatocytes (MPHs), LASS2 overexpression significantly decreased intracellular lipid content compared with the control cells. LASS2 overexpression also significantly upregulated lipolysis-related proteins, such as ATGL and HSL, and inhibited lipogenesis-related proteins, such as SREBP1 and FAS. In addition, the phosphorylation levels of AMPK and ACC increased significantly. On the contrary, LASS2 knockdown in FFAs-treated hepatocytes aggravated lipid accumulation via facilitating lipogenesis and inhibiting lipolysis. Further, co-IP and LC-MS analysis found that LASS2 might interacted with NDUFS2 to inhibit lipogenesis. The production of mitochondrial reactive oxygen species (mtROS) may be related to the interaction between LASS2 and NDUFS2. Collectively, we are the first time to showed that LASS2 might promote the phosphorylation of AMPK via mtROS produced by interaction with NDUFS2/OXPHOS, thus inhibiting lipogenesis.


Assuntos
Fígado Gorduroso/metabolismo , Fígado Gorduroso/patologia , Hepatócitos/metabolismo , NADH Desidrogenase/metabolismo , Fosforilação Oxidativa , Esfingosina N-Aciltransferase/metabolismo , Adenilato Quinase/metabolismo , Animais , Linhagem Celular , Regulação para Baixo , Ácidos Graxos/biossíntese , Fígado Gorduroso/genética , Regulação da Expressão Gênica , Técnicas de Silenciamento de Genes , Metabolismo dos Lipídeos , Fígado/metabolismo , Fígado/patologia , Masculino , Camundongos Endogâmicos C57BL , Camundongos Obesos , Mitocôndrias Hepáticas/metabolismo , Ligação Proteica , Espécies Reativas de Oxigênio/metabolismo , Transdução de Sinais
5.
Cell ; 180(6): 1130-1143.e20, 2020 03 19.
Artigo em Inglês | MEDLINE | ID: mdl-32160528

RESUMO

Fatty acid synthases (FASs) are central to metabolism but are also of biotechnological interest for the production of fine chemicals and biofuels from renewable resources. During fatty acid synthesis, the growing fatty acid chain is thought to be shuttled by the dynamic acyl carrier protein domain to several enzyme active sites. Here, we report the discovery of a γ subunit of the 2.6 megadalton α6-ß6S. cerevisiae FAS, which is shown by high-resolution structures to stabilize a rotated FAS conformation and rearrange ACP domains from equatorial to axial positions. The γ subunit spans the length of the FAS inner cavity, impeding reductase activities of FAS, regulating NADPH turnover by kinetic hysteresis at the ketoreductase, and suppressing off-pathway reactions at the enoylreductase. The γ subunit delineates the functional compartment within FAS. As a scaffold, it may be exploited to incorporate natural and designed enzymatic activities that are not present in natural FAS.


Assuntos
Ácido Graxo Sintases/química , Ácido Graxo Sintases/metabolismo , Proteína de Transporte de Acila/química , Proteína de Transporte de Acila/metabolismo , Aciltransferases/metabolismo , Sítios de Ligação , Domínio Catalítico , Microscopia Crioeletrônica/métodos , Cristalografia por Raios X/métodos , Ácidos Graxos/biossíntese , Ácidos Graxos/química , Modelos Moleculares , Subunidades Proteicas/química , Subunidades Proteicas/isolamento & purificação , Saccharomyces cerevisiae/metabolismo , Proteínas de Saccharomyces cerevisiae/metabolismo , Relação Estrutura-Atividade
6.
Sci Rep ; 10(1): 4107, 2020 03 05.
Artigo em Inglês | MEDLINE | ID: mdl-32139775

RESUMO

Although Alzheimer's disease (AD) is a central nervous system disease and type 2 diabetes MELLITUS (T2DM) is a metabolic disorder, an increasing number of genetic epidemiological studies show clear link between AD and T2DM. The current approach to uncovering the shared pathways between AD and T2DM involves association analysis; however such analyses lack power to discover the mechanisms of the diseases. As an alternative, we developed novel causal inference methods for genetic studies of AD and T2DM and pipelines for systematic multi-omic casual analysis to infer multilevel omics causal networks for the discovery of common paths from genetic variants to AD and T2DM. The proposed pipelines were applied to 448 individuals from the ROSMAP Project. We identified 13 shared causal genes, 16 shared causal pathways between AD and T2DM, and 754 gene expression and 101 gene methylation nodes that were connected to both AD and T2DM in multi-omics causal networks.


