Fatty Acid Elongase Research Articles

Metabolic Profiling Reveals an Abnormal Pattern of Serum Fatty Acids in MRL/lpr Mice Under Treatment With Prednisone.

Glucocorticoids (GCs) are commonly used to treat systemic lupus erythematosus (SLE). Unfortunately, excessive GCs can induce many side effects associated with disordered fatty acid (FA) metabolism. Although an increased level of total FA has been found after GCs treatment, it is not clear whether all FA species increased or only certain FA species were altered. A gas chromatography–mass spectrometry-based FA profiling approach was performed to reveal the alterations of FA species in SLE model mice (MRL/lpr) after treatment with 5 mg/kg of prednisone. The study showed a distinct FA profile in MRL/lpr mice compared to the controls, mainly manifested by elevated polyunsaturated FAs (arachidonate, docosahexaenoate, etc.), which are related to the inflammatory state; and altered (product FA/precursor FA) ratios representing the estimated activities of FA desaturase and elongase (higher activities of multiple elongases, △4 desaturase, △5 desaturase, △6 desaturase, and lower activity of △8 desaturase). Treatment with 5 mg/kg of prednisone decreased the total level of n-6 polyunsaturated FA in MRL/lpr mice; in particular, the level of arachidonate and estimated activity of △5 desaturase were reduced to the control level. Moreover, prednisone induced additional perturbations in FAs, including not only saturated FAs, but also monounsaturated FAs and n-3 polyunsaturated FAs, indicating that there was a strong effect of prednisone on FA metabolism. These results may be valuable for further studies of the side effects of GCs treatment.

Hepatocyte ELOVL Fatty Acid Elongase 6 Determines Ceramide Acyl-Chain Length and Hepatic Insulin Sensitivity in Mice.

Dysfunctional hepatic lipid metabolism is a cause of nonalcoholic fatty liver disease (NAFLD), the most common chronic liver disorder worldwide, and is closely associated with insulin resistance and type 2 diabetes. ELOVL fatty acid elongase 6 (Elovl6) is responsible for converting C16 saturated and monounsaturated fatty acids (FAs) into C18 species. We have previously shown that Elovl6 contributes to obesity-induced insulin resistance by modifying hepatic C16/C18-related FA composition. To define the precise molecular mechanism by which hepatic Elovl6 affects energy homeostasis and metabolic disease, we generated liver-specific Elovl6 knockout (LKO) mice. Unexpectedly, LKO mice were not protected from high-fat diet-induced insulin resistance. Instead, LKO mice exhibited higher insulin sensitivity than controls when consuming a high-sucrose diet (HSD), which induces lipogenesis. Hepatic patatin-like phospholipase domain-containing protein 3 (Pnpla3) expression was down-regulated in LKO mice, and adenoviral Pnpla3 restoration reversed the enhancement in insulin sensitivity in HSD-fed LKO mice. Lipidomic analyses showed that the hepatic ceramide(d18:1/18:0) content was lower in LKO mice, which may explain the effect on insulin sensitivity. Ceramide(d18:1/18:0) enhances protein phosphatase 2A (PP2A) activity by interfering with the binding of PP2A to inhibitor 2 of PP2A, leading to Akt dephosphorylation. Its production involves the formation of an Elovl6-ceramide synthase 4 (CerS4) complex in the endoplasmic reticulum and a Pnpla3-CerS4 complex on lipid droplets. Consistent with this, liver-specific Elovl6 deletion in ob/ob mice reduced both hepatic ceramide(d18:1/18:0) and PP2A activity and ameliorated insulin resistance. Our study demonstrates the key role of hepatic Elovl6 in the regulation of the acyl-chain composition of ceramide and that C18:0-ceramide is a potent regulator of hepatic insulin signaling linked to Pnpla3-mediated NAFLD.

RETRACTED: Impaired lipid metabolism by age-dependent DNA methylation alterations accelerates aging

Epigenetic alterations and metabolic dysfunction are two hallmarks of aging. However, the mechanism of how their interaction regulates aging, particularly in mammals, remains largely unknown. Here we show ELOVL fatty acid elongase 2 (Elovl2), a gene whose epigenetic alterations are most highly correlated with age prediction, contributes to aging by regulating lipid metabolism. Impaired Elovl2 function disturbs lipid synthesis with increased endoplasmic reticulum stress and mitochondrial dysfunction, leading to key accelerated aging phenotypes. Restoration of mitochondrial activity can rescue age-related macular degeneration (AMD) phenotypes induced by Elovl2 deficiency in human retinal pigmental epithelial (RPE) cells. We revealed an epigenetic-metabolism axis contributing to aging and potentially to antiaging therapy.

