Polyunsaturated Fatty Acid Biosynthesis Research Articles

Hepatic transcriptome analysis reveals that elovl5 deletion promotes PUFA biosynthesis and deposition

The safe and low-cost acquisition of polyunsaturated fatty acids (PUFAs) has become a research hotspot. Fatty acyl elongase 5 (Elovl5), a rate-limiting enzyme for fatty acid elongation, is principally in charge of extending C18 and C20 PUFA substrates. However, the role of elovl5 in regulating pathways and genes involved in PUFA synthesis remain largely unknown. Here, hepatic transcriptome analysis of wild-type and elovl5 knockout (elovl5−/−) zebrafish was performed to identify the potential regulatory targets related to PUFA deposition and synthesis. There were 1579 differentially expressed genes (DEGs), of which 787 had their expression levels increased while 792 had the opposite effect. Peroxisome proliferators-activated receptors (PPAR) signaling pathway was considerably enriched in DEGs, according to the KEGG analysis, in which fatp2, fabp7, and pparδ were engaged in PUFA absorption and deposition. Additionally, transcriptome analysis also revealed that cyp46a1 and cyp2r1 were implicated in the synthesis of bile acids and the metabolism of vitamin D, thus indirectly participating in PUFA biosynthesis and deposition. Finally, the DEGs, which improve PUFA level following elovl5 deletion, were verified through feeding experiment with two prepared diets soybean oil diet and linolenic acid oil diet. This study revealed potential regulatory targets that improve PUFA level after elovl5 deletion in teleosts.

Abstract 1250: Aberrant polyunsaturated fatty acids desaturation impairs survival and anti-tumor immunity of tumor resident memory CD8+ T cell in hepatocellular carcinoma

Abstract Background & Aim: Aberrant polyunsaturated fatty acids (PUFAs) metabolism is closely associated with the immunosuppressive tumor microenvironment (TME). Tissue-resident memory T cell (Trm) plays a central role against solid tumors, including HCC, depending on the uptake of exogenous fatty acids to maintain survival and anti-tumor immunity. Here we aimed to explore the potential interaction in TME between HCC-derived PUFAs and Trm function in a series of integrated clinical, basic and translational studies. Methods: Tissue interstitial fluids (TIFs) were isolated and analyzed from fresh paired adjacent tissues and HCC for detecting the PUFAs alterations in the HCC TME. Integrated analyses were performed in multiple public databases and local cohorts. Fatty acid desaturase 1 (FADS1) knockdown (KD) HCC cell line was generated to observe the tumor biology and metabolic function. The effects of FADS1 knockout (KO) or overexpression were investigated in multiple spontaneous HCC mouse models via hydrodynamic tail vein injection (HTVI) in vivo. Mutation landscape analysis was used to explore the mechanism of FADS1 transcription. Therapeutic interventions by downregulating FADS1 were also evaluated in vivo. Results: Clinically, we found that the arachidonic acid (ARA) level was enriched in HCC TIFs. A rate-limiting enzyme controlling PUFA biosynthesis, especially ARA production from dihomo-γ-linolenic acid (DGLA), FADS1, was overexpressed in HCC patients. High expression of FADS1 predicted poor survival and was negatively correlated with CD8+CD103+Trm infiltration. Moreover, in the local clinical trial, we noticed that the expression of FADS1 was upregulated in non-response patients with anti-PD-1 mAb treatment. Functionally, we demonstrated that the ARA level was decreased in the conditioned medium (CM) from FADS1 KD cells. Applying CM prolonged the survival of CD8+CD103+Trm ex vivo. FADS1 KO inhibited tumor development by increasing intratumor infiltration and function of CD8+CD103+Trm. In contrast, overexpression of FADS1 promoted HCC progression by restraining immune surveillance in vivo. Mechanistically, we found that the expression of FADS1 was upregulated in the TP53 mutation group by analyzing the mutation landscape of HCC. Mutant P53 protein could bind to the promoter region of FADS1. Therapeutically, recombinant adeno-associated 8 (rAAV8)-shRNAs delivery and anti-PD-1 mAb administration inhibited the tumor growth of anti-PD-1 mAb treatment-resistant HCC model driven by CTNNB1N90 and MYC. Conclusions: We demonstrated that mutant P53 protein directly promoted FADS1 transcription, increasing ARA accumulation in HCC TME. FADS1 remodeling the metabolic patterns of PUFAs impaired CD8+CD103+Trm anti-tumor immunity, possibly leading to HCC progression. Citation Format: Tao Ding, Li Pang, Runying Long, Shinuan Zeng, Kevin TP Ng, Qingmei Zhang, Zhiwei Chen, Kwan Man. Aberrant polyunsaturated fatty acids desaturation impairs survival and anti-tumor immunity of tumor resident memory CD8+ T cell in hepatocellular carcinoma [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2023; Part 1 (Regular and Invited Abstracts); 2023 Apr 14-19; Orlando, FL. Philadelphia (PA): AACR; Cancer Res 2023;83(7_Suppl):Abstract nr 1250.

