scholarly journals Genetic regulation of the bacterial omega-3 polyunsaturated fatty acid biosynthesis pathway

2020 ◽  
Author(s):  
Marco N. Allemann ◽  
Eric E. Allen
Keyword(s):  
Fatty Acids ◽  
Fatty Acid ◽  
Polyunsaturated Fatty Acids ◽  
Fatty Acid Synthesis ◽  
Fatty Acid Synthase ◽  
Genetic Regulation ◽  
Transcriptional Regulator ◽  
Acid Synthesis ◽  
Biosynthesis Pathway ◽  
Omega 3

AbstractA characteristic among many marine Gammaproteobacteria is the biosynthesis and incorporation of omega-3 polyunsaturated fatty acids into membrane phospholipids. Biosynthesis of eicosapentaenoic (EPA) and/or docosahexaenoic (DHA) acids is accomplished using a polyketide/fatty acid synthase mechanism encoded by a set of five genes pfaABCDE. This unique fatty acid synthesis (FAS) pathway co-exists with the canonical Type II dissociated fatty acid synthesis pathway, which is responsible for the biosynthesis of saturated, monounsaturated, and hydroxylated fatty acids used in phospholipid and lipid A biosynthesis. In this work, a genetic approach was undertaken to elucidate genetic regulation of the pfa genes in the model marine bacterium Photobacterium profundum SS9. Using a reporter gene fusion, we showed that expression of the pfa operon is down regulated in response to exogenous fatty acids, particularly long chain monounsaturated fatty acids. This regulation occurs independently of the canonical fatty acid regulators, FabR and FadR, present in P. profundum SS9. Transposon mutagenesis and screening of a library of mutants identified a novel transcriptional regulator, which we have designated pfaF, to be responsible for the observed regulation of the pfa operon in P. profundum SS9. Gel mobility shift and DNase I footprinting assays confirmed that PfaF binds the pfaA promoter and identified the PfaF binding site.ImportanceThe production of polyunsaturated fatty acids (PUFA) by marine Gammaproteobacteria, particularly those from deep-sea environments, has been known for decades. These unique fatty acids are produced by a polyketide-type mechanism and subsequently incorporated into the phospholipid membrane. While much research has focused on the biosynthesis genes, their products and the phylogenetic distribution of these gene clusters, no prior studies have detailed the genetic regulation of this pathway. This study describes how this pathway is regulated under various culture conditions and has identified and characterized a fatty acid responsive transcriptional regulator specific to the PUFA biosynthesis pathway.

scholarly journals Genetic Regulation of the Bacterial Omega-3 Polyunsaturated Fatty Acid Biosynthesis Pathway

2020 ◽  
Vol 202 (16) ◽  
Author(s):  
Marco N. Allemann ◽  
Eric E. Allen
Keyword(s):  
Fatty Acids ◽  
Fatty Acid ◽  
Fatty Acid Synthesis ◽  
Genetic Regulation ◽  
Transcriptional Regulator ◽  
Acid Synthesis ◽  
Omega 3 ◽  
Content Type ◽  
Fatty Acid Synthesis Pathway ◽  
Long Chain

ABSTRACT A characteristic among many marine Gammaproteobacteria is the biosynthesis and incorporation of omega-3 polyunsaturated fatty acids into membrane phospholipids. The biosynthesis of eicosapentaenoic acid (EPA) and/or docosahexaenoic acid (DHA) is mediated by a polyketide/fatty acid synthase mechanism encoded by a set of five genes, pfaABCDE. This unique fatty acid synthesis pathway coexists with the principal type II dissociated fatty acid synthesis pathway, which is responsible for the biosynthesis of core saturated, monounsaturated, and hydroxylated fatty acids used in phospholipid and lipid A biosynthesis. In this work, a genetic approach was undertaken to elucidate genetic regulation of the pfa genes in the model marine bacterium Photobacterium profundum SS9. Using a reporter gene fusion, we showed that expression of the pfa operon is downregulated in response to exogenous fatty acids, particularly long-chain monounsaturated fatty acids. This regulation occurs independently of the canonical fatty acid regulators, FabR and FadR, present in P. profundum SS9. Transposon mutagenesis and screening of a library of mutants identified a novel transcriptional regulator, which we have designated pfaF, to be responsible for the observed regulation of the pfa operon in P. profundum SS9. Gel mobility shift and DNase I footprinting assays confirmed that PfaF binds the pfaA promoter and identified the PfaF binding site. IMPORTANCE The production of long-chain omega-3 polyunsaturated fatty acids (PUFA) by marine Gammaproteobacteria, particularly those from deep-sea environments, has been known for decades. These unique fatty acids are produced by a polyketide-type mechanism and subsequently incorporated into the phospholipid membrane. While much research has focused on the biosynthesis genes, their products, and the phylogenetic distribution of these gene clusters, no prior studies have detailed the genetic regulation of this pathway. This study describes how this pathway is regulated under various culture conditions and has identified and characterized a fatty acid-responsive transcriptional regulator specific to PUFA biosynthesis.