Assuntos
Doença de Alzheimer/etiologia , Diabetes Mellitus Tipo 2/etiologia , Doença de Alzheimer/genética , Doença de Alzheimer/metabolismo , Ácidos e Sais Biliares/biossíntese , Proteína de Ligação a CREB/metabolismo , Causalidade , Simulação por Computador , Metilação de DNA , Diabetes Mellitus Tipo 2/genética , Diabetes Mellitus Tipo 2/metabolismo , Neurônios Dopaminérgicos/metabolismo , Ácidos Graxos/biossíntese , Estudos de Associação Genética , Proteínas de Homeodomínio/metabolismo , Humanos , Cinesina/metabolismo , Quinases de Proteína Quinase Ativadas por Mitógeno/metabolismo , Fatores do Domínio POU/metabolismo , Transdução de Sinais
7.
Sci Rep ; 10(1): 3749, 2020 02 28.
Artigo em Inglês | MEDLINE | ID: mdl-32111914

RESUMO

Lipoyl synthases are key enzymes in lipoic acid biosynthesis, a co-factor of several enzyme complexes involved in central metabolism. Plant pyruvate dehydrogenase complex (PDH), located in mitochondria and plastids, catalyses the first step of fatty acid biosynthesis in these organelles. Among their different components, the E2 subunit requires the lipoic acid prosthetic group to be active. De novo lipoic acid biosynthesis is achieved by the successive action of two enzymes on octanoyl-ACP: octanoyltransferase (LIP2) and lipoyl synthase (LIP1). In this study, two plastidial lipoyl synthase genes from sunflower (Helianthus annuus L.) were identified (HaLIP1p1 and HaLIP1p2), sequenced and cloned in a heterologous production system (Escherichia coli). Gene expression studies revealed similar expression patterns for both isoforms, with a slight predominance of HaLIP1p1 in vegetative tissues and mature seeds. Tertiary structural models for these enzymes indicate they both have the same theoretical catalytic sites, using lipoyl-lys and 5-deoxyadenosine as docking substrates. The fatty acid profile of E. coli cells overexpressing HaLIP1p1 and HaLIP1p2 did not present major differences, and the in vivo activity of both proteins was confirmed by complementation of an E. coli JW0623 mutant in which lipoyl synthase is defective. Although no significant differences were detected in the total fatty acid composition of transgenic Arabidopsis thaliana seeds overexpressing any of both proteins, a lipidomic analysis revealed a redistribution of the glycerolipid species, accompanied with increased phosphatidylethanolamine (PE) content and a decrease in diacyglycerols (DAG) and phosphatidylcholine (PC). Depletion of the SAM co-factor caused by HaLIP1p1 and HaLIP1p2 overexpression in transgenic plants could explain this remodelling through its effects on PC synthesis.


Assuntos
Aciltransferases , Arabidopsis , Ácidos Graxos , Helianthus/genética , Proteínas de Plantas , Plantas Geneticamente Modificadas , Sulfurtransferases , Aciltransferases/biossíntese , Aciltransferases/genética , Arabidopsis/genética , Arabidopsis/metabolismo , Ácidos Graxos/biossíntese , Ácidos Graxos/genética , Helianthus/enzimologia , Proteínas de Plantas/biossíntese , Proteínas de Plantas/genética , Plantas Geneticamente Modificadas/genética , Plantas Geneticamente Modificadas/metabolismo , Sementes/genética , Sementes/metabolismo , Sulfurtransferases/biossíntese , Sulfurtransferases/genética
8.
Appl Microbiol Biotechnol ; 104(6): 2537-2544, 2020 Mar.
Artigo em Inglês | MEDLINE | ID: mdl-32025762

RESUMO

The oleaginous yeast Lipomyces starkeyi is a potential cost-effective source for the production of microbial lipids. Fatty acid elongases have vital roles in the syntheses of long-chain fatty acids. In this study, two genes encoding fatty acid elongases of L. starkeyi, LsELO1, and LsELO2 were identified and characterized. Heterologous expression of these genes in Saccharomyces cerevisiae revealed that LsElo1 is involved in the production of saturated long-chain fatty acids with 24 carbon atoms (C24:0) and that LsElo2 is involved in the conversion of C16 fatty acids to C18 fatty acids. In addition, both LsElo1 and LsElo2 were able to elongate polyunsaturated fatty acids. LsElo1 elongated linoleic acid (C18:2) to eicosadienoic acid (C20:2), and LsElo2 elongated α-linolenic acid (C18:3) to eicosatrienoic acid (C20:3). Overexpression of LsElo2 in L. starkeyi caused a reduction in C16 fatty acids, such as palmitic and palmitoleic acids, and an accumulation of C18 fatty acids such as oleic and linoleic acids. Our findings have the potential to contribute to the remodeling of fatty acid composition and the production of polyunsaturated long-chain fatty acids in oleaginous yeasts.