Functional and Compositional Changes in the Fecal Microbiome of a Shorebird during Migratory Stopover.

Shorebirds migrate long distances twice annually, which requires intense physiological and morphological adaptations, including the ability to rapidly gain weight via fat deposition at stopover locations. The role of the microbiome in weight gain in avian hosts is unresolved, but there is substantial evidence to support the hypothesis that the microbiome is involved with host weight from mammalian microbiome literature. Here, we collected 100 fecal samples of Ruddy Turnstones to investigate microbiome composition and function during stopover weight gain in Delaware Bay, USA. Using 16S rRNA sequencing on 90 of these samples and metatranscriptomic sequencing on 22, we show that taxonomic composition of the microbiome shifts during weight gain, as do functional aspects of the metatranscriptome. We identified 10 genes that are associated with weight class, and polyunsaturated fatty acid biosynthesis in the microbiota is significantly increasing as birds gain weight. Our results support that the microbiome is a dynamic feature of host biology that interacts with both the host and the environment and may be involved in the rapid weight gain of shorebirds. IMPORTANCE Many animals migrate long distances annually, and these journeys require intense physiological and morphological adaptations. One such adaptation in shorebirds is the ability to rapidly gain weight at stopover locations in the middle of their migrations. The role of the microbiome in weight gain in birds is unresolved but is likely to play a role. Here, we collected 100 fecal samples from Ruddy Turnstones to investigate microbiome composition (who is there) and function (what they are doing) during stopover weight gain in Delaware Bay, USA. Using multiple molecular methods, we show that both taxonomic composition and function of the microbiome shifts during weight gain. We identified 10 genes that are associated with weight class, and polyunsaturated fatty acid biosynthesis in the microbiota is significantly increasing as birds gain weight. Our results support that the microbiome is a dynamic feature of host biology that interacts with both the host and the environment and may be involved in the rapid weight gain of shorebirds.

RNAseq reveals modulation of genes involved in fatty acid biosynthesis in chicken liver according to genetic background, sex, and diet.

Increases in chicken production are mainly due to specialised breeds. However, local breeds are of increasing importance, known for ability to adapt to the environment and unique products. Conventional poultry products contain lower levels of n-3 fatty acids (FAs) compared to those obtained from local breeds, therefore the aim of this study was to evaluate the modulation of expression of genes involved in long-chain polyunsaturated FA (PUFA) biosynthesis pathways according to genetic background, diet conditions, and sex. Animals from two local breeds and a commercial line were fed different diets: control and experimental diet (10% linseed supplementation). For each breed and diet group, both sexes were reared. The RNA was extracted from 36 liver samples and sequenced by RNAseq method. Bioinformatic analysis was carried out to find differentially expressed genes from comparisons between experimental groups. Results showed low impact of diet on differentially expressed genes related to FA biosynthesis, but linseed diet increased percentage of n-3 FAs of liver. Sex and genetic background determined the differential expression of genes related to long-chain PUFA biosynthesis. Specifically, females of local breeds shared 23 up-regulated genes when compared to their respective commercial line groups. Some of the shared genes had a role in de novo triglyceride biosynthesis (MTTPL and GPAM), and in de novo FA biosynthesis (ACACA and SCD) was detected. In conclusion, local breeds are able to better adapt to a diet rich in PUFA, by triggering certain transcriptomic shifts in the liver that allow birds to process the high PUFA content provided by diet.

GPR120-ERK1-Srebp1c signaling pathway regulates long-chain polyunsaturated fatty acids biosynthesis in marine teleost Siganus canaliculatus.

