Δ6 Desaturase Research Articles

Associations of erythrocyte fatty acid compositions with FADS1 gene polymorphism in Japanese mothers and infants

Long-chain polyunsaturated fatty acids (LC-PUFAs) are involved in the fetal growth in utero, and are essential for the development of visual and cognitive functions during infancy. The purpose of this study was to examine the associations of erythrocyte fatty acid compositions with FADS1 gene polymorphism in Japanese mothers and infants. The subjects were 383 mothers who participated in an adjunct birth cohort study of the Japan Environment and Children's Study (JECS). In maternal FADS1 SNP genotypes, the precursor fatty acids composition of the Δ5 desaturase in the maternal blood showed significant differences in levels among the groups, and showed increasing values in the order of TT < TC < CC genotype groups. On the other hand, many product fatty acids levels were significantly reduced in the order of TT > TC > CC genotype groups, and DHA levels were significantly lower in the CC genotype group relative to the other groups. Likewise, the relationship between fetal genotype group and fatty acid composition in cord blood was very similar to the maternal relationship. These results indicate the maternal and fetal blood fatty acid compositions are strongly influenced by the FADS1 genotypes. With respect to the cord blood DHA composition, the levels in the fetal CC genotype group showed a trend toward lower values in the maternal CC genotype group pair (p = 0.066) compared to the maternal TC genotype group pair. However, in the fetal TT and TC genotype groups (p = 0.131, p = 0.729, respectively), the maternal genotype did not have a significant effect. The DHA composition was more influenced by the maternal genotype in the fetal CC genotype group than in the fetal TT and TC genotype groups. It was shown that DHA transport via the placenta from the mother might be promoted in the fetal CC genotype compared to the other fetal genotype groups. In conclusion, differences in the FADS1 SNP genotypes of pregnant women and their children may greatly affect the supply of LC-PUFAs. Further studies on the involvement of the FADS1 polymorphisms and the fetal LC-PUFA levels in the fetal growth and development are warranted.

Dietary Fish Oil Alters DNA Methylation of Genes Involved in Polyunsaturated Fatty Acid Biosynthesis in Muscle and Liver of Atlantic Salmon (Salmo salar)

Adequate dietary supply of eicosapentaenoic acid (20:5n-3) and docosahexaenoic acid (22:6n-3) is required to maintain health and growth of Atlantic salmon (Salmo salar). However, salmon can also convert α-linolenic acid (18:3n-3) into eicosapentaenoic acid (20:5n-3) and docosahexaenoic acid (22:6n-3) by sequential desaturation and elongation reactions, which can be modified by 20:5n-3 and 22:6n-3 intake. In mammals, dietary 20:5n-3 + 22:6n-3 intake can modify Fads2 expression (Δ6 desaturase) via altered DNA methylation of its promoter. Decreasing dietary fish oil (FO) has been shown to increase Δ5fad expression in salmon liver. However, it is not known whether this is associated with changes in the DNA methylation of genes involved in polyunsaturated fatty acid synthesis. To address this, we investigated whether changing the proportions of dietary FO and vegetable oil altered the DNA methylation of Δ6fad_b, Δ5fad, Elovl2, and Elovl5_b promoters in liver and muscle from Atlantic salmon and whether any changes were associated with mRNA expression. Higher dietary FO content increased the proportions of 20:5n-3 and 22:6n-3 and decreased Δ6fad_b mRNA expression in liver, but there was no effect on Δ5fad, Elovl2, and Elovl5_b expression. There were significant differences between liver and skeletal muscle in the methylation of individual CpG loci in all four genes studied. Methylation of individual Δ6fad_b CpG loci was negatively related to its expression and to proportions of 20:5n-3 and 22:6n-3 in the liver. These findings suggest variations in dietary FO can induce gene-, CpG locus-, and tissue-related changes in DNA methylation in salmon.

Molecular and functional characterisation of a putative elovl4 gene and its expression in response to dietary fatty acid profile in Atlantic bluefin tuna (Thunnus thynnus).