Assuntos
Elongases de Ácidos Graxos/metabolismo , Lipomyces/enzimologia , Lipomyces/genética , Ácidos Eicosanoicos/análise , Elongases de Ácidos Graxos/genética , Elongases de Ácidos Graxos/isolamento & purificação , Ácidos Graxos/biossíntese , Ácido Linoleico/análise , Ácido Oleico/análise , Saccharomyces cerevisiae/genética
9.
Sci Rep ; 10(1): 2731, 2020 02 17.
Artigo em Inglês | MEDLINE | ID: mdl-32066792

RESUMO

The aim of this work was to enhance the biodiesel quality and hydrocarbon content of green microalga B. braunii strain KMITL 2 cultivated outdoor under several salinity conditions in a batch production. The enhancement would be such that the microalgal biodiesel qualities met or exceeded the current standard so that it would be a good raw material for biodiesel production. The microalga production was in 300 L open oval ponds, among various salinity levels tested (0-20 ppt), 5 ppt was the best for hydrocarbon production, yielding 54.2 ± 0.9% hydrocarbon content and 5.1 ± 0.4 g L-1 biomass. As the microalga production was scaled up by cultivation in 3,675 L open raceway pond under the 5 ppt condition, the microalga yielded a bit higher hydrocarbon content (58.8 ± 2.9%) but much lower biomass (2.5 ± 0.5 g L-1). The production in both oval and raceway ponds yielded a nearly identical biodiesel property (61.06-67.42 cetane number) which exceeded the value specified in international standards. Therefore, it was concluded that B. braunii strain KMITL 2 can be batch cultivated in an open pond at optimum salinity to yield sufficient hydrocarbon and biomass for biodiesel production.


Assuntos
Clorófitas/efeitos dos fármacos , Ácidos Graxos/biossíntese , Hidrocarbonetos/metabolismo , Microalgas/efeitos dos fármacos , Sais/farmacologia , Biocombustíveis , Biomassa , Clorófitas/crescimento & desenvolvimento , Clorófitas/metabolismo , Meios de Cultura/química , Meios de Cultura/farmacologia , Ácidos Graxos/agonistas , Humanos , Hidrocarbonetos/agonistas , Microalgas/crescimento & desenvolvimento , Microalgas/metabolismo , Tanques , Salinidade , Tailândia
10.
Mar Biotechnol (NY) ; 22(2): 180-193, 2020 Apr.
Artigo em Inglês | MEDLINE | ID: mdl-32006128

RESUMO

In fish breeding practices, gamete maturity of females is vital to reproductive success. For some species, it is possible to estimate the female maturation status based on abdomen observation, but quite difficult for some species which mature at big size. To screen out the potential biomarker in fish blood relating to female maturation, we employed the approach integrating the UPLC-MS/MS and RNA-seq techniques to investigate the metabolites and genes reflecting the sexual maturation and spawning of female blunt snout bream Megalobrama amblycephala. The study included four groups, 1-year-old immature female individuals, 2-year-old immature female individuals, 2-year-old sexually mature female individuals, and 2-year-old sexually mature female individuals after 24 h of successful spawning. The upregulated metabolites in mature females were involved in "steroid hormone biosynthesis," "metabolic pathways," "glycerophospholipid metabolism," etc. compared with those of immature individuals. As the key intermediate of steroid hormone biosynthesis, 17α-hydroxypregnenolone exhibited the highest level in 2-year-old mature females than in the immature females. Meanwhile, the metabolites (i.e., dodecanoic acid and myristic acid) participating in fatty acid synthesis exhibited much lower levels in the females after spawning than those before spawning. In addition to the metabolites, the genes involved in ovarian steroidogenesis were significantly upregulated in the 2-year-old immature females compared to the 1-year-old immature females, indicating that the ovarian steroidogenesis plays important roles in ovarian development of M. amblycephala at the early stages. The significant upregulation of genes (i.e., itpr1, camk2, and mekk2) involved in the "GnRH signaling pathway" was observed in the mature females compared with the immature females, which indicated that the estrogen levels increased after female maturation in M. amblycephala. Moreover, many genes (e.g., gck, creb1, tf2-9, ryr2, asgr1, and creb1) regulating insulin secretion and thyroid hormone synthesis were significantly downregulated after female spawning. The dynamics of gene expression and metabolites observed in this study provide novel cues for guiding fish practical artificial reproduction.