The rabbitfish Siganus canaliculatus is the first marine teleost reported to possess long-chain polyunsaturated fatty acids (LC-PUFA) biosynthetic ability; its regulatory mechanisms have been investigated at the transcriptional and posttranscriptional levels, but little is known about its regulation at the cellular signaling level. The present study investigated the regulatory role of the G-protein-coupled receptor 120 (GPR120) signaling pathway in LC-PUFA biosynthesis in rabbitfish. S. canaliculatus hepatocyte line (SCHL) cells treated with GRP120 agonists (TUG891 and GW9508) showed significantly lower docosahexaenoic acid (DHA) content and mRNA levels of the key genes involved in LC-PUFA biosynthesis, encoding Δ6/Δ5 Fads2, Elovl5, and transcriptional factor Srebp1c. Transcriptome analysis of the treated SCHL cells showed significantly lower mRNA levels of genes encoding extracellular signal-regulated kinase 1 (ERK1), AMP-activated protein kinase (AMPKα2), target of rapamycin (TORC2) and Srebp1c, suggesting that these proteins are potentially involved in the GRP120 signaling pathway. Moreover, treatment of SCHL cells with signaling chemicals of ERK1, AMPKα2, TORC2, and Srebp1c confirmed the involvement of the ERK1-Srebp1c signaling pathway in the regulation of LC-PUFA biosynthesis. The mRNA levels of Srebp1c, Δ6/Δ5 fads2 and elovl5 were significantly lower in cells treated with PUFAs (linoleic acid, α-linolenic acid, arachidonic acid, eicosapentaenoic acid, DHA) but higher in those treated with ERK1 inhibitors (U0126 and CI-1040). CI-1040-treated cells showed significantly higher DHA content, but the other treatment groups (except PD98059) showed significantly lower DHA content. These results indicate that the GPR120-ERK1-Srebp1c signaling pathway regulates rabbitfish LC-PUFA biosynthesis, representing a novel regulatory mechanism in vertebrates.

Function of the Polyketide Synthase Domains of Schizochytrium sp. on Fatty Acid Synthesis in Yarrowia lipolytica.

It is well known that polyunsaturated fatty acids (PUFAs) in Schizochytrium sp. are mainly synthesized via the polyketide synthase (PKS) pathway. However, the specific mechanism of PKS in fatty acid synthesis is still unclear. In this work, the functions of ORFA, ORFB, ORFC, and their individual functional domain genes on fatty acid synthesis were investigated through heterologous expression in Yarrowia lipolytica. The results showed that the expression of ORFA, ORFB, ORFC, and their individual functional domains all led to the increase of the very long-chain PUFA content (mainly eicosapentaenoic acid). Furthermore, the transcriptomic analysis showed that except for the 3-ketoacyl-ACP synthase (KS) domain of ORFB, the expression of an individual functional domain, including malonyl-CoA: ACP acyltransferase, 3-hydroxyacyl-ACP dehydratase (DH), 3-ketoacyl-ACP reductase, and KS domains of ORFA, acyltransferase domains of ORFB, and two DH domains of ORFC resulted in upregulation of the tricarboxylic acid cycle and pentose phosphate pathway, downregulation of the triacylglycerol biosynthesis, fatty acid synthesis pathway, and β-oxidation in Yarrowia lipolytica. These results provide a theoretical basis for revealing the function of PKS in fatty acid synthesis in Y. lipolytica and elucidate the possible mechanism for PUFA biosynthesis.

Goat FADS2 controlling fatty acid metabolism is directly regulated by SREBP1 in mammary epithelial cells.

Goat milk provides benefits to human health due to its richness in bioactive components, such as polyunsaturated fatty acids (PUFAs). The fatty acid desaturase 2 (FADS2) is the first rate-limiting enzyme in PUFAs biosynthesis. However, its role and transcriptional regulation mechanisms in fatty acid metabolism in dairy goat remain unclear. Here, our study revealed that the FADS2 gene was highly expressed during the peak lactation compared with the dry period, early lactation, and late lactation. The content of triacylglycerol (TAG) was enhanced with the increasing mRNA expression of TAG synthesis genes (diacylglycerol acyltransferase 1/2, DGAT1/2) in FADS2-overexpressed goat mammary epithelial cells (GMECs). Overexpression of FADS2 was positively correlated with the elevated concentrations of dihomo-gamma-linolenic acid (DGLA) and docosahexaenoic acid (DHA) in GMECs. BODIPY staining showed that FADS2 promoted lipid droplet accumulation in GMECs. To clarify the transcriptional regulatory mechanisms of FADS2, 2,226 bp length of FADS2 promoter was obtained. Deletion mutation assays revealed that the core region of FADS2 promoter was located between the -375 and -26 region, which contained SRE1 (-361/-351) and SRE2 (-191/-181) cis-acting elements of transcription factor sterol regulatory element-binding protein 1 (SREBP1). Overexpression of SREBP1 enhanced relative luciferase activity of the single mutant of SRE1 or SRE2, vice versa, and failed to alter the relative luciferase activity of the joint mutant of SRE1 and SRE2. Chromatin immunoprecipitation (ChIP) and site-directed mutation assays further demonstrated that SREBP1 regulated the transcription of the FADS2 gene by binding to SRE sites in vivo and in vitro. In addition, the mRNA levels of FADS2 were significantly decreased by targeting SRE1 and SRE2 sites in the genome via the CRISPR interference (CRISPRi) system. These findings establish a direct role for FADS2 regulating TAG and fatty acid synthesis by SREBP1 transcriptional regulation in dairy goat, providing new insights into fatty acid metabolism in mammary gland of ruminants.