Identification and characterization of two fatty acid elongases in Lipomyces starkeyi.

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.

Dysregulation of microRNA-125a contributes to obesity-associated insulin resistance and dysregulates lipid metabolism in mice

Obesity is associated with an increased risk of developing insulin resistance (IR) and type 2 diabetes (T2D). A diverse group of factors including miRNA has been implicated in the pathogenesis of these two metabolic conditions, although underlying molecular mechanisms involved are not well defined. Here, we provide evidence that hepatic miR-125a levels are diminished in both genetic as well as dietary mouse models of obesity. Overexpression of miR-125a enhanced insulin signaling and attenuated cellular lipid accumulation in HepG2 cells and Hepa1–6 cells. Likewise, treatment of mice with ago-miR-125a increased insulin sensitivity, similar to overexpression of miR-125a, whereas treatment of mice with antago-miR-125a blunted the insulin sensitivity. Furthermore, overexpression of miR-125a in mice previously fed a high-fat diet (HFD), significantly improved insulin sensitivity, and attenuated obesity-linked hepatic steatosis and hepatocyte lipid accumulation. In addition, we show that ELOVL fatty acid elongase 6 (Elovl6) is a direct target of miR-125a, and participates in miR-125a mediated regulation of insulin sensitivity and lipid metabolism. These data led us to conclude that dysregulated miR-125a expression augments the development of obesity-induced IR and that miR-125a might serve as a therapeutic target for the development of new drug(s) in the clinical management of metabolic diseases.

Genome-Wide Association Study for Serum Omega-3 and Omega-6 Polyunsaturated Fatty Acids: Exploratory Analysis of the Sex-Specific Effects and Dietary Modulation in Mediterranean Subjects with Metabolic Syndrome.

Many early studies presented beneficial effects of polyunsaturated fatty acids (PUFA) on cardiovascular risk factors and disease. However, results from recent meta-analyses indicate that this effect would be very low or nil. One of the factors that may contribute to the inconsistency of the results is that, in most studies, genetic factors have not been taken into consideration. It is known that fatty acid desaturase (FADS) gene cluster in chromosome 11 is a very important determinant of plasma PUFA, and that the prevalence of the single nucleotide polymorphisms (SNPs) varies greatly between populations and may constitute a bias in meta-analyses. Previous genome-wide association studies (GWAS) have been carried out in other populations and none of them have investigated sex and Mediterranean dietary pattern interactions at the genome-wide level. Our aims were to undertake a GWAS to discover the genes most associated with serum PUFA concentrations (omega-3, omega-6, and some fatty acids) in a scarcely studied Mediterranean population with metabolic syndrome, and to explore sex and adherence to Mediterranean diet (MedDiet) interactions at the genome-wide level. Serum PUFA were determined by NMR spectroscopy. We found strong robust associations between various SNPs in the FADS cluster and omega-3 concentrations (top-ranked in the adjusted model: FADS1-rs174547, p = 3.34 × 10−14; FADS1-rs174550, p = 5.35 × 10−14; FADS2-rs1535, p = 5.85 × 10−14; FADS1-rs174546, p = 6.72 × 10−14; FADS2-rs174546, p = 9.75 × 10−14; FADS2-rs174576, p = 1.17 × 10−13; FADS2-rs174577, p = 1.12 × 10−12, among others). We also detected a genome-wide significant association with other genes in chromosome 11: MYRF (myelin regulatory factor)-rs174535, p = 1.49 × 10−12; TMEM258 (transmembrane protein 258)-rs102275, p = 2.43 × 10−12; FEN1 (flap structure-specific endonuclease 1)-rs174538, p = 1.96 × 10−11). Similar genome-wide statistically significant results were found for docosahexaenoic fatty acid (DHA). However, no such associations were detected for omega-6 PUFAs or linoleic acid (LA). For total PUFA, we observed a consistent gene*sex interaction with the DNTTIP2 (deoxynucleotidyl transferase terminal interacting protein 2)-rs3747965 p = 1.36 × 10−8. For adherence to MedDiet, we obtained a relevant interaction with the ME1 (malic enzyme 1) gene (a gene strongly regulated by fat) in determining serum omega-3. The top-ranked SNP for this interaction was ME1-rs3798890 (p = 2.15 × 10−7). In the regional-wide association study, specifically focused on the FADS1/FASD2/FADS3 and ELOVL (fatty acid elongase) 2/ELOVL 5 regions, we detected several statistically significant associations at p < 0.05. In conclusion, our results confirm a robust role of the FADS cluster on serum PUFA in this population, but the associations vary depending on the PUFA. Moreover, the detection of some sex and diet interactions underlines the need for these associations/interactions to be studied in all specific populations so as to better understand the complex metabolism of PUFA.