Elongation of very long-chain fatty acid 4 (Elovl4) proteins are involved in the biosynthesis of very long-chain (>C24) fatty acids and in many teleost fish species they are key enzymes in the pathway for the production of docosahexaenoic acid (DHA; 22:6n-3) from eicosapentaenoic acid (EPA; 20:5n-3). Therefore, Elovl4 may be particularly important in Atlantic bluefin tuna (ABT; Thunnus thynnus) characterised by having high DHA to EPA ratios. The present study cloned and characterised both the function and expression of an elovl4 cDNA from ABT. The Elovl4 had an open reading frame of 915 base pairs encoding a putative protein of 304 amino acids. Alignment and phylogenetic analyses indicated that the Elovl4 isoform identified in the present study was an Elovl4b. Functional characterisation demonstrated that the Elovl4b enzyme had elongase activity towards all the polyunsaturated fatty acid (PUFA) substrates assayed. The ABT Elovl4b contributed to DHA biosynthesis by elongation of EPA and DPA to 24:5n-3, the latter being desaturated to 24:6n-3 by the action of fads2 (Δ6 desaturase). Additionally, the ABT Elovl4b has a role in the biosynthesis of very long-chain PUFA up to C34, compounds of key structural roles in neural tissues such as eye and brain, which had high levels of elovl4b transcripts. Surprisingly, while the relative expression of fads2, required for the production of DHA from EPA, was increased in liver of ABT fed a diet with reduced levels of EPA and DHA, expression of elovl4b was reduced. Results indicated that ABT has enzymes necessary for endogenous production of DHA from EPA and demonstrate that Elovl4b can effectively compensate for absence of Elovl2.

Changes in emergence phenology, fatty acid composition, and xenobiotic-metabolizing enzyme expression is associated with increased insecticide resistance in the Colorado potato beetle.

The Colorado potato beetle (Leptinotarsa decemlineata) is a major agricultural pest of solanaceous crops. An effective management strategy employed by agricultural producers to control this pest species is the use of systemic insecticides. Recent emphasis has been placed on the use of neonicotinoid insecticides. Despite efforts to curb resistance development through integrated pest management approaches, resistance to neonicotinoids in L. decemlineata populations continues to increase. One contributing factor may be alterations in insect fatty acids, which have multiple metabolic functions and are associated with the synthesis of xenobiotic‐metabolizing enzymes to mitigate the effects of insecticide exposure. In this study, we analyzed the fatty acid composition of L. decemlineata populations collected from an organic production field and from a commercially managed field to determine if fatty acid composition varied between the two populations. We demonstrate that a population of L. decemlineata that has a history of systemic neonicotinoid exposure (commercially managed) has a different lipid composition and differential expression of known metabolic detoxification mechanisms relative to a population that has not been exposed to neonicotinoids (organically managed). The fatty acid data indicated an upregulation of Δ6 desaturase in the commercially managed L. decemlineata population and suggest a role for eicosanoids and associated metabolic enzymes as potential modulators of insecticide resistance. We further observed a pattern of delayed emergence within the commercially managed population compared with the organically managed population. Variations in emergence timing together with specific fatty acid regulation may significantly influence the capacity of L. decemlineata to develop insecticide resistance.

Dietary flaxseed and turmeric is a novel strategy to enrich chicken meat with long chain ω-3 polyunsaturated fatty acids with better oxidative stability and functional properties

The purpose of the present study was to elucidate the effects of feeding flaxseed meal (FSM) and turmeric rhizome powder (TRP) supplementation on tissue lipid profile, lipid metabolism, health indices, oxidative stability, and physical properties of broiler chicken meat. The 100 g FSM along with 10.0 g TRP supplementation significantly increased the ω-3 PUFA, particularly ALA, EPA, DPA, and DHA of broiler chicken meat due to the corresponding increase ∆9 and Δ5 + Δ6 desaturase activities. The increased activities of the desaturases resulted in significantly better health indices of the broiler chicken meat. The feeding of 100 g FSM along with 10.0 g TRP supplementation reduced the atherogenic and thrombogenic indices of broiler chicken meat. The 100 g FSM feeding reduced the oxidative stability, water holding capacity, extract release volume of broiler chicken meat and increased drip loss, whereas, 10.0 g TRP supplementation reversed these negative effects of FSM.