Features of the metabolism of polyunsaturated fatty acids in type 2 diabetes mellitus

Type 2 diabetes mellitus (T2DM) is a widespread disease with a high risk of cardiovascular complications, disability and mortality. The progression of T2DM is closely related to lipid metabolism disorders, caused both by insufficient intake of polyunsaturated fatty acids (PUFAs), and by a violation of their endogenous metabolism. Desaturase enzymes, FADS1/2, are involved in the regulation of PUFA metabolism. Violation of the functioning of FADS1/2 and their genes leads to a change in the biosynthesis of PUFAs and the fatty acid composition of cell membranes. The purpose of this research was to summarize the data of modern literature on the metabolism of PUFAs and the effect of FADS genetic variants on the fatty acid composition of cell membranes in T2DM. Material and methods. The search and analysis of publications was carried out using the PubMed, MEDLINE, Web of Science databases, mainly for the last 10 years, using the search keywords: polyunsaturated fatty acids, fatty acid desaturases, desaturase genes, type 2 diabetes mellitus. Results. In the pathogenesis of T2DM and its complications, many factors play a role, including impaired PUFA metabolism. A large evidence base has been accumulated on the effect of PUFAs on cardiometabolic risk factors in T2DM. The effect of desaturase activity on the fatty acid composition of cells was identified as the most important link in the metabolism of PUFAs. Focusing on the modulation of desaturase activity and studying the polymorphism of fatty acid desaturase genes may be a useful therapeutic option in the treatment of patients with T2DM and its complications. Conclusion. A promising direction of scientific research in the treatment and prevention of T2DM and its complications is the study of genetic mechanisms associated with the metabolism of PUFAs and their metabolites.

Effect of Overexpression of γ-Tocopherol Methyltransferase on α-Tocopherol and Fatty Acid Accumulation and Tolerance to Salt Stress during Seed Germination in Brassica napus L.

Rapeseed (Brassica napus L.) is an important oil crop and a major source of tocopherols, also known as vitamin E, in human nutrition. Enhancing the quality and composition of fatty acids (FAs) and tocopherols in seeds has long been a target for rapeseed breeding. The gene γ-Tocopherol methyltransferase (γ-TMT) encodes an enzyme catalysing the conversion of γ-tocopherol to α-tocopherol, which has the highest biological activity. However, the genetic basis of γ-TMT in B. napus seeds remains unclear. In the present study, BnaC02.TMT.a, one paralogue of Brassica napus γ-TMT, was isolated from the B. napus cultivar "Zhongshuang11" by nested PCR, and two homozygous transgenic overexpression lines were further characterised. Our results demonstrated that the overexpression of BnaC02.TMT.a mediated an increase in the α- and total tocopherol content in transgenic B. napus seeds. Interestingly, the FA composition was also altered in the transgenic plants; a reduction in the levels of oleic acid and an increase in the levels of linoleic acid and linolenic acid were observed. Consistently, BnaC02.TMT.a promoted the expression of BnFAD2 and BnFAD3, which are involved in the biosynthesis of polyunsaturated fatty acids during seed development. In addition, BnaC02.TMT.a enhanced the tolerance to salt stress by scavenging reactive oxygen species (ROS) during seed germination in B. napus. Our results suggest that BnaC02.TMT.a could affect the tocopherol content and FA composition and play a positive role in regulating the rapeseed response to salt stress by modulating the ROS scavenging system. This study broadens our understanding of the function of the Bnγ-TMT gene and provides a novel strategy for genetic engineering in rapeseed breeding.

Lipid desaturation regulates the balance between self-renewal and differentiation in mouse blastocyst-derived stem cells

Stem cells are defined by their ability to self-renew and differentiate, both shown in multiple studies to be regulated by metabolic processes. To decipher metabolic signatures of self-renewal in blastocyst-derived stem cells, we compared early differentiating embryonic stem cells (ESCs) and their extra-embryonic counterparts, trophoblast (T)SCs to their self-renewing counterparts. A metabolomics analysis pointed to the desaturation of fatty acyl chains as a metabolic signature of differentiating blastocyst-derived SCs via the upregulation of delta-6 desaturase (D6D; FADS2) and delta-5 desaturase (D5D; FADS1), key enzymes in the biosynthesis of polyunsaturated fatty acids (PUFAs). The inhibition of D6D or D5D by specific inhibitors or SiRNA retained stemness in ESCs and TSCs, and attenuated endoplasmic reticulum (ER) stress-related apoptosis. D6D inhibition in ESCs upregulated stearoyl-CoA desaturase-1 (Scd1), essential to maintain ER homeostasis. In TSCs, however, D6D inhibition downregulated Scd1. TSCs show higher Scd1 mRNA expression and high levels of monounsaturated fatty acyl chain products in comparison to ESCs. The addition of oleic acid, the product of Scd1 (essential for ESCs), to culture medium, was detrimental to TSCs. Interestingly, TSCs express a high molecular mass variant of Scd1 protein, hardly expressed by ESCs. Taken together, our data suggest that lipid desaturation is a metabolic regulator of the balance between differentiation and self-renewal of ESCs and TSCs. They point to lipid polydesaturation as a driver of differentiation in both cell types. Monounsaturated fatty acids (MUFAs), essential for ESCs are detrimental to TSCs.