Fatty acid synthesis: from CO2 to functional genomics

2000 ◽  
Vol 28 (6) ◽  
pp. 567-574 ◽  
Author(s):  
J. Ohlrogge ◽  
M. Pollard ◽  
X. Bao ◽  
M. Focke ◽  
T. Girke ◽  
...  
Keyword(s):  
Fatty Acids ◽  
Fatty Acid ◽  
Fatty Acid Synthesis ◽  
Regulatory Networks ◽  
Fatty Acid Synthase ◽  
Expression Patterns ◽  
Acid Synthesis ◽  
Acetyl Coa ◽  
Specific Expression ◽  
Direct Production

For over 25 years there has been uncertainty over the pathway from CO2, to acetyl-CoA in chloroplasts. On the one hand, free acetate is the most effective substrate for fatty acid synthesis by isolated chloroplasts, and free acetate concentrations reported in leaf tissue (0.1–1 mM) appear adequate to saturate fatty acid synthase. On the other hand, a clear mechanism to generate sufficient free acetate for fatty acid synthesis is not established and direct production of acetyl-CoA from pyruvate by a plastid pyruvate dehydrogenase seems a more simple and direct path. We have re-examined this question and attempted to distinguish between the alternatives. The kinetics of 13CO2 and 14CO2 movement into fatty acids and the absolute rate of fatty acid synthesis in leaves was determined in light and dark. Because administered 14C appears in fatty acids within < 2–3 min our results are inconsistent with a large pool of free acetate as an intermediate in leaf fatty acid synthesis. In addition, these studies provide an estimate of the turnover rate of fatty acid in leaves. Studies similar to the above are more complex in seeds, and some questions about the regulation of plant lipid metabolism seem difficult to solve using conventional biochemical or molecular approaches. For example, we have little understanding of why or how some seeds produce >50%, oil whereas other seeds store largely carbohydrate or protein. Major control over complex plant biochemical pathways may only become possible by understanding regulatory networks which provide ‘global’ control over these pathways. To begin to discover such networks and provide a broad analysis of gene expression in developing oilseeds, we have produced micro-arrays that display approx. 5000 seed-expressed Arabidopsis genes. Sensitivity of the arrays was 1–2 copies of mRNA/cell. The arrays have been hybridized with probes derived from seeds, leaves and roots, and analysis of expression ratios between the different tissues has allowed the tissue-specific expression patterns of many hundreds of genes to be described for the first time. Approx. 10% of the genes were expressed at ratios ≥ 10-fold higher in seeds than in leaves or roots. Included in this list are a large number of proteins of unknown function, and potential regulatory factors such as protein kinases, phosphatases and transcription factors. The arrays were also found to be useful for analysis of Brassica seeds.

scholarly journals Dependence of Arbuscular-Mycorrhizal Fungi on Their Plant Host for Palmitic Acid Synthesis