Microbial lipids from industrial wastes using xylose-utilizing Ashbya gossypii strains

This work presents the exploitation of waste industrial by-products as raw materials for the production of microbial lipids in engineered strains of the filamentous fungus Ashbya gossypii. A lipogenic xylose-utilizing strain was used to apply a metabolic engineering approach aiming at relieving regulatory mechanisms to further increase the biosynthesis of lipids. Three genomic manipulations were applied: the overexpression of a feedback resistant form of the acetyl-CoA carboxylase enzyme; the expression of a truncated form of Mga2, a regulator of the main Δ9 desaturase gene; and the overexpression of an additional copy of DGA1 that codes for diacylglycerol acyltransferase. The performance of the engineered strain was evaluated in culture media containing mixed formulations of corn-cob hydrolysates, sugarcane molasses or crude glycerol. Our results demonstrate the efficiency of the engineered strains, which were able to accumulate about 40% of cell dry weight (CDW) in lipid content using organic industrial wastes as feedstocks.

Genetic Silencing of Fatty Acid Desaturases Modulates α-Synuclein Toxicity and Neuronal Loss in Parkinson-Like Models of C. elegans.

The molecular basis of Parkinson’s disease (PD) is currently unknown. There is increasing evidence that fat metabolism is at the crossroad of key molecular pathways associated with the pathophysiology of PD. Fatty acid desaturases catalyze synthesis of saturated fatty acids from monounsaturated fatty acids thereby mediating several cellular mechanisms that are associated with diseases including cancer and metabolic disorders. The role of desaturases in modulating age-related neurodegenerative manifestations such as PD is poorly understood. Here, we investigated the effect of silencing Δ9 desaturase enzyme encoding fat-5 and fat-7 genes which are known to reduce fat content, on α-synuclein expression, neuronal morphology and dopamine-related behaviors in transgenic PD-like models of Caenorhabditis elegans (C. elegans). The silencing of the fat-5 and fat-7 genes rescued both degeneration of dopamine neurons and deficits in dopamine-dependent behaviors, including basal slowing and ethanol avoidance in worm models of PD. Similarly, silencing of these genes also decreased the formation of protein aggregates in a nematode model of PD expressing α-synuclein in the body wall muscles and rescued deficits in resistance to heat and osmotic stress. On the contrary, silencing of nhr-49 and tub-1 genes that are known to increase total fat content did not alter behavioral and pathological endpoints in the PD worm strains. Interestingly, the genetic manipulation of all four selected genes resulted in differential fat levels in the PD models without having significant effect on the lifespan, further indicating a complex fat homeostasis unique to neurodegenerative pathophysiology. Overall, we provide a comprehensive understanding of how Δ9 desaturase can alter PD-like pathology due to environmental exposures and proteotoxic stress, suggesting new avenues in deciphering the disease etiology and possible therapeutic targets.

XBP-1 Remodels Lipid Metabolism to Extend Longevity

SummaryThe endoplasmic reticulum unfolded protein response (UPRER) is a cellular stress response that maintains homeostasis within the secretory pathway, regulates glucose and lipid metabolism, and influences longevity. To ask whether this role in lifespan determination depends upon metabolic intermediaries, we metabotyped C. elegans expressing the active form of the UPRER transcription factor XBP-1, XBP-1s, and found many metabolic changes. These included reduced levels of triglycerides and increased levels of oleic acid (OA), a monounsaturated fatty acid associated with lifespan extension in C. elegans. Here, we show that constitutive XBP-1s expression increases the activity of lysosomal lipases and upregulates transcription of the Δ9 desaturase FAT-6, which is required for the full lifespan extension induced by XBP-1s. Dietary OA supplementation increases the lifespan of wild-type, but not xbp-1s-expressing animals and enhances proteostasis. These results suggest that modulation of lipid metabolism by XBP-1s contributes to its downstream effects on protein homeostasis and longevity.