Dominant Elongase Activity of Elovl5a but Higher Expression of Elovl5b in Common Carp (Cyprinus carpio)

Most diploid freshwater and marine fish encode one elovl5 elongase, having substrate specificity and activities towards C18, C20 and C22 polyunsaturated fatty acids (PUFAs). The allo-tetraploid common carp is hypothesized to encode two duplicated elovl5 genes. How these two elovl5 genes adapt to coordinate the PUFA biosynthesis through elongase function and expression divergence requires elucidation. In this study, we obtained the full-length cDNA sequences of two elovl5 genes in common carp, named as elovl5a and elovl5b. Functional characterization showed that both enzymes had elongase activity towards C18, C20 and C22 PUFAs. Especially, the activities of these two enzymes towards C22 PUFAs ranged from 3.87% to 8.24%, higher than those in most freshwater and marine fish. The Elovl5a had higher elongase activities than Elovl5b towards seven substrates. The spatial-temporal expression showed that both genes co-transcribed in all tissues and development stages. However, the expression levels of elovl5b were significantly higher than those of elovl5a in all examined conditions, suggesting that elovl5b would be the dominantly expressed gene. These two genes had different potential transcriptional binding sites. These results revealed the complicated roles of elovl5 on PUFA synthesis in common carp. The data also increased the knowledge of co-ordination between two homoeologs of the polyploid fish through function and expression divergence.

Evaluation of cottonseed oil as a substitute for dietary fish oil of juvenile black seabream (Acanthopagrus schlegelii): Based on growth, lipid metabolism, antioxidant capacity and pi3k/akt pathway

An eight weeks feeding trial was conducted to evaluate the potential of cottonseed oil (CSO) as a substitute for fish oil (FO) on juvenile black seabream (Acanthopagrus schlegelii) (2.29 ± 0.01 g). Five diets were formulated with pure FO (FO) or partial CSO replacement of FO: 20% CSO (CSO20), 40% CSO (CSO40), 60% CSO (CSO60) and 80% CSO (CSO80), respectively. Results showed that fish fed CSO60 diet significantly increased growth performance compared to control group. Hepatic expression of genes related to long-chain polyunsaturated fatty acid biosynthesis were significantly up-regulated by dietary CSO60 compared to FO group. Hepatic paraffin section and serum biochemical indices showed that fish fed dietary CSO80 could increase vacuolar fat drops and cause liver damage. The expression levels of peroxisome proliferators-activated receptor alpha and its downstream lipolysis related genes were markedly up-regulated with the increasing of replacement levels of CSO, opposite results were recorded in sterol regulatory element-binding protein-1c and lipogenesis related genes. The malondialdehyde content in liver was decreased with the increasing replacement levels of CSO, whereas the antioxidant parameters were significantly higher in CSO60 group compared to FO group. Phosphatidylinositol-3 kinase and protein kinase B expression levels were significantly up-regulated by dietary CSO80, triggering inflammation by activating nuclear factor kappa b. Dietary CSO60 dramatically promote target of rapamycin expression level. With the increasing replacement levels of FO by CSO, the expression level of ribosomal protein S6 kinase 1 was significantly increased, whereas opposite result was found for the eukaryotic translation initiation factor 4E binding protein 2. Collectively, based on weight gain, double broken line model analysis showed that a maximum growth occurred at 59.13% replacement of dietary FO by CSO. These findings demonstrate that dietary CSO60 can improve growth and antioxidant capacity. However, dietary CSO80 can result in lipid deposition, liver injury and inflammation, hence, suppress growth performance.