Skin permeability barrier formation by the ichthyosis-causative gene FATP4 through formation of the barrier lipid ω-O-acylceramide

The epidermis-specific lipid acylceramide plays a pivotal role in the formation of the permeability barrier in the skin; abrogation of its synthesis causes the skin disorder ichthyosis. However, the acylceramide synthetic pathway has not yet been fully elucidated: Namely, the acyl-CoA synthetase (ACS) involved in this pathway remains to be identified. Here, we hypothesized it to be encoded by FATP4/ACSVL4, the causative gene of ichthyosis prematurity syndrome (IPS). In vitro experiments revealed that FATP4 exhibits ACS activity toward an ω-hydroxy fatty acid (FA), an intermediate of the acylceramide synthetic pathway. Fatp4 knockout (KO) mice exhibited severe skin barrier dysfunction and morphological abnormalities in the epidermis. The total amount of acylceramide in Fatp4 KO mice was reduced to ∼10% of wild-type mice. Decreased levels and shortening of chain lengths were observed in the saturated, nonacylated ceramides. FA levels were not decreased in the epidermis of Fatp4 KO mice. The expression levels of the FA elongase Elovl1 were reduced in Fatp4 KO epidermis, partly accounting for the reduction and shortening of saturated, nonacylated ceramides. A decrease in acylceramide levels was also observed in human keratinocytes with FATP4 knockdown. From these results, we conclude that skin barrier dysfunction observed in IPS patients and Fatp4 KO mice is caused mainly by reduced acylceramide production. Our findings further elucidate the molecular mechanism governing acylceramide synthesis and IPS pathology.

The NASA Twins Study: The Effect of One Year in Space on Long-Chain Fatty Acid Desaturases and Elongases

Background: At present, there is no clear understanding of the effect of long-duration spaceflight on the major enzymes that govern the metabolism of omega-6 and omega-3 fatty acids. To address this gap in knowledge, we used data from the NASA Twins Study, which includes a multiscale omics investigation of the changes that occurred during a year-long (340 days) human spaceflight. Embedded within the NASA Twins data are specific analytes associated with fatty acid metabolism. Objectives: To examine the long-chain fatty acid desaturases and elongases in a single human during 1 year in space. Method: One male twin was on board the International Space Station (ISS) for 1 year, while his monozygotic twin served as a genetically matched ground control. Longitudinal assessments included the genome, epige­nome, transcriptome, proteome, metabolome, microbiome, and immunome during the mission, as well as 6 months before and after. The gene-specific fatty acid desaturase and elongase transcriptome data (FADS1, FADS2, ELOVL2, and ELOVL5) were extracted from untargeted RNA-seq measurements derived from white blood cell fractions. Results: Most data from the elongases and desaturases exhibited relatively similar expression profiles (R² >0.6) over time for the CD8, CD19, and lymphocyte-depleted (LD) cell fractions, indicating overall conservation of function within and between the subjects. Both cell-type and temporal specificity was observed in some cases, and some differences were also apparent between the polyadenylated (polyA) fraction of processed RNAs versus the ribodepleted (ribo–) fraction. The flight subject showed a stronger enrichment of the fatty acid metabolic process pathway across almost all cell types (columns, CD4, CD8, CPT, and LD), most especially in the ribodepleted fraction of RNA, but also with the polyA+ fraction of RNA. Gene set enrichment analysis (GSEA) measures across three related fatty acid metabolism pathways showed a differential between the ground and the flight subject. Conclusions: There appears to be no persistent alteration of desaturase and elongase gene expression associated with 1 year in space. However, these data provide evidence that cellular lipid metabolism can be responsive and dynamic to spaceflight, even though it appears cell-type and context specific, most notably in terms of the fraction of RNA measured and the collection protocols. These results also provide new evidence of mid-flight spikes in expression of selected genes, which may indicate transient responses to specific insults during spaceflight.