Isolation of Gram‐positiveFirmibacteriaas major eicosapentaenoic acid producers from subtropical marine sediments

In this study, a total of 172 putative omega-3 producers were isolated from 28 sediment samples from the Arabian Gulf employing a selective isolation procedure using marine agar containing 0·1% triphenyl tetrazolium chloride (TTC). Out of these 172 isolates, 19 isolates produced eicosapentaenoic acid (EPA) as confirmed by Gas Chromatography-Mass Spectrometry (GC-MS). The EPA content of the isolated bacterial strain varied from 1·76 to 6·52% of total fatty acids. Among the 19 isolates of EPA producers, while 17 isolates harboured both pfaA gene and Δ6 desaturase gene, only five isolates harboured Δ5 desaturase gene. Two of the EPA positive strains harbour none of the three genes tested. The 16s RNA identification of these isolates revealed that except one, all the EPA producers were Gram-positive marine bacteria belonging to the phylum Firmicutes, family Bacillacea, genera Bacillus and Oceanobacillus. Halomonas pacifica was the only Gram-negative Gamma-Proteobacteria detected to produce EPA from this region. SIGNIFICANCE AND IMPACT OF THE STUDY: Recently, marine bacteria are considered as a promising source of polyunsaturated fatty acid (PUFA) over marine fishes and microalgae. PUFA producers reported from polar and deep-sea sources were restricted to five well-known marine genera under two distinct domains of bacteria such as proteobacteria (Shewanella, Colwellia, and Moritella) and cytophaga group (Flexibacter, Psychroflexus). This study revealed that subtropical marine environment could also be the source of PUFA producing bacteria, and they predominantly belonged to the class of Firmibacteria. This finding opens up new avenue for research to study the inherent mechanism and physiology of such organisms from this unique environment.

Accumulation of γ-linolenic acid and stearidonic acid in rapeseeds that heterologously express the Phytophthora citrophthora Δ6 desaturase gene

Most oilseeds contain polyunsaturated fatty acids, such as linoleic acid (LA) and α-linolenic acid (ALA) that are important essential fatty acids for human health. γ-Linolenic acid (GLA) and stearidonic acid (SDA), synthesized from LA and ALA by Δ6 desaturase, respectively, are mainly obtained from algae and fish. Given the benefits of GLA and SDA to human health, many researchers have investigated ways to produce these fatty acids in oilseed crops. Canola-type oilseed rape (Brassica napus), widely cultivated in temperate and microthermal zones, accumulates oleic acid to a relatively greater extent (60–70% oleic acid content) than do other oilseed crops. We transformed the Phytophthora citrophthora Δ6 desaturase (PcD6DES) gene under the control of seed-specific vicilin into canola-type oilseed rape ‘Youngsan’. D6DES products (GLA, SDA, and 18:2Δ6,9) accumulated in mature PcD6DES-transformed rapeseeds and leaves. D6DES products of PcD6DES rapeseeds were over 20% in T1 and more than 25% in T2. Expression levels of the PcD6DES gene and the content of D6DES products coincided with each other and were related to expression levels and fatty acid composition in leaves and developing seeds 15, 25, 35, and 45 days after flowering. Seed weights of PcD6DES rapeseeds were not lower than those of Youngsan. In this study, PcD6DES oilseed rapes accumulated GLA, SDA, and putative 18:2Δ6,9 content to a maximum of 25% in the seed oil. Results show that PcD6DES rapeseed oil can potentially be used as a health food to improve human health.

Stearoyl-ACP Δ9 Desaturase 6 and 8 (GhA-SAD6 and GhD-SAD8) Are Responsible for Biosynthesis of Palmitoleic Acid Specifically in Developing Endosperm of Upland Cotton Seeds.