Contribution of anthocyanin and polyunsaturated fatty acid biosynthesis to cold tolerance during bud sprouting in tree peony

Tree peony is an economically important crop that is widely used in the food, floriculture, and pharmaceutical industries. However, the buds of tree peony are susceptible to low temperature stress. To elaborate the metabolic and transcriptional changes during bud sprouting, we performed metabolite and transcriptome analyses of the buds of Paeonia suffruticosa cultivar ‘Yulouchun’ at three developmental stages under low temperature treatment. The results showed that the contents of anthocyanin and α-linolenic acid (ALA) increased, concomitant with increased production of sucrose, gibberellic acid, indolylacetic acid, trans -zeatin riboside, and jasmonic acid as well as reduced accumulation of abscisic acid. Transcriptome profiling revealed 6355 differentially expressed genes (DEGs) during bud growth. We identified many structural DEGs whose expression was correlated with the accumulation of anthocyanin, ALA, sucrose, and hormones. In particular, the expression of transcription factors (TFs) PsMYB113 and PsMYB123 was found to be positively correlated with anthocyanin biosynthetic genes flavonoid 3′,5′-Hydroxylase , dihydroflavonol 4-reductase ( DFR ), anthocyanidin synthase ( ANS ), and two UDP-glycosylatransferase s. A positive correlation between a bZIP TF PsDPBF3 and an ALA biosynthetic gene fatty acid desaturase3 ( FAD3 ) was also found. Transcript abundances of these TFs and structural genes changed under sucrose and hormone treatments. Silencing of PsMYB113 and PsMYB123 resulted in decreased anthocyanin production and tolerance to freezing in tree peony buds. In addition, PsDPBF3 -silenced buds displayed reduced ALA levels and low temperature hardiness. In vivo promoter-binding assays verified that both PsMYB113 and PsMYB123 trans-activated the expression of DFR and ANS , while PsDPBF3 specifically regulated the transcription of FAD3 . Taken together, our data suggest that the production of anthocyanin and polyunsaturated fatty acid is closely implicated in cold tolerance of tree peony buds. • Cold stress increased various metabolite contents during bud sprouting of tree peony. • Transcriptome profiling revealed 6355 DEGs in cold-treated tree peony buds. • Silencing of PsMYB113 , PsMYB123 , and PsDPBF3 led to reduced tolerance to cold stress. • PsMYB113 and PsMYB123 bound to the promoters of anthocyanin-related DFR and ANS. • PsDPBF3 specifically trans-activated the expression of PUFA-associated FAD3 .

Effects of low salinities on growth, fatty acid composition, and transcriptome in Florida pompano (Trachinotus carolinus) at early developmental stages

The Florida pompano, Trachinotus carolinus, is a prime marine finfish candidate for commercial production in the United States (U.S.). Low salinity conditions (i.e., < 30 parts per thousand, ppt) can affect the early development of T. carolinus by increasing osmoregulatory stress. To address the knowledge gap regarding the impact of low salinity on Florida pompano health, we conducted an experiment to determine the optimal salinity for the culture of fingerlings. Larvae were reared in three salinities (10, 20, and 30 ppt) at local producers' farms and growth, fatty acid profiles, and transcriptome responses were quantified every three days after hatching (3, 6, 9, 12, 15, 18, and 24 days post hatch) and results were compared across salinities. Larvae reared at 10 ppt grew at a normal rate and the developmental stage had a greater impact than salinity on fatty acid composition and larval gene expression. Although there were few differentially expressed genes between salinities (n = 17), there was an upregulation of gene sets associated with ribosomes and oxidative phosphorylation and a downregulation of glycerophospholipid and carbon metabolism at lower salinities in comparison to higher salinities. In the early days of development, the fatty acids linoleic acid (LA), alpha linoleic acid (LNA), and 18:3 n-6 were proven to be important as potential energy fuels and precursors for long-chain polyunsaturated fatty acids (LC-PUFA) biosynthesis, which was supported by the whole transcriptomics analysis. The gene expression of fatty acid synthesis genes coincided with changes in the fatty acid levels suggesting the ability of Florida pompano larvae to biosynthesize LC-PUFAs from LA and LNA precursors. Overall, this study suggests that it is possible to raise Florida pompano larvae for 24 DPH under low salinity conditions as low as 10 ppt. However, it is important to run this study for a longer period to investigate the long-term effects of low salinity on larval health and development. Importantly, this research serves as a model study for future on-farm collaborations and helps build a bridge between scientists and farmers in aquaculture.

Computationally Guided Enzymatic Studies on Schizochytrium-Sourced Malonyl-CoA:ACP Transacylase.