Expression of enzymes involved in polyunsaturated fatty acid synthesis in the stallion testis and epididymis.

The aim of the present study was to characterise key enzymes involved in polyunsaturated fatty acid (PUFA) synthesis in the testis and epididymis collected from 2-year-old healthy warmblood stallions (n=10). The mRNA expression of fatty acid synthase, the Δ9-, Δ6-, Δ5- and Δ4-desaturases and elongases 6, 5 and 2 (encoded by the fatty acid synthase (FASN), the stearoyl-CoA desaturase (SCD), the fatty acid desaturase 2 (FADS2), the fatty acid desaturase 1 (FADS1), the delta 4-desaturase, sphingolipid 1 (DEGS1), ELOVL fatty acid elongase 6(ELOVL6), ELOVL fatty acid elongase 5 (ELOVL5), ELOVL fatty acid elongase 2 (ELOVL2) genes respectively) was determined in equine testis and epididymis. All enzymes were present in testicular tissue and along the epididymis, but mRNA expression differed among localisations. The protein localisation of FADS1, FADS2 and ELOVL5 was determined by immunohistochemistry. In the testes, FADS1 was expressed in the germinal cells and ELOVL5 was expressed in germinal and Leydig cells; FADS2 was not detected. In the epididymis, FADS1 and FADS2 were expressed in the principal and basal cells, whereas ELOVL5 was found only in the principal cells of the caput. All three enzymes were present in epididymal vesicles secreted by an apocrine mechanism. These results suggest active PUFA metabolism during spermatogenesis and epididymal sperm maturation in stallions.

Leucosceptroid B from glandular trichomes of Leucosceptrum canum reduces fat accumulation in Caenorhabditis elegans through suppressing unsaturated fatty acid biosynthesis

Obesity that is highly associated with numerous metabolic diseases has become a global health issue nowdays. Plant sesterterpenoids are an important group of natural products with great potential; thus, their bioactivities deserve extensive exploration. RNA-seq analysis indicated that leucosceptroid B, a sesterterpenoid previously discovered from the glandular trichomes of Leucosceptrum canum, significantly regulated the expression of 10 genes involved in lipid metabolism in Caenorhabditis elegans. Furthermore, leucosceptroid B was found to reduce fat storage, and downregulate the expression of two stearoyl-CoA desaturase (SCD) genes fat-6 and fat-7, and a fatty acid elongase gene elo-2 in wild-type C. elegans. In addition, leucosceptroid B significantly decreased fat accumulation in both fat-6 and fat-7 mutant worms but did not affect the fat storage of fat-6; fat-7 double mutant. These findings indicated that leucosceptroid B reduced fat storage depending on the downregulated expression of fat-6, fat-7 and elo-2 and thereby inhibiting the biosynthesis of the corresponding unsaturated fatty acid. These findings provide new insights into the development and utilization of plant sesterterpenoids as potential antilipemic agents.

MYCN and PRC1 cooperatively repress docosahexaenoic acid synthesis in neuroblastoma via ELOVL2