Palmitoleic acid (16:1Δ9) is one kind of ω-7 fatty acids (ω-7 FAs) widely used in food, nutraceutical and industry. However, such high-valued ω-7 FA only has a trace level in mature seeds of cotton and other common oil crops. We found that palmitoleic acid (>10.58 Mol%) was specially enriched in developing cotton endosperm which is disappeared in its mature seed. The present study was conducted to investigate the mechanism underlying high accumulation of palmitoleic acid in developing endosperm but not in embryo of upland cotton (Gossypium hirsutum L.) seed. Of 17 stearoyl-ACP Δ9 desaturases (SAD) gene family members identified in upland cotton, six GhSADs may specifically work in the desaturation of palmitic acid (16:0-ACP) to produce palmitoleic acid (16:1Δ9-ACP), which were revealed by examining the key amino acids in the catalytic center and their cis-elements. Gene expression analysis showed that spatial patterns of these GhSADs were different in developing ovules, with GhA-SAD6 and GhD-SAD8 preferentially expressed in developing endosperms. Functional analysis by transient expression in Nicotiana benthamiana leaves and genetic complementary assay using yeast mutant BY4389 strain unable to synthesize unsaturated fatty acids demonstrated that GhA-SAD6 and GhD-SAD8 have strong substrate specificity for 16:0-ACP. In contrast, GhA-SAD5 and GhA-SAD7 exhibited high specific activity on 18:0-ACP. Taken together, these data evidence that GhA-SAD6 and GhD-SAD8 are responsible for making palmitoleic acid in developing cotton endosperms, and provide endogenous gene targets for genetic modification to enrich ω-7 FAs in cotton seed oil required for sustainable production of functionality-valued products.

Balanced omega-3 and -6 vegetable oil of Amazonian sacha inchi act as LC-PUFA precursors in rainbow trout juveniles: Effects on growth and fatty acid biosynthesis

The aim of this study was to evaluate the metabolic responses to fish oil (FO) replacement with different inclusion levels of sacha inchi (SI) oil (0FO, 40FO, 60FO, and 100FO, representing the % of FO inclusion) in the diet of O. mykiss juveniles. Growth rates, energetic substrates, enzymatic pathways involved in the synthesis of fatty acids (FAs), and the FA profile in the muscle, liver, and plasma were investigated. Fish (n = 320) were distributed into eight tanks (four groups in duplicate) and fed with the four experimental diets for 90 days. Biometric data were obtained every 30 days, and after 45 and 90 days, ten fish from each experimental group were sampled. Somatic indices, biochemical parameters, and the mRNA expression levels of three genes involved in FA elongation and desaturation, Δ6 desaturase (Δ6), elongase 2 (elovl2), and elongase 5 (elovl5), were analyzed. The substitution of FO with SI oil did not alter growth rates, regardless of inclusion level and time point. Fish fed with SI oil had increased hepatosomatic and viscerosomatic indexes (HSI and VSI) after 45 days, while the VSI increased after 90 days in the 0FO group. Fish fed exclusively with SI oil had increased total protein content in their muscle tissue after 90 days. The 40FO, 60FO, and 100FO groups had similar ARA and DHA percentages in their livers at 45 days, which is reflective of rainbow trout being able to synthesize LC-PUFA from dietary C18PUFAs. Although replacing FO with SI oil resulted in higher expression levels of the desaturase and elongase genes at both time points, this increase was insufficient to maintain the high levels of muscular LC-PUFA in the 40FO and 0FO groups. The percentages of C18PUFAs in the muscle of fish fed with the 0FO diet increased, while the percentage of n-3 LC-PUFA and the n-3:n-6 ratio decreased. Although the 0FO group exhibited higher transcription levels for the enzymes involved in LC-PUFA biosynthesis, the most promising results in terms of muscle FA profile were observed in the 60FO group. This finding indicates that the inclusion of 40% of SI into the diet of O. mykiss did not affect the percentage of ARA, EPA, and DHA. Taken together, these data suggest that SI represents a viable, promising replacement for FO in the feed for salmonids and potentially other aquacultured fish species as well.