Assuntos
Biomarcadores/sangue , Cyprinidae/fisiologia , Maturidade Sexual/fisiologia , Animais , Cyprinidae/sangue , Cyprinidae/genética , Ácidos Graxos/biossíntese , Feminino , Hormônios Esteroides Gonadais/biossíntese , Metaboloma , Ovário/fisiologia , Transdução de Sinais/genética , Transcriptoma
11.
Enzyme Microb Technol ; 134: 109487, 2020 Mar.
Artigo em Inglês | MEDLINE | ID: mdl-32044034

RESUMO

Haematococcus pluvialis could accumulate large amounts of triacylglycerol (TAG) and astaxanthin under various environmental stresses. To gain insights into the multiple defensive systems for carbon metabolism against nitrogen starvation, transcriptome analysis was performed. It was found that the genes related to carbon fixation, glycolysis, fatty acid and carotenoid biosynthesis pathways were up-regulated remarkably. Glyceraldehyde 3-phosphate (G3P) biosynthesis was accelerated with the enhanced C3 and C4 pathway. Meanwhile, the pyruvate kinase (PK) and pyruvate dehydrogenase E2 component (aceF) genes were significantly increased 12.9-fold and 13.9-fold, respectively, resulting more pyruvate and acetyl-CoA generation, which were beneficial to carotenoids and fatty acid biosynthesis. Methylerythritol 4-phosphate (MEP) pathway mediated carotenoid precursor isopentenyl diphosphate (IPP) synthesis, as the all eight related genes were up-regulated. The carbon flux toward astaxanthin biosynthesis with the increased astaxanthin pathway genes. The redistribution of carbon was also promoted for TAG accumulation. In addition, the up-regulation of diacylglycerol acyltransferase (DGAT) and phospholipid: diacylglycerol acyltransferase (PDAT) genes indicated that both acyl-CoA dependent and independent pathway regulated TAG accumulation. Therefore, this work reveals the multiple defensive mechanism for carbon metabolism in response to nitrogen starvation, which extended our understanding on the carotenoids, TAG and other important metabolites synthesis.


Assuntos
Vias Biossintéticas/genética , Clorófitas/genética , Clorófitas/metabolismo , Perfilação da Expressão Gênica , Nitrogênio/metabolismo , Ciclo do Carbono , Carotenoides/metabolismo , Ácidos Graxos/biossíntese , Glicólise , Regulação para Cima
12.
World J Microbiol Biotechnol ; 36(3): 35, 2020 Feb 22.
Artigo em Inglês | MEDLINE | ID: mdl-32088779

RESUMO

Odd-chain fatty acids (OCFAs) naturally occur in bacteria, higher animals, and in plants. During recent years, they have received increasing attention due to their unique pharmacological properties and usefulness for agricultural and industrial applications. Recently, OCFAs have been identified and quantified in a few organisms, and new pharmacological functions of OCFAs have been reported. Some of the publications are related to the optimization of OCFA production through fermentation and genetic engineering. The present review aims to provide a summary on the recent progress in the field of microbial-derived OCFAs. More specifically, we outline the publications of OCFAs related to (i) different sources of OCFAs; (ii) endogenous synthesis of OCFAs; (iii) production of OCFAs through fermentation; (iv) genetic engineering related to OCFA; and (v) role of OCFAs in human health and disease. Finally, some areas that require further research are discussed.


Assuntos
Bactérias/metabolismo , Ácidos Graxos/biossíntese , Fermentação , Engenharia Genética
13.
Cell ; 180(1): 176-187.e19, 2020 01 09.
Artigo em Inglês | MEDLINE | ID: mdl-31923394

RESUMO

In response to biotic stress, plants produce suites of highly modified fatty acids that bear unusual chemical functionalities. Despite their chemical complexity and proposed roles in pathogen defense, little is known about the biosynthesis of decorated fatty acids in plants. Falcarindiol is a prototypical acetylenic lipid present in carrot, tomato, and celery that inhibits growth of fungi and human cancer cell lines. Using a combination of untargeted metabolomics and RNA sequencing, we discovered a biosynthetic gene cluster in tomato (Solanum lycopersicum) required for falcarindiol production. By reconstituting initial biosynthetic steps in a heterologous host and generating transgenic pathway mutants in tomato, we demonstrate a direct role of the cluster in falcarindiol biosynthesis and resistance to fungal and bacterial pathogens in tomato leaves. This work reveals a mechanism by which plants sculpt their lipid pool in response to pathogens and provides critical insight into the complex biochemistry of alkynyl lipid production.