BackgroundThe MYCN amplification is a defining hallmark of high-risk neuroblastoma. Due to irregular oncogenes orchestration, tumor cells exhibit distinct fatty acid metabolic features from non-tumor cells. However, the function of MYCN in neuroblastoma fatty acid metabolism reprogramming remains unknown.MethodsGas Chromatography-Mass Spectrometer (GC-MS) was used to find the potential target fatty acid metabolites of MYCN. Real-time PCR (RT-PCR) and clinical bioinformatics analysis was used to find the related target genes. The function of the identified target gene ELOVL2 on cell growth was detected through CCK-8 assay, Soft agar colony formation assay, flow Cytometry assay and mouse xenograft. Chromatin immunoprecipitation (ChIP) and Immunoprecipitation-Mass Spectrometer (IP-MS) further identified the target gene and the co-repressor of MYCN.ResultsThe fatty acid profile of MYCN-depleted neuroblastoma cells identified docosahexaenoic acid (DHA), an omega-3 polyunsaturated fatty acid with anti-tumor activity, significantly increased after MYCN depletion. Compared with MYCN single-copy neuroblastoma cells, DHA level was significantly lower in MYCN-amplified neuroblastoma cells. RT-PCR and clinical bioinformatics analysis discovered that MYCN interfered DHA accumulation via ELOVL fatty acid elongase 2 (ELOVL2) which is a rate-limiting enzyme of cellular DHA synthesis. Enforced ELOVL2 expression in MYCN-amplified neuroblastoma cells led to decreased cell growth and counteracted the growth-promoting effect of MYCN overexpression both in vitro and vivo. ELOVL2 Knockdown showed the opposite effect in MYCN single-copy neuroblastoma cells. In primary neuroblastoma, high ELOVL2 transcription correlated with favorable clinical tumor biology and patient survival. The mechanism of MYCN-mediated ELOVL2 inhibition contributed to epigenetic regulation. MYCN recruited PRC1 (Polycomb repressive complex 1), catalysed H2AK119ub (histone 2A lysine 119 monoubiquitination) and inhibited subsequent ELOVL2 transcription.ConclusionsThe tumor suppressive properties of DHA and ELOVL2 are repressed by the MYCN and PRC1 jointly, which suggests a new epigenetic mechanism of MYCN-mediated fatty acid regulation and indicates PRC1 inhibition as a potential novel strategy to activate ELOVL2 suppressive functions.

Reduced chain length in myelin sphingolipids and poorer motor coordination in mice deficient in the fatty acid elongase Elovl1.

Very‐long‐chain fatty acids, with a chain length of >C20, are abundant in myelin sphingolipids. Recently, a de novo mutation in the ELOVL1 gene, which encodes fatty acid elongase, was identified in patients with neurocutaneous disorders involving skin ichthyosis and multiple neurological abnormalities, including hypomyelination, spastic paraplegia, and high‐frequency deafness. However, the consequences of ELOVL1 deficiency for lipid composition and detailed pathological changes in the brain remain unclear. Here, we analyzed Elovl1 mutant mice as a model of human ELOVL1 deficiency. The mice exhibited a decreased postnatal survival rate, and some died of startle epilepsy. The acyl chain length of sphingolipids such as galactosylceramides, sulfatides, sphingomyelins, and ceramides in the brains of these mice was markedly shortened. Moreover, the mice exhibited reduced levels of galactosylceramides, which are important for myelin formation and stability. Electron microscope analysis of the corpus callosum in Elovl1 mutant mice revealed modest hypomyelination, especially in large‐diameter axons. Furthermore, behavioral testing of the mice revealed deficits such as poorer motor coordination and reduced acoustic startle response to high‐intensity stimulus. These findings provide clues to the molecular mechanism of the neurological symptoms of patients with the ELOVL1 mutation.

Agrobacterium-Mediated Transient Gene Expression in Developing Ricinus communis Seeds: A First Step in Making the Castor Oil Plant a Chemical Biofactory.

The castor oil plant represents a promising platform to produce oils with industrial applications. However, its use in biotechnology is limited by the absence of a well-established procedure to transform it, and a poor understanding of gene regulation and promoter use in this species. As such, a method has been developed to express proteins or hairpin-RNA in this plant, a method based on the direct injection of Agrobacterium into the developing endosperm of castor oil fruit, enabling different constructs and promoters to be tested. This method produces a high rate of transformation and a good proportion of viable seeds that express reporter genes for up to 20 days after infiltration (DAI). Gene expression under the control of different promoters was tested by quantitative real-time polymerase chain reaction and by directly assaying the activity of the galactouronidase reporter gene, which proved to be strongest when driven by the glycinin promoter. Constructs expressing a fatty acid elongase from Lesquerella fendleri were tested, the expression of which provoked an important increase in the lesquerolic acid in the castor oil endosperm at 5 and 10 DAI, although this fatty acid did not accumulate significantly in the final mature seeds. The nature of this response could reflect the poor availability of substrates for this enzyme. In the light of this data, the potential of this technique to test promoters and different constructs in castor oil plants and other oilseeds is discussed.