The malonyl-CoA:ACP transacylase (MAT) domain is responsible for the selection and incorporation of malonyl building blocks in the biosynthesis of polyunsaturated fatty acids (PUFAs) in eukaryotic microalgae (Schizochytrium) and marine bacteria (Moritella marina, Photobacterium profundum, and Shewanella). Elucidation of the structural basis underlying the substrate specificity and catalytic mechanism of the MAT will help to improve the yield and quality of PUFAs. Here, a methodology guided by molecular dynamics simulations was carried out to identify and mutate specificity-conferring residues within the MAT domain of Schizochytrium. Combining mutagenesis, cell-free protein synthesis, and in vitro biochemical assay, we dissected nearby interactions and molecular mechanisms relevant for binding and catalysis and found that the reorientation of the Ser154 Cβ-Oγ bond establishes distinctive proton-transfer chains (His153-Ser154 and Asn235-His153-Ser154) for catalysis. Gln66 can be replaced by tyrosine to shorten the distance between His153 (Nε2) and Ser154 (Oγ), which facilitates a faster proton-transfer rate, allowing better use of acyl substrates than the wild type. Furthermore, we screened a mutant that displayed an 18.4% increase in PUFA accumulation. These findings provide important insights into the study of MAT through protein engineering and will benefit dissecting the molecular mechanisms of other PUFA-related catalytic domains.

Long-chain polyunsaturated fatty acid biosynthesis in a land-crab with advanced terrestrial adaptations: Molecular cloning and functional characterization of two fatty acyl elongases.

Depending on the presence and activities of the front-end fatty acyl desaturases and elongation of very long-chain fatty acid (Elovl) enzymes, animals have different capacities for long-chain (≥C20) polyunsaturated fatty acids (LC-PUFA) biosynthesis. Successful land colonisation in brachyuran crabs requires a shift towards terrestrial food chain with limited LC-PUFA availability. We cloned and functionally characterised two elovl genes from the purple land crab Gecarcoidea lalandii. The two Elovl contained all the necessary motifs of a typical polyunsaturated fatty acids (PUFA) Elovl and phylogenetically clustered in the Elovl1 and Elovl6 clades, respectively. The G. lalandii Elovl1 elongated saturated fatty acids, with low activities towards C20 and C22 PUFA substrates. Moreover, the G. lalandii Elovl6 was particularly active in the elongation of C18 PUFA, although it also recognised monounsaturated fatty acids as substrates for elongation. Collectively, the herein characterised G. lalandii elovl paralogues fulfil all the elongation steps involved in the LC-PUFA biosynthetic pathways. Tissue distribution of the G. lalandii elovl genes, along with the FA composition analyses, suggest the hepatopancreas and gill as key metabolic sites for fatty acid elongation. However, current data suggest that G. lalandii is unable to rely solely on biosynthesis to fulfil LC-PUFA requirements, since front-end desaturase appears to be absent in this species and other decapods.

Fatty acid desaturase 1 (FADS1) is a cancer marker for patient survival and a potential novel target for precision cancer treatment.

Fatty Acid Desaturase-1 (FADS1) or delta 5 desaturase (D5D) is a rate-limiting enzyme involved in the biosynthesis of long-chain polyunsaturated fatty acids (LC-PUFAs), i.e., arachidonic acid (ARA) and eicosapentaenoic (EPA). These LC-PUFAs and their metabolites play essential and broad roles in cancer cell proliferation, metastasis, and tumor microenvironment. However, the role of FADS1 in cancers remains incompletely understood. Utilizing The Cancer Genome Atlas (TCGA) database, we explored the role of FADS1 across different cancer types using multiple bioinformatics and statistical tools. Moreover, we studied the impact of a FADS1 inhibitor (D5D-IN-326) on proliferation of multiple cancer cell lines. We identified that FADS1 gene is a predictor for cancer survival in multiple cancer types. Compared to normal tissue, the mRNA expression of FADS1 is significantly increased in primary tumors while even higher in metastatic and recurrent tumors. Mechanistically, pathway analysis demonstrated that FADS1 is associated with cholesterol biosynthesis and cell cycle control genes. Interestingly, FADS1 expression is higher when TP53 is mutated. Tumors with increased FADS1 expression also demonstrated an increased signatures of fibroblasts and macrophages infiltration among most cancer types. Our in vitro assays showed that D5D-IN-326 significantly inhibited cell proliferation of kidney, colon, breast, and lung cancer cell lines in a dose-dependent manner. Lastly, single nucleotide polymorphisms (SNPs) which are well-established expression quantitative trait loci (eQTLs) for FADS1 in normal human tissues are also significantly correlated with FADS1 expression in tumors of multiple tissue types, potentially serving as a marker to stratify cancer patients with high/low FADS1 expression in their tumor tissue. Our study suggests that FADS1 plays multiple roles in cancer biology and is potentially a novel target for precision cancer treatment.