2005 ◽  
Vol 71 (9) ◽  
pp. 5341-5347 ◽  
Author(s):  
Martin Trépanier ◽  
Guillaume Bécard ◽  
Peter Moutoglis ◽  
Claude Willemot ◽  
Serge Gagné ◽  
...  
Keyword(s):  
Fatty Acids ◽  
Fatty Acid ◽  
Arbuscular Mycorrhizal Fungi ◽  
Fatty Acid Synthesis ◽  
Mycorrhizal Fungi ◽  
Fatty Acid Synthase ◽  
Arbuscular Mycorrhizal ◽  
Acid Synthesis ◽  
Fungal Mycelium ◽  
Plant Host

ABSTRACT Lipids are the major form of carbon storage in arbuscular-mycorrhizal fungi. We studied fatty acid synthesis by Glomus intraradices and Gigaspora rosea. [14C]Acetate and [14C]sucrose were incorporated into a synthetic culture medium to test fatty acid synthetic ability in germinating spores (G. intraradices and G. rosea), mycorrhized carrot roots, and extraradical fungal mycelium (G. intraradices). Germinating spores and extraradical hyphae could not synthesize 16-carbon fatty acids but could elongate and desaturate fatty acids already present. The growth stimulation of germinating spores by root exudates did not stimulate fatty acid synthesis. 16-Carbon fatty acids (16:0 and 16:1) were synthesized only by the fungi in the mycorrhized roots. Our data strongly suggest that the fatty acid synthase activity of arbuscular-mycorrhizal fungi is expressed exclusively in the intraradical mycelium and indicate that fatty acid metabolism may play a major role in the obligate biotrophism of arbuscular-mycorrhizal fungi.

Energy balances of beef tallow and soyabean oil in broiler chickens

2007 ◽  
Vol 2007 ◽  
pp. 240-240
Author(s):  
Sasiphan Wongsuthavas ◽  
Chalermpol Yuangklang ◽  
Jamlong Mitchaothai ◽  
Kraisit Vasupen ◽  
Anton Beynen
Keyword(s):  
Fatty Acids ◽  
Fatty Acid ◽  
Polyunsaturated Fatty Acids ◽  
Body Fat ◽  
Fatty Acid Synthesis ◽  
Beef Tallow ◽  
Broiler Chickens ◽  
Saturated Fatty Acids ◽  
Acid Synthesis ◽  
Abdominal Fat

From previous reports indicate that broiler chickens fed diets enriched with polyunsaturated fatty acids have less abdominal fat or total body fat (Sanz et al., 1999) deposition than do broiler chickens fed diets containing saturated fatty acids. In general, body fat accumulation may be considered the net result of the balance among dietary absorbed fat, endogenous fat synthesis (lipogenesis) and fat catabolism via β-oxidation (lypolysis). Thus, if the amount of absorbed fat is the same, lower body fat deposition may be attributed to increased fat catabolism or diminished endogenous fatty acid synthesis or to both process. Differences in lipid oxidation rates seem to be the main mechanism involved in this effect. Supplementation of unsaturated fatty acids in diets can be enhanced fat catabolism and reduced fatty acid synthesis were reported to occur in rats fed polyunsaturated fatty acid rich in diets compared with rats fed diets enriched with saturated fatty acids (Crespo and Esteve-Garcia, 2002). Polyunsaturated fatty acids rich in diet effect on fat absorption were increased. In, contrast amount of abdominal fat deposition was decreased. It may be that the polyunsaturated fatty acid was higher rate fat oxidation or lower rate of fatty acid synthesis or both. The present study was undertaken to determine the effect of amount of beef tallow versus soybean oil absorbed is the same on energy intake, energy excretion, energy retention and energy expenditure of broiler chickens.

scholarly journals Comparison of the Substrate Preferences of ω3 Fatty Acid Desaturases for Long Chain Polyunsaturated Fatty Acids

2019 ◽  
Vol 20 (12) ◽  
pp. 3058 ◽  
Author(s):  
Pushkar Shrestha ◽  
Xue-Rong Zhou ◽  
Sapna Vibhakaran Pillai ◽  
James Petrie ◽  
Robert de Feyter ◽  
...  
Keyword(s):  
Fatty Acids ◽  
Fatty Acid ◽  
Polyunsaturated Fatty Acids ◽  
Nicotiana Benthamiana ◽  
Animal Species ◽  
Biosynthesis Pathway ◽  
Omega 3 ◽  
Fatty Acid Desaturases ◽  
Substrate Preferences ◽  
Long Chain