Human Cytomegalovirus pUL37x1 Is Important for Remodeling of Host Lipid Metabolism.

Human cytomegalovirus (HCMV) replication requires host metabolism. Infection alters the activity in multiple metabolic pathways, including increasing fatty acid elongation and lipid synthesis. The virus-host interactions regulating the metabolic changes associated with replication are essential for infection. While multiple host factors, including kinases and transcription factors, important for metabolic changes that occur following HCMV infection have been identified, little is known about the viral factors required to alter metabolism. In this study, we tested the hypothesis that pUL37x1 is important for the metabolic remodeling that is necessary for HCMV replication using a combination of metabolomics, lipidomics, and metabolic tracers to measure fatty acid elongation. We observed that fibroblast cells infected with wild-type (WT) HCMV had levels of metabolites similar to those in cells infected with a mutant virus lacking the UL37x1 gene, subUL37x1. However, we found that relative to WT-infected cells, subUL37x1-infected cells had reduced levels of two host proteins that were previously demonstrated to be important for lipid metabolism during HCMV infection: fatty acid elongase 7 (ELOVL7) and the endoplasmic reticulum (ER) stress-related kinase PERK. Moreover, we observed that HCMV infection results in an increase in phospholipids with very-long-chain fatty acid tails (PL-VLCFAs) that contain 26 or more carbons in one of their two tails. The levels of many PL-VLCFAs were lower in subUL37x1-infected cells than in WT-infected cells. Overall, we conclude that although pUL37x1 is not necessary for network-wide metabolic changes associated with HCMV infection, it is important for the remodeling of a subset of metabolic changes that occur during infection.IMPORTANCE Human cytomegalovirus (HCMV) is a common pathogen that asymptomatically infects most people and establishes a lifelong infection. However, HCMV can cause end-organ disease that results in death in the immunosuppressed and is a leading cause of birth defects. HCMV infection depends on host metabolism, including lipid metabolism. However, the viral mechanisms for remodeling of metabolism are poorly understood. In this study, we demonstrate that the viral UL37x1 protein (pUL37x1) is important for infection-associated increases in lipid metabolism, including fatty acid elongation to produce very-long-chain fatty acids (VLCFAs). Furthermore, we found that HCMV infection results in a significant increase in phospholipids, particularly those with VLCFA tails (PL-VLCFAs). We found that pUL37x1 was important for the high levels of fatty acid elongation and PL-VLCFA accumulation that occur in HCMV-infected cells. Our findings identify a viral protein that is important for changes in lipid metabolism that occur following HCMV infection.

The impact of reducing fatty acid desaturation on the composition and thermal stability of rapeseed oil.

SummaryOilseed rape (Brassica napus) is the third largest source of vegetable oil globally. In addition to food uses, there are industrial applications that exploit the ability of the species to accumulate the very‐long‐chain fatty acid (VLCFA) erucic acid in its seed oil, controlled by orthologues of FATTY ACID ELONGASE 1 (Bna.FAE1.A8 and Bna.FAE1.C3). The proportion of polyunsaturated fatty acids (PUFAs) in rapeseed oil is predicted to affect its thermal stability and is controlled by orthologues of FATTY ACID DESATURASE 2, particularly Bna.FAD2.C5. Our aim was to develop rapeseed lines combining high erucic and low PUFA characters and to assess the impact on thermal stability of the oil they produce. The new type of rapeseed oil (high erucic low polyunsaturate; HELP) contained a substantially greater proportion of erucic acid (54%) compared with high erucic rapeseed oil (46%). Although the total VLCFA content was greater in oil from HELP lines (64%) than from high erucic rapeseed (57%), analysis of triacylglycerol composition showed negligible incorporation of VLCFAs into the sn‐2 position. Rancimat analysis showed that the thermal stability of rapeseed oil was improved greatly as a consequence of reduction of PUFA content, from 3.8 and 4.2 h in conventional low erucic and high erucic rapeseed oils, respectively, to 11.3 and 16.4 h in high oleic low PUFA (HOLP) and HELP oils, respectively. Our results demonstrate that engineering of the lipid biosynthetic pathway of rapeseed, using traditional approaches, enables the production of renewable industrial oils with novel composition and properties.

Seipin negatively regulates sphingolipid production at the ER-LD contact site.