A chromosome-level genome assembly of the jade perch (Scortum barcoo)

Endemic to Australia, jade perch (Scortum barcoo) is a highly profitable freshwater bass species. It has extraordinarily high levels of omega-3 polyunsaturated fatty acids (PUFAs), which detailed genes involved in are largely unclear. Meanwhile, there were four chromosome-level bass species have been previous sequenced, while the bass ancestor genome karyotypes have not been estimated. Therefore, we sequenced, assembled and annotated a genome of jade perch to characterize the detailed genes for biosynthesis of omega-3 PUFAs and to deduce the bass ancestor genome karyotypes. We constructed a chromosome-level genome assembly with 24 pairs of chromosomes, 657.7 Mb in total length, and the contig and the scaffold N50 of 4.8 Mb and 28.6 Mb respectively. We also identified repetitive elements (accounting for 19.7% of the genome assembly) and predicted 26,905 protein-coding genes. Meanwhile, we performed genome-wide localization and characterization of several important genes encoding some key enzymes in the biosynthesis pathway of PUFAs. These genes may contribute to the high concentration of omega-3 in jade perch. Moreover, we conducted a series of comparative genomic analyses among four representative bass species at a chromosome level, resulting in a series of sequences of a deductive bass ancestor genome.

The MAP-Kinase HOG1 Controls Cold Adaptation in Rhodosporidium kratochvilovae by Promoting Biosynthesis of Polyunsaturated Fatty Acids and Glycerol.

The aim of this study was to investigate the role of RKHog1 in the cold adaptation of Rhodosporidium kratochvilovae strain YM25235 and elucidate the correlation of biosynthesis of polyunsaturated fatty acids (PUFAs) and glycerol with its cold adaptation. The YM25235 strain was subjected to salt, osmotic, and cold stress tolerance analyses. mRNA levels of RKhog1, Δ12/15-fatty acid desaturase gene (RKD12), RKMsn4, HisK2301, and RKGPD1 in YM25235 were detected by reverse transcription quantitative real-time PCR. The contents of PUFAs, such as linoleic acid (LA) and linolenic acid (ALA) was measured using a gas chromatography-mass spectrometer, followed by determination of the growth rate of YM25235 and its glycerol content at low temperature. The RKHog1 overexpression, knockout, and remediation strains were constructed. Stress resistance analysis showed that overexpression of RKHog1 gene increased the biosynthesis of glycerol and enhanced the tolerance of YM25235 to cold, salt, and osmotic stresses, respectively. Inversely, the knockout of RKHog1 gene decreased the biosynthesis of glycerol and inhibited the tolerance of YM25235 to different stresses. Fatty acid analysis showed that the overexpression of RKHog1 gene in YM25235 significantly increased the content of LA and ALA, but RKHog1 gene knockout YM25235 strain had decreased content of LA and ALA. In addition, the mRNA expression level of RKD12, RKMsn4, RKHisK2301, and RKGPD1 showed an increase at 15°C after RKHog1 gene overexpression but were unchanged at 30°C. RKHog1 could regulate the growth adaptability and PUFA content of YM25235 at low temperature and this could be helpful for the cold adaptation of YM25235.

Polyunsaturated fatty acids promote the rapid fusion of lipid droplets in Caenorhabditis elegans

Lipid droplets (LDs) are intracellular organelles that dynamically regulate lipids and energy homeostasis in the cell. LDs can grow through either local lipid synthesis or LD fusion. However, how lipids involving in LD fusion for LD growth is largely unknown. Here, we show that genetic mutation of acox-3 (acyl-CoA oxidase), maoc-1 (enoyl-CoA hydratase), dhs-28 (3-hydroxylacyl-CoA dehydrogenase), and daf-22 (3-ketoacyl-CoA thiolase), all involved in the peroxisomal β-oxidation pathway in Caenorhabditis elegans, led to rapid fusion of adjacent LDs to form giant LDs (gLDs). Mechanistically, we show that dysfunction of peroxisomal β-oxidation results in the accumulation of long-chain fatty acid-CoA and phosphocholine, which may activate the sterol-binding protein 1/sterol regulatory element–binding protein to promote gLD formation. Furthermore, we found that inactivation of either FAT-2 (delta-12 desaturase) or FAT-3 and FAT-1 (delta-15 desaturase and delta-6 desaturase, respectively) to block the biosynthesis of polyunsaturated fatty acids (PUFAs) with three or more double bonds (n≥3-PUFAs) fully repressed the formation of gLDs; in contrast, dietary supplementation of n≥3-PUFAs or phosphocholine bearing these PUFAs led to recovery of the formation of gLDs in peroxisomal β-oxidation–defective worms lacking PUFA biosynthesis. Thus, we conclude that n≥3-PUFAs, distinct from other well-known lipids and proteins, promote rapid LD fusion leading to LD growth.