Omega-3 long chain polyunsaturated fatty acids (ω3 LC-PUFAs) such as eicosapentaenoic acid (EPA; 20:5ω3) and docosahexaenoic acid (DHA; 22:6ω3) are important fatty acids for human health. These ω3 LC-PUFAs are produced from their ω3 precursors by a set of desaturases and elongases involved in the biosynthesis pathway and are also converted from ω6 LC-PUFA by omega-3 desaturases (ω3Ds). Here, we have investigated eight ω3-desaturases obtained from a cyanobacterium, plants, fungi and a lower animal species for their activities and compared their specificities for various C18, C20 and C22 ω6 PUFA substrates by transiently expressing them in Nicotiana benthamiana leaves. Our results showed hitherto unreported activity of many of the ω3Ds on ω6 LC-PUFA substrates leading to their conversion to ω3 LC-PUFAs. This discovery could be important in the engineering of EPA and DHA in heterologous hosts.

scholarly journals Genetic suppression of lethal mutations in fatty acid biosynthesis mediated by a secondary lipid synthase

2021 ◽  
Author(s):  
Marco N. Allemann ◽  
Eric E. Allen
Keyword(s):  
Fatty Acids ◽  
Fatty Acid ◽  
Polyunsaturated Fatty Acids ◽  
Fatty Acid Synthase ◽  
Fatty Acid Biosynthesis ◽  
Biosynthetic Pathways ◽  
Omega 3 ◽  
Genetic Redundancy ◽  
Acid Biosynthesis ◽  
Malonyl Coa

The biosynthesis and incorporation of polyunsaturated fatty acids into phospholipid membranes is a unique feature of certain marine Gammaproteobacteria inhabiting high-pressure and/or low temperature environments. In these bacteria, monounsaturated and saturated fatty acids are produced via the classical dissociated Type II fatty acid synthase mechanism, while omega-3 polyunsaturated fatty acids such as EPA (20:5n-3) and DHA (22:6n-3) are produced by a hybrid polyketide/fatty acid synthase – encoded by the pfa genes - also referred to as the secondary lipid synthase mechanism. In this work, phenotypes associated with partial or complete loss of monounsaturated biosynthesis are shown to be compensated for by several-fold increased production of polyunsaturated fatty acids in the model marine bacterium Photobacterium profundum SS9. One route to suppression of these phenotypes could be achieved by transposition of insertion sequences within or upstream of the fabD, malonyl CoA-acyl carrier protein transacylase, coding sequence. Genetic experiments in this strain indicated that fabD is not an essential gene, yet mutations in fabD and pfaA are synthetically lethal. Based on these results, we speculated that the malonyl-CoA transacylase domain within PfaA compensates for loss of FabD activity. Heterologous expression of either pfaABCD from P. profundum SS9 or pfaABCDE from Shewanella pealeana in Escherichia coli complemented the loss of the chromosomal copy of fabD in vivo. The co-occurrence of independent, yet compensatory fatty acid biosynthetic pathways in select marine bacteria may provide genetic redundancy to optimize fitness under extreme conditions. Importance A defining trait among many cultured piezophilic and/or psychrophilic marine Gammaproteobacteria is the incorporation of both monounsaturated and polyunsaturated fatty acids into membrane phospholipids. The biosynthesis of these different classes of fatty acid molecules is linked to two genetically distinct co-occurring pathways that utilize the same pool of intracellular precursors. Using a genetic approach, new insights have been gained into the interactions between these two biosynthetic pathways. Specifically, core fatty acid biosynthesis genes previously thought to be essential were found to be non-essential in strains harboring both pathways due to functional overlap between the two pathways. These results provide new routes to genetically optimize long-chain omega-3 polyunsaturated fatty acid biosynthesis in bacteria and reveal a possible ecological role for maintaining multiple pathways for lipid synthesis in a single bacterium.

scholarly journals Stimulation of fatty acid synthesis in vitro by gonadotrophin-induced testicular ribonucleic acid