Assuntos
Di-Inos/metabolismo , Ácidos Graxos/biossíntese , Álcoois Graxos/metabolismo , Lycopersicon esculentum/genética , Resistência à Doença/genética , Di-Inos/química , Ácidos Graxos/metabolismo , Álcoois Graxos/química , Regulação da Expressão Gênica de Plantas/genética , Metabolômica , Família Multigênica/genética , Doenças das Plantas/microbiologia , Folhas de Planta/metabolismo , Proteínas de Plantas/metabolismo , Plantas Geneticamente Modificadas , Estresse Fisiológico/genética
14.
Int J Mol Sci ; 21(2)2020 Jan 15.
Artigo em Inglês | MEDLINE | ID: mdl-31952262

RESUMO

Hyperlipidemia is a chronic disorder that plays an important role in the development of cardiovascular diseases, type II diabetes, atherosclerosis, hypertension, and non-alcoholic fatty liver disease. Hyperlipidemias have created a worldwide health crisis and impose a substantial burden not only on personal health but also on societies and economies. Transcription factors in the sterol regulatory element binding protein (SREBP) family are key regulators of the lipogenic genes in the liver. SREBPs regulate lipid homeostasis by controlling the expression of a range of enzymes required for the synthesis of endogenous cholesterol, fatty acids, triacylglycerol, and phospholipids. Thereby, SREBPs have been considered as targets for the treatment of metabolic diseases. The aim of this study was to investigate the beneficial functions and the possible underlying molecular mechanisms of SREBP decoy ODN, which is a novel inhibitor of SREBPs, in high-fat diet (HFD)-fed hyperlipidemic mice. Our studies using HFD-induced hyperlipidemia animal model revealed that SREBB decoy ODN inhibited the increased expression of fatty acid synthetic pathway, such as SREBP-1c, FAS, SCD-1, ACC1, and HMGCR. In addition, SREBP decoy ODN decreased pro-inflammatory cytokines, including TNF-α, IL-1ß, IL-8, and IL-6 expression. These results suggest that SREBP decoy ODN exerts its anti-hyperlipidemia effects in HFD-induced hyperlipidemia mice by regulating their lipid metabolism and inhibiting lipogenesis through inactivation of the SREPB pathway.


Assuntos
Modelos Animais de Doenças , Hiperlipidemias/prevenção & controle , Oligodesoxirribonucleotídeos/farmacologia , Proteína de Ligação a Elemento Regulador de Esterol 1/antagonistas & inibidores , Animais , Vias Biossintéticas/efeitos dos fármacos , Vias Biossintéticas/genética , Citocinas/genética , Citocinas/metabolismo , Dieta Hiperlipídica/efeitos adversos , Ácidos Graxos/biossíntese , Regulação da Expressão Gênica/efeitos dos fármacos , Hiperlipidemias/etiologia , Hiperlipidemias/genética , Mediadores da Inflamação/metabolismo , Metabolismo dos Lipídeos/efeitos dos fármacos , Metabolismo dos Lipídeos/genética , Lipogênese/efeitos dos fármacos , Lipogênese/genética , Masculino , Camundongos Endogâmicos C57BL , Oligodesoxirribonucleotídeos/genética , Proteína de Ligação a Elemento Regulador de Esterol 1/genética , Proteína de Ligação a Elemento Regulador de Esterol 1/metabolismo
15.
BMC Plant Biol ; 20(1): 21, 2020 Jan 13.
Artigo em Inglês | MEDLINE | ID: mdl-31931712

RESUMO

BACKGROUND: Triacylglycerols (TAGs) are the main composition of plant seed oil. Long-chain acyl-coenzyme A synthetases (LACSs) catalyze the synthesis of long-chain acyl-coenzyme A, which is one of the primary substrates for TAG synthesis. In Arabidopsis, the LACS gene family contains nine members, among which LACS1 and LACS9 have overlapping functions in TAG biosynthesis. However, functional characterization of LACS proteins in rapeseed have been rarely reported. RESULTS: An orthologue of the Arabidopsis LACS2 gene (BnLACS2) that is highly expressed in developing seeds was identified in rapeseed (Brassica napus). The BnLACS2-GFP fusion protein was mainly localized to the endoplasmic reticulum, where TAG biosynthesis occurs. Interestingly, overexpression of the BnLACS2 gene resulted in significantly higher oil contents in transgenic rapeseed plants compared to wild type, while BnLACS2-RNAi transgenic rapeseed plants had decreased oil contents. Furthermore, quantitative real-time PCR expression data revealed that the expression of several genes involved in glycolysis, as well as fatty acid (FA) and lipid biosynthesis, was also affected in transgenic plants. CONCLUSIONS: A long chain acyl-CoA synthetase, BnLACS2, located in the endoplasmic reticulum was identified in B. napus. Overexpression of BnLACS2 in yeast and rapeseed could increase oil content, while BnLACS2-RNAi transgenic rapeseed plants exhibited decreased oil content. Furthermore, BnLACS2 transcription increased the expression of genes involved in glycolysis, and FA and lipid synthesis in developing seeds. These results suggested that BnLACS2 is an important factor for seed oil production in B. napus.