Seipin is known for its critical role in controlling lipid droplet (LD) assembly at the LD-forming subdomain of the endoplasmic reticulum (ER). Here, we identified a new function of seipin as a negative regulator for sphingolipid production. We show that yeast cells lacking seipin displayed altered sensitivity to sphingolipid inhibitors, accumulated sphingoid precursors and intermediates, and increased serine palmitoyltransferase (SPT) and fatty acid (FA) elongase activities. Seipin associated with SPT and FA elongase, and the interaction was reduced by inhibitors for sphingolipid synthesis in a concentration-dependent manner. We further show that the interactions of seipin with SPT and FA elongase occurred at ER-LD contacts and were likely regulated differentially. Further evidence indicated that LD biogenesis was intact when SPT activity was blocked, whereas excess sphingoid intermediates may affect LD morphology. Expression of human seipin rescued the altered sphingolipids in yeast seipin mutants, suggesting that the negative regulation of sphingolipid synthesis by seipin is likely an evolutionarily conserved process.

Maternal seafood consumption during pregnancy and child attention outcomes: a cohort study with gene effect modification by PUFA-related genes.

There is a need to test the fetal programming theoretical framework in nutritional epidemiology. We evaluated whether maternal seafood intake during pregnancy was associated with 8-year-old attention outcomes after adjusting for previous child seafood intake and cognitive function. We also explored effect modification by several single nucleotide polymorphisms (SNPs) related with polyunsaturated fatty acid (PUFA) metabolism. Our final analyses included 1644 mother-child pairs from the prospective INMA (INfancia y Medio Ambiente) cohort study (Spain, recruitment between 2003 and 2008). We used food frequency questionnaires to assess prenatal and postnatal seafood consumption of the mother-child pairs. We evaluated attention function of the children through the computer-based Attention Network Test (ANT) and we used the number of omission errors and the hit reaction time standard error (HRT-SE). Parents reported child attention deficit hyperactivity disorder (ADHD) symptoms using the Revised Conners' Parent Rating Scale Short Form (CPRS-R: S). We measured seven candidate SNPs in a subsample of 845 children. We estimated associations using regression models, adjusting for family characteristics, child seafood intake and cognitive functions at early ages, and to explore SNP effect modifications. Higher total seafood intake during early pregnancy was associated with a reduction of child ANT omission errors, 5th quintile (median = 854 g/week) vs 1st quintile (median = 195 g/week), incidence risk ratio (IRR) 0.76; 95% CI = 0.61, 0.94. Similar results were observed after adjusting the models for child seafood intake and previous cognitive status. Lean, large and small fatty fish showed similar results, and generally similar but less robust associations were observed with the other attention outcomes. Shellfish and canned tuna showed weaker associations. The association patterns were weaker in late pregnancy and null in child seafood consumption. Child rs1260326 (glucokinase regulator, GCKR) and child/maternal rs2281591 (fatty acid elongase 2, ELOVL2) polymorphisms showed nominal P-value for interactions <0.10 between total seafood intake and ANT outcomes. After adjusting for previous child cognitive functions and child seafood intake, high pregnancy consumption (total, lean, small and large fatty fish) was independently associated with improvements of some 8-year-old attention outcomes. Genetic effect modification analyses suggest PUFA intake from seafood as a potential biological mechanism of such association.

Membrane Disruption by Very Long Chain Fatty Acids during Necroptosis.

Necroptosis is a form of regulated cell death which results in loss of plasma membrane integrity, release of intracellular contents, and an associated inflammatory response. We previously found that saturated very long chain fatty acids (VLCFAs), which contain ≥20 carbons, accumulate during necroptosis. Here, we show that genetic knockdown of Fatty Acid (FA) Elongase 7 (ELOVL7) reduces accumulation of specific very long chain FAs during necroptosis, resulting in reduced necroptotic cell death and membrane permeabilization. Conversely, increasing the expression of ELOVL7 increases very long chain fatty acids and membrane permeabilization. In vitro, introduction of the VLCFA C24 FA disrupts bilayer integrity in liposomes to a greater extent than a conventional C16 FA. To investigate the microscopic origin of these observations, atomistic Molecular Dynamics (MD) simulations were performed. MD simulations suggest that fatty acids cause clear differences in bilayers based on length and that it is the interdigitation of C24 FA between the individual leaflets that results in disorder in the region and, consequently, membrane disruption. We synthesized clickable VLCFA analogs and observed that many proteins were acylated by VLCFAs during necroptosis. Taken together, these results confirm the active role of VLCFAs during necroptosis and point to multiple potential mechanisms of membrane disruption including direct permeabilization via bilayer disruption and permeabilization by targeting of proteins to cellular membranes by fatty acylation.