1968 ◽  
Vol 108 (1) ◽  
pp. 147-152 ◽  
Author(s):  
Ajit Goswami ◽  
James K. Skipper ◽  
William L. Williams
Keyword(s):  
Fatty Acids ◽  
Fatty Acid ◽  
Luteinizing Hormone ◽  
Polyunsaturated Fatty Acids ◽  
Fatty Acid Synthesis ◽  
Follicle Stimulating Hormone ◽  
Acid Synthesis ◽  
Stimulation Of ◽  
Stimulating Hormone

RNA from testes of hypophysectomized rats treated with follicle-stimulating hormone and luteinizing hormone markedly stimulates in vitro the incorporation of acetate and malonate (as CoA derivatives) into polyunsaturated fatty acids. The system in vitro contains the components necessary for both protein and fatty acid synthesis. That the RNA is a hormone-induced messenger type that causes enzyme synthesis that then causes fatty acid synthesis is supported by the following observations: (1) the stimulation of RNA synthesis by follicle-stimulating hormone and luteinizing hormone is decreased by injection of the animals with actinomycin D; (2) puromycin in the system in vitro decreases the synthesis of polyunsaturated fatty acids; (3) the activity of the RNA preparation is destroyed by digestion with ribonuclease; in fact, the digest is inhibitory, which is a characteristic of messenger-RNA-mediated protein synthesis; (4) protein that might be denatured enzyme is virtually absent from the effective RNA preparations.

scholarly journals Reassessment of the Genetic Regulation of Fatty Acid Synthesis in Escherichia coli: Global Positive Control by the Dual Functional Regulator FadR

2015 ◽  
Vol 197 (11) ◽  
pp. 1862-1872 ◽  
Author(s):  
L. My ◽  
N. Ghandour Achkar ◽  
J. P. Viala ◽  
E. Bouveret
Keyword(s):  
Escherichia Coli ◽  
Fatty Acid ◽  
Fatty Acid Synthesis ◽  
Genetic Regulation ◽  
Transcriptional Regulator ◽  
Acid Synthesis ◽  
Biofuel Production ◽  
Global Regulator ◽  
Content Type ◽  
E Coli

ABSTRACTInEscherichia coli, the FadR transcriptional regulator represses the expression of fatty acid degradation (fad) genes. However, FadR is also an activator of the expression offabAandfabB, two genes involved in unsaturated fatty acid synthesis. Therefore, FadR plays an important role in maintaining the balance between saturated and unsaturated fatty acids in the membrane. We recently showed that FadR also activates the promoter upstream of thefabHgene (L. My, B. Rekoske, J. J. Lemke, J. P. Viala, R. L. Gourse, and E. Bouveret, J Bacteriol 195:3784–3795, 2013, doi:10.1128/JB.00384-13). Furthermore, recent transcriptomic and proteomic data suggested that FadR activates the majority of fatty acid (FA) synthesis genes. In the present study, we tested the role of FadR in the expression of all genes involved in FA synthesis. We found that FadR activates the transcription of all tested FA synthesis genes, and we identified the FadR binding site for each of these genes. This necessitated the reassessment of the transcription start sites foraccAandaccBgenes described previously, and we provide evidence for the presence of multiple promoters driving the expression of these genes. We showed further that regulation by FadR impacts the amount of FA synthesis enzymes in the cell. Our results show that FadR is a global regulator of FA metabolism inE. coli, acting both as a repressor of catabolism and an activator of anabolism, two directly opposing pathways.IMPORTANCEIn most bacteria, a transcriptional regulator tunes the level of FA synthesis enzymes. Oddly, such a global regulator still was missing forE. coli, which nonetheless is one of the prominent model bacteria used for engineering biofuel production using the FA synthesis pathway. Our work identifies the FadR functional dual regulator as a global activator of almost all FA synthesis genes inE. coli. Because FadR also is the repressor of FA degradation, FadR acts both as a repressor and an activator of the two opposite pathways of FA degradation and synthesis. Our results show that there are still discoveries waiting to be made in the understanding of the genetic regulation of FA synthesis, even in the very well-known bacteriumE. coli.

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