Assuntos
Brassica napus , Coenzima A Ligases , Sementes/metabolismo , Triglicerídeos/biossíntese , Brassica napus/genética , Brassica napus/metabolismo , Coenzima A Ligases/genética , Coenzima A Ligases/metabolismo , Ácidos Graxos/biossíntese , Regulação da Expressão Gênica de Plantas , Genes de Plantas , Glicólise/genética , Metabolismo dos Lipídeos/genética , Óleos Vegetais/metabolismo , Plantas Geneticamente Modificadas/genética , Interferência de RNA , Triglicerídeos/genética
16.
Artigo em Inglês | MEDLINE | ID: mdl-31759173

RESUMO

In many γ-proteobacteria, FadR is recognized as a global transcriptional regulator: in addition to being the most prominent regulator for FA biosynthesis and degradation, the protein also mediates expression of many genes in diverse biological processes. In Shewanella oneidensis, a bacterium renowned for its respiratory versatility, FadR directly controls only a few genes. However, the FadR loss substantially increases BCFA contents and impairs growth. In this study, we showed that FadR is required to activate a number of important FA biosynthesis genes, including fabA, fabB, and fabH1. Although most of these genes are controlled by FadR in a direct manner, they are not critically responsible for the phenotypes resulting from the FadR depletion. Subsequent investigations identified BKD encoded by the bkd operon as the critical factor for enhanced BCFA production. In the absence of FadR, the bkd operon is derepressed, resulting in elevated conversion of 3MOP to 3-methylbutanoyl-CoA, one of the direct substrates for BCFA synthesis. We further showed that the growth defect of the fadR mutant is due to BCAA shortage, a scenario also attributable to excessive BKD: 3MOP, the common substrate for both BCFA and BCAA, is disproportionately used for BCFA synthesis, leading to reduced production of BCAA. Collectively, our findings reveal that the S. oneidensis FadR regulon is surely larger than previously proposed and a new mechanism by which FadR impacts bacterial physiology.


Assuntos
3-Metil-2-Oxobutanoato Desidrogenase (Lipoamida)/metabolismo , Proteínas de Bactérias/metabolismo , Ácidos Graxos/biossíntese , Regulação Bacteriana da Expressão Gênica/fisiologia , Proteínas Repressoras/metabolismo , Shewanella/fisiologia , 3-Metil-2-Oxobutanoato Desidrogenase (Lipoamida)/genética , Aminoácidos de Cadeia Ramificada/biossíntese , Proteínas de Bactérias/genética , Vias Biossintéticas/genética , Isoleucina/metabolismo , Mutação , Óperon/genética , Regulon/fisiologia , Proteínas Repressoras/genética
17.
Prostate ; 80(2): 162-172, 2020 02.
Artigo em Inglês | MEDLINE | ID: mdl-31769890

RESUMO

BACKGROUND: Prostate cancer (PC) remains a leading cause of cancer mortality and the most successful chemopreventative and treatment strategies for PC come from targeting the androgen receptor (AR). Although AR plays a key role, it is likely that other molecular pathways also contribute to PC, making it essential to identify and develop drugs against novel targets. Recent studies have identified peroxisome proliferator-activated receptor gamma (PPARγ), a nuclear receptor that regulates fatty acid (FA) metabolism, as a novel target in PC, and suggest that inhibitors of PPARγ could be used to treat existing disease. We hypothesized that PPARγ acts through AR-dependent and independent mechanisms to control PC development and growth and that PPARγ inhibition is a viable PC treatment strategy. METHODS: Immunohistochemistry was used to determine expression of PPARÒ¯ in a cohort of patients with PC. Standard molecular techniques were used to investigate the PPARÒ¯ signaling in PC cells as well a xenograft mouse model to test PPARÒ¯ inhibition in vivo. Kaplan-Meier curves were created using cBioportal. RESULTS: We confirmed the expression of PPARÒ¯ in human PC. We then showed that small molecule inhibition of PPARγ decreases the growth of AR-positive and -negative PC cells in vitro and that T0070907, a potent PPARγ antagonist, significantly decreased the growth of human PC xenografts in nude mice. We found that PPARγ antagonists or small interfering RNA (siRNA) do not affect mitochondrial activity nor do they cause apoptosis; instead, they arrest the cell cycle. In AR-positive PC cells, antagonists and siRNAs reduce AR transcript and protein levels, which could contribute to growth inhibition. AR-independent effects on growth appear to be mediated by effects on FA metabolism as the specific FASN inhibitor, Fasnall, inhibited PC cell growth but did not have an additive effect when combined with PPARγ antagonists. Patients with increased PPARÒ¯ target gene expression, but not alterations in PPARÒ¯ itself, were found to have significantly worse overall survival. CONCLUSIONS: Having elucidated the direct cancer cell effects of PPARγ inhibition, our studies have helped to determine the role of PPARγ in PC growth, and support the hypothesis that PPARγ inhibition is an effective strategy for PC treatment.