Modulating Heterologous Pathways and Optimizing Culture Conditions for Biosynthesis of trans-10, cis-12 Conjugated Linoleic Acid in Yarrowia lipolytica.

A novel recombinant strain has been constructed for converting glycerol into a specific conjugated linoleic acid isomer (trans-10, cis-12 CLA) using Yarrowia lipolytica as host. The lipid accumulation pathway was modified for increasing lipid content. Overexpression of the diacylglycerol transferase (DGA1) gene improved the intracellular lipid yield by approximately 45% as compared to the original strain. The corresponding intracellular lipid yield of recombinant strain WXYL037 reached 52.2% of the cell dry weight. In combination with integration of Δ12 desaturase from Mortierella alpina (MA12D) and DGA1, the linoleic acid (LA) production content reached 0.88 g/L, which was 2-fold that of the original strain. Furthermore, with overexpressed DGA1, MA12D and Propionibacterium acnes isomerase (PAI), the titer of trans-10, cis-12 CLA in WXYL037 reached 110.6 mg/L after 72 h of shake flask culture, representing a 201.8% improvement when compared with that attained in the WXYL030 strain, which manifested overexpressed PAI. With optimal medium, the maximum CLA content and lipid yield of Y. lipolytica Po1g were 132.6 mg/L and 2.58 g/L, respectively. This is the first report of the production of trans-10, cis-12 CLA by the oleaginous yeast Y. lipolytica using glycerol as the sole carbon source through expression of DGA1 combined with MA12D and PAI.

Sources of microbial oils with emphasis to Mortierella (Umbelopsis) isabellina fungus.

The last years a constantly rising number of publications have appeared in the literature in relation to the production of oils and fats deriving from microbial sources (the "single cell oils"-SCOs). SCOs can be used as precursors for the synthesis of lipid-based biofuels or employed as substitutes of expensive oils rarely found in the plant or animal kingdom. In the present review-article, aspects concerning SCOs (economics, biochemistry, substrates, technology, scale-up), with emphasis on the potential of Mortierella isabellina were presented. Fats and hydrophilic substrates have been used as carbon sources for cultivating Zygomycetes. Among them, wild-type M. isabellina strains have been reported as excellent SCO-producers, with conversion yields on sugar consumed and lipid in DCW values reported comparable to the maximum ones achieved for genetically engineered SCO-producing strains. Lipids produced on glucose contain γ-linolenic acid (GLA), a polyunsaturated fatty acid (PUFA) of high dietary and pharmaceutical importance, though in low concentrations. Nevertheless, due to their abundance in oleic acid, these lipids are perfect precursors for the synthesis of 2nd generation biodiesel, while GLA can be recovered and directed to other usages. Genetic engineering focusing on over-expression of Δ6 and Δ12 desaturases and of C16 elongase may improve the fatty acid composition (viz. increasing the concentration of GLA or other nutritionally important PUFAs) of these lipids.

De novo transcriptomic analysis of gonad of Strongylocentrotus nudus and gene discovery for biosynthesis of polyunsaturated fatty acids.

Strongylocentrotus nudus is an important cultured sea urchin species in north China, because its gonad is rich in unsaturated fatty acids, particularly long polyunsaturated fatty acids (LC-PUFAs). These PUFAs play pleiotropic and crucial roles in a wide range of biological process. However, the genes contributing to biosynthesis PUFAs have not been elucidated yet, and the molecular mechanism relative to the difference in PUFA composition between male and female gonad as been revealed but the corresponding has not been understood. In this paper, solexa sequencing based transcriptomic approach was used to identify and characterize the key genes relative to PUFA synthesis and further conducted different expressed genes between male and female gonad. A total of 130,124 transcripts and 189330 unigenes were de novo assembled from 64.32Gb data. Next, these unigenes were subjected to functional annotation by mapping to six public databases, and this process revealed a lot of genes involving in lipid metabolism. In addition, three types of fatty acids front-end desaturase and three species of very long fatty acids elongase were identified and the pathway for PUFA biosynthesis was hypothesized. Last, comparative analysis revealed the higher expression level of Δ5 desaturase, Δ6 desaturase, ELOVL-4, -6 and -7 in male gonad compared with female. This results could plausible explain the differ in composition of PUFAs between male and female gonad of sea urchin.

