Long-chain acyl-CoA Synthetase Research Articles

Evolution, characterization, and immune response function of long-chain acyl-CoA synthetase genes in rainbow trout (Oncorhynchus mykiss) under hypoxic stress.

Long-chain acyl-CoA synthetases (Acsls), members of the acyl-activating enzyme superfamily, haves been systematically characterized in mammals and certain fishes, but the research on their involvement in reproductive development and hypoxic stress response in rainbow trout remains limited. In this study, we investigated the acsl gene structure and physical and chemical characteristics and the evolutionary relationship among acsl genes using the NCBI/Ensembl database. Using hypoxia treatment experiment, acsl gene expression in various organs and its regulation were investigated. A total of 11 acsl genes were identified in rainbow trout. Phylogenetic analyses found that acsl genes in rainbow trout were clustered into two clades: acsl3/4 and acsl1/2/5/6, and the additional gene duplication observed resulted from the third round of genome duplication unique to teleosts. Multiple sequence alignment and conserved motif analyses showed that the sequence of acsl proteins was highly conserved. Real-time quantitative PCR (RT-qPCR) showed that the acsl genes were highly expressed in immune tissues (liver and head kidney). Under hypoxia, the expression of acsl genes was upregulated, suggesting that they enhance the tolerance to hypoxia and are involved in the immune response in rainbow trout. Our study provides valuable insights into teleost evolution and effects of hypoxia on biological immunity and form a basis for further research on the functional characteristics of acsl genes.

Role of fatty acid transport protein 4 in metabolic tissues: insights into obesity and fatty liver disease.

Fatty acid (FA) metabolism is a series of processes that provide structural substances, signalling molecules and energy. Ample evidence has shown that FA uptake is mediated by plasma membrane transporters including FA transport proteins (FATPs), caveolin-1, fatty-acid translocase (FAT)/CD36, and fatty-acid binding proteins. Unlike other FA transporters, the functions of FATPs have been controversial because they contain both motifs of FA transport and fatty acyl-CoA synthetase (ACS). The widely distributed FATP4 is not a direct FA transporter but plays a predominant function as an ACS. FATP4 deficiency causes ichthyosis premature syndrome in mice and humans associated with suppression of polar lipids but an increase in neutral lipids including triglycerides (TGs). Such a shift has been extensively characterized in enterocyte-, hepatocyte-, and adipocyte-specific Fatp4-deficient mice. The mutants under obese and non-obese fatty livers induced by different diets persistently show an increase in blood non-esterified free fatty acids and glycerol indicating the lipolysis of TGs. This review also focuses on FATP4 role on regulatory networks and factors that modulate FATP4 expression in metabolic tissues including intestine, liver, muscle, and adipose tissues. Metabolic disorders especially regarding blood lipids by FATP4 deficiency in different cell types are herein discussed. Our results may be applicable to not only patients with FATP4 mutations but also represent a model of dysregulated lipid homeostasis, thus providing mechanistic insights into obesity and development of fatty liver disease.

Correction: Myeloid-Specific Deficiency of Long-Chain Acyl CoA Synthetase 4 Reduces Inflammation by Remodeling Phospholipids and Reducing Production of Arachidonic Acid-Derived Proinflammatory Lipid Mediators.

Andrew R. Reeves, Brian E. Sansbury, Meixia Pan, Xianlin Han, Matthew Spite, and Andrew S. Greenberg. 2021. Myeloid-Specific Deficiency of Long-Chain Acyl CoA Synthetase 4 Reduces Inflammation by Remodeling Phospholipids and Reducing Production of Arachidonic Acid–Derived Proinflammatory Lipid

Expression of Physaria longchain acyl-CoA synthetases and hydroxy fatty acid accumulation in transgenic Arabidopsis

Hydroxy fatty acids (HFA) are industrially useful chemical feedstocks that accumulate in seed-storage triacylglycerols (TAG) of several plant species, including castor (Ricinus communis) and Physaria (Physaria fendleri). For researchers, HFA also offer a unique opportunity to trace fatty acid metabolism and modification. Past work producing HFA in Arabidopsis (Arabidopsis