Assuntos
PPAR gama/metabolismo , Neoplasias da Próstata/metabolismo , Receptores Androgênicos/metabolismo , Animais , Benzamidas/farmacologia , Pontos de Checagem do Ciclo Celular/efeitos dos fármacos , Processos de Crescimento Celular/efeitos dos fármacos , Processos de Crescimento Celular/fisiologia , Linhagem Celular Tumoral , Ácidos Graxos/biossíntese , Humanos , Masculino , Camundongos , Camundongos Nus , Terapia de Alvo Molecular , PPAR gama/antagonistas & inibidores , Neoplasias da Próstata/tratamento farmacológico , Neoplasias da Próstata/genética , Neoplasias da Próstata/patologia , Piridinas/farmacologia , Receptores Androgênicos/genética , Transcrição Genética , Ensaios Antitumorais Modelo de Xenoenxerto
18.
Mol Cell Biochem ; 464(1-2): 11-20, 2020 Jan.
Artigo em Inglês | MEDLINE | ID: mdl-31677030

RESUMO

Sirtuin 3 (SIRT3) modulates mitochondria-localized processes and is implicated in the metabolic reprogramming of cancer cells, especially fatty acid (FA) synthesis. However, the relationship between SIRT3 and aberrant lipid synthesis in cervical cancer remains unclear. Here, we investigated the clinical relevance of SIRT3 expression in cervical squamous cell carcinoma (CSCC), cervical intraepithelial neoplasia (CIN), and normal tissues. To analyze the role of SIRT3 in CCSC in vitro, endogenous SIRT3 levels were up- and down-regulated in SiHa and C33a cells, respectively, via lentiviral-based transfection. Levels were quantified using qRT-PCR. Acetylation levels for acetyl-coA carboxylase (ACC1) were measured with the anti-acetyllysine antibody. Knockdown of SIRT3 reduced levels of cellular lipid content in cells. To investigate the role of SIRT3 in cell proliferation, nude mice were xenografted with SIRT3-overexpressing or SIRT3-knockdown CCSC cells. Overall, the results demonstrate that SIRT3 significantly contributed to the reprogramming of FA synthesis in CCSC by up-regulating ACC1 to promote de novo lipogenesis by SIRT3 deacetylation. Moreover, the findings show that the SIRT3-mediated regulation of FA synthesis played a critical role in the proliferation and metastasis of CCSC cells, suggesting that SIRT3 has therapeutic potential in CCSC treatment.


Assuntos
Carcinoma de Células Escamosas/enzimologia , Ácido Graxo Sintase Tipo I/metabolismo , Proteínas de Neoplasias/metabolismo , Sirtuína 3/metabolismo , Neoplasias do Colo do Útero/enzimologia , Animais , Carcinoma de Células Escamosas/genética , Carcinoma de Células Escamosas/patologia , Linhagem Celular Tumoral , Ácido Graxo Sintase Tipo I/genética , Ácidos Graxos/biossíntese , Ácidos Graxos/genética , Feminino , Humanos , Camundongos , Camundongos Nus , Invasividade Neoplásica , Metástase Neoplásica , Proteínas de Neoplasias/genética , Sirtuína 3/genética , Neoplasias do Colo do Útero/genética , Neoplasias do Colo do Útero/patologia
19.
Gut ; 69(1): 177-186, 2020 01.
Artigo em Inglês | MEDLINE | ID: mdl-30954949