Novel Cellular Functions of Very Long Chain-Fatty Acids: Insight From ELOVL4 Mutations.

Elongation of Very Long chain fatty acids-4 (ELOVL4) protein is a member of the ELOVL family of fatty acid elongases that is collectively responsible for catalyzing formation of long chain fatty acids. ELOVL4 is the only family member that catalyzes production of Very Long Chain Saturated Fatty Acids (VLC-SFA) and Very Long Chain Polyunsaturated Fatty Acids (VLC-PUFA) with chain lengths ≥28 carbons. ELOVL4 and its VLC-SFA and VLC-PUFA products are emerging as important regulators of synaptic signaling and neuronal survival in the central nervous system (CNS). Distinct sets of mutations in ELOVL4 cause three different neurological diseases in humans. Heterozygous inheritance of one set of autosomal dominant ELOVL4 mutations that leads to truncation of the ELOVL4 protein causes Stargardt-like macular dystrophy (STGD3), an aggressive juvenile-onset retinal degeneration. Heterozygous inheritance of a different set of autosomal dominant ELOVL4 mutations that leads to a full-length protein with single amino acid substitutions causes spinocerebellar ataxia 34 (SCA34), a late-onset neurodegenerative disease characterized by gait ataxia and cerebellar atrophy. Homozygous inheritance of a different set of ELOVL4 mutations causes a more severe disease with infantile onset characterized by seizures, spasticity, intellectual disability, ichthyosis, and premature death. ELOVL4 is expressed widely in the CNS and is found primarily in neurons. ELOVL4 is expressed in cell-specific patterns within different regions of the CNS that are likely to be related to disease symptoms. In the retina, ELOVL4 is expressed exclusively in photoreceptors and produces VLC-PUFA that are incorporated into phosphatidylcholine and enriched in the light sensitive membrane disks of the photoreceptor outer segments. VLC-PUFA are enzymatically converted into “elovanoid” compounds that appear to provide paracrine signals that promote photoreceptor and neuronal survival. In the brain, the main ELOVL4 products are VLC-SFA that are incorporated into sphingolipids and enriched in synaptic vesicles, where they regulate kinetics of presynaptic neurotransmitter release. Understanding the function of ELOVL4 and its VLC-SFA and VLC-PUFA products will advance our understanding of basic mechanisms in neural signaling and has potential for developing novel therapies for seizure and neurodegenerative diseases.

De novo transcriptome sequencing of Monodopsis subterranea CCALA 830 and identification of genes involved in the biosynthesis of eicosapentanoic acid and triacylglycerol

Monodopsis subterranea, a unicellular yellow-green freshwater microalga, is widely known for its ability to produce high amount of therapeutically beneficial ω-3 polyunsaturated fatty acid eicosapentanoic acid (EPA; C20:5Δ5,8,11,14,17). Currently, no genomic information is available on M. subterranea despite its nutraceutical and commercial applications. Analysis on fatty acid methyl esters from M. subterranea strain CCALA 830 demonstrated accumulation of 28% EPA. In order to obtain better understanding of EPA biosynthesis and to identify genes involved in the process of lipid metabolism and accumulation in this alga, de novo transcriptome sequencing and assembly was performed using Illumina Hiseq 2000 sequencing. A total of 35,954 transcripts were obtained through final transcriptome assembly with an average transcript length of 769.36 bp. BLAST similarity searches for assembled transcripts were performed followed by annotation using Gene ontology and Kyoto Encyclopedia of Genes and Genomes orthology identifiers. Transcripts involved in lipid biosynthesis including various fatty acid desaturases and elongases involved in PUFAs biosynthesis were identified during the study. In addition, sequences for several transcription factors and a number of simple sequence repeats were also ascertained which can be used as powerful genetic markers for further genetic analysis. This study would provide a useful resource for future research on M. subterranea genome.