A Δ9 desaturase (SlitDes11) is associated with the biosynthesis of ester sex pheromone components in Spodoptera litura

Sex pheromone biosynthesis in moths relies on the activity of multiple enzymes, including Δ9 desaturase, which plays an important role in catalyzing desaturation at the Δ9 position of the carbon chain. However, the physiological function of moth Δ9 desaturase has not been elucidated in vivo. In this study, we used the CRISPR/Cas9 system to knockout the Δ9 desaturase gene (SlitDes11) of Spodoptera litura to analyze its role in sex pheromone biosynthesis. First, through the direct injection of SlitDes11-single guide RNA (sgRNA)/Cas9 messenger RNA into newly laid eggs, gene editing was induced in around 30% of eggs 24 h after injection and was induced in 20.8% of the resulting adult moths. Second, using a sibling-crossing strategy, insects with mutant SlitDes11 (bearing a premature stop codon) were selected, and homozygous mutants were obtained in the G5 generation. Third, pheromone gland extracts of adult female homozygous SlitDes11 mutants were analyzed using Gas chromatography (GC). The results showed that titers of all three ester sex pheromone components; Z9, E11-14:Ac, Z9,E12-14:Ac, and Z9-14:Ac; were reduced by 62.40%, 78.50%, and 72.50%, respectively. This study provides the first direct evidence for the role of SlitDes11 in sex pheromone biosynthesis in S. litura, and indicates the gene could be as potential target to disrupt sexual communication in S. litura for developing a new pollution-free insecticide.

Pseudomonas putida Δ9-fatty acid desaturase: Gene cloning, expression, and function in the cationic surfactants stress.

Pseudomonas putida counteract the fluidizing effect of cationic surfactants decreasing the content of membrane unsaturated fatty acid (UFA). A Δ9-fatty acid desaturase gene (desA) from P. putida was isolated, cloned, and successfully expressed in Escherichia coli, a Δ9 desaturase deficient organism. desA consists of 1185 bp and codes for 394 amino acids. The deduced amino acid sequence reveals three histidine clusters and a hydropathy profile, typical of membrane-bound desaturases. Validating desA expression in E. coli cells, the amount of palmitoleic acid increased from 2.05 to 7.36%, with the concomitant increase in membrane fluidity (fluorescence polarization value decrease from 0.13 ± 0.03 to 0.09 ± 0.02). Also, when DesA activity was assayed in vivo, the percentage of UFA obtained from exogenous palmitic acid [1-14 C] increased 10-fold. In contrast, when cells expressing desA were exposed 15 min at sublethal concentration of cationic surfactants, the amount of UFA was 82% lower than that detected in cells non-exposed. Thus, the decrease in UFA content to counteract the fluidizing effect of cationic surfactants can be correlated with reduction of DesA activity.

Long-Chain Polyunsaturated Fatty Acids in the Green Microalga Lobosphaera incisa Contribute to Tolerance to Abiotic Stresses.