RESUMO

OBJECTIVE: Increased de novo fatty acid (FA) synthesis and cholesterol biosynthesis have been independently described in many tumour types, including hepatocellular carcinoma (HCC). DESIGN: We investigated the functional contribution of fatty acid synthase (Fasn)-mediated de novo FA synthesis in a murine HCC model induced by loss of Pten and overexpression of c-Met (sgPten/c-Met) using liver-specific Fasn knockout mice. Expression arrays and lipidomic analysis were performed to characterise the global gene expression and lipid profiles, respectively, of sgPten/c-Met HCC from wild-type and Fasn knockout mice. Human HCC cell lines were used for in vitro studies. RESULTS: Ablation of Fasn significantly delayed sgPten/c-Met-driven hepatocarcinogenesis in mice. However, eventually, HCC emerged in Fasn knockout mice. Comparative genomic and lipidomic analyses revealed the upregulation of genes involved in cholesterol biosynthesis, as well as decreased triglyceride levels and increased cholesterol esters, in HCC from these mice. Mechanistically, loss of Fasn promoted nuclear localisation and activation of sterol regulatory element binding protein 2 (Srebp2), which triggered cholesterogenesis. Blocking cholesterol synthesis via the dominant negative form of Srebp2 (dnSrebp2) completely prevented sgPten/c-Met-driven hepatocarcinogenesis in Fasn knockout mice. Similarly, silencing of FASN resulted in increased SREBP2 activation and hydroxy-3-methyl-glutaryl-CoA (HMG-CoA) reductase (HMGCR) expression in human HCC cell lines. Concomitant inhibition of FASN-mediated FA synthesis and HMGCR-driven cholesterol production was highly detrimental for HCC cell growth in culture. CONCLUSION: Our study uncovers a novel functional crosstalk between aberrant lipogenesis and cholesterol biosynthesis pathways in hepatocarcinogenesis, whose concomitant inhibition might represent a therapeutic option for HCC.


Assuntos
Carcinoma Hepatocelular/metabolismo , Colesterol/biossíntese , Ácido Graxo Sintase Tipo I/metabolismo , Ácidos Graxos/biossíntese , Neoplasias Hepáticas/metabolismo , Animais , Vias Biossintéticas/efeitos dos fármacos , Vias Biossintéticas/genética , Carcinogênese/genética , Carcinoma Hepatocelular/genética , Linhagem Celular Tumoral , Ácido Graxo Sintase Tipo I/genética , Feminino , Técnicas de Silenciamento de Genes , Inativação Gênica , Genômica , Humanos , Hidroximetilglutaril-CoA Redutases/genética , Hidroximetilglutaril-CoA Redutases/metabolismo , Lipidômica , Neoplasias Hepáticas/genética , Masculino , Camundongos , Camundongos Knockout , PTEN Fosfo-Hidrolase/metabolismo , Proteínas Proto-Oncogênicas c-met/metabolismo , Proteína de Ligação a Elemento Regulador de Esterol 2/genética , Proteína de Ligação a Elemento Regulador de Esterol 2/metabolismo , Transcriptoma
20.
Genes (Basel) ; 10(12)2019 12 01.
Artigo em Inglês | MEDLINE | ID: mdl-31805727

RESUMO

Maize is an important oil seed crop and a major food crop in different parts of the world. Since maize has relatively lower seed oil content as compared to other oil crops, efforts are continuing to improve its oil content percentage. In this study, we analyzed two contrasting maize genotypes with differential oil accumulation percentages. High oil-content (HOC) maize had 11% oil content while low oil-content (LOC) maize had significantly lower oil content (5.4%). Transmission electron microscopy revealed a higher accumulation of oil bodies in the HOC maize embryo as compared to LOC maize. Comparative RNA-sequencing analysis at different developmental stages of the seed embryos identified 739 genes that are constantly differentially expressed (DEGs) at all the six developmental stages from 15 days after pollination (DAP) to 40 DAP. Kyoto Encyclopedia of Genes and Genomes (KEGG) enrichment analysis identified fatty acid metabolism and fatty acid biosynthesis as the most enriched biological pathways contributed by these DEGs. Notably, transcriptional changes are more intense at the early stages of embryo development as compared to later stages. In addition, pathways related to oil biosynthesis and their corresponding genes were more enriched at 30 DAP, which seems to be the key stage for oil accumulation. The study also identified 33 key DEGs involved in fatty acid and triacylglycerols biosynthesis, most of which were up-regulated in HOC, that may shape the differential oil contents in the two contrasting maize. Notably, we discovered that both acyl-CoA-dependent and acyl-CoA-independent processes are essential for the high oil accumulation in maize embryo.


Assuntos
Ácidos Graxos/genética , Regulação da Expressão Gênica de Plantas/fisiologia , Genes de Plantas , Genótipo , Sementes/genética , Zea mays/genética , Ácidos Graxos/biossíntese , Óleos Vegetais/metabolismo , Sementes/embriologia , Especificidade da Espécie , Zea mays/crescimento & desenvolvimento
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