Lobosphaera incisa is a green microalga that accumulates high levels of the valuable omega-6 long-chain polyunsaturated fatty acids (LC-PUFA) arachidonic acid (ARA, 20:4n-6) in triacylglycerols (TAG) under nitrogen (N) starvation. LC-PUFA accumulation is a rare trait in photosynthetic microalgae with insufficiently understood physiological significance. In this study, RNAi was attempted, for the first time in L. incisa, to produce knockdown lines for the Δ5 desaturase gene. Two lines, termed modified lines, which were isolated during screening for transgenic events, demonstrated alterations in their LC-PUFA profile, ARA-biosynthesis gene expression and lipid class distribution. In line M5-78, which appeared to carry a mutation in the Δ6 elongase gene, LC-PUFA were substituted by 18:3n-6 in all glycerolipids. Line M2-35, for which the exact genetic background has not been established, displayed a dramatic reduction in 20:4n-6, concomitant with an augmented proportion of 18:1n-9, in particular in the extraplastidial membrane lipids and TAG. The physiological responses of the modified lines to stressful conditions were compared with the wild type and the Δ5 desaturase mutant. In the N-replete cells of modified lines, the frequency of lipid droplets was reduced, while a number of starch grains increased, suggesting altered partitioning of assimilated carbon into reserve products. Furthermore, both lines exhibited reduced ability to accumulate TAG under N deprivation and recover from N starvation. Both lines demonstrated lower photosynthetic pigment contents, impairments in photosynthesis under a range of stressful conditions, and less efficient functioning of photoprotection under optimal conditions. Possible implications of fatty acids modifications in the stress response of L. incisa are addressed.

Conditional deletion of Des1 in the mouse retina does not impair the visual cycle in cones.

Cone photoreceptors are essential for vision under moderate to high illuminance and allow color discrimination. Their fast dark adaptation rate and resistance to saturation are believed to depend in part on an intraretinal visual cycle that supplies 11-cis-retinaldehyde to cone opsins. Candidate enzymes of this pathway have been reported, but their physiologic contribution to cone photoresponses remains unknown. Here, we evaluate the role of a candidate retinol isomerase of this pathway, sphingolipid δ4 desaturase 1 (Des1). Single-cell RNA sequencing analysis revealed Des1 expression not only in Müller glia but also throughout the retina and in the retinal pigment epithelium. We assessed cone functional dependence on Müller cell–expressed Des1 through a conditional knockout approach. Floxed Des1 mice, on a guanine nucleotide-binding protein subunit α transducin 1 knockout (Gnat1−/−) background to allow isolated recording of cone-driven photoresponses, were bred with platelet-derived growth factor receptor α (Pdgfrα)-Cre mice to delete Des1 in Müller cells. Conditional knockout of Des1 expression, as shown by tissue-selective Des1 gene recombination and reduced Des1 catalytic activity, caused no gross changes in the retinal structure and had no effect on cone sensitivity or dark adaptation but did slightly accelerate the rate of cone phototransduction termination. These results indicate that Des1 expression in Müller cells is not required for cone visual pigment regeneration in the mouse.—Kiser, P. D., Kolesnikov, A.V., Kiser, J. Z., Dong, Z., Chaurasia, B., Wang, L., Summers, S. A., Hoang, T., Blackshaw, S., Peachey, N. S., Kefalov, V. J., Palczewski, K. Conditional deletion of Des1 in the mouse retina does not impair the visual cycle in cones.

Genome-wide identification and expression of eight fatty acid desaturase genes, and the fatty acid profile, in the marine rotifer Brachionus koreanus fed the alga Tetraselmis suecica

Fatty acid desaturases catalyze the formation of a C=C double bond from a C–C single bond in fatty acids, leading to diversification of the fatty acid pool. In this study, we identified the fatty acid desaturase genes in the monogonont marine rotifer Brachionus koreanus, measured the messenger RNA (mRNA) expression, and quantified fatty acid composition under feeding with the alga Tetraselmis suecica. Eight fatty acid desaturase genes were identified and classified by phylogenetic analysis. Fatty acid desaturases in the rotifer B. koreanus were categorized into two Δ4 desaturases, five Δ5/6 desaturases, and one Δ9 desaturase. mRNA expression of B. koreanus under the alga T. suecica-fed condition clearly indicated that transcriptional levels of desaturase genes were increased compared to the non-T. suecica-fed group. Also, the distribution of fatty acids of B. koreanus fed T. suecica was more balanced compared to that of T. suecica. These results provide a better understanding of the role of fatty acid desaturases in B. koreanus fed the alga T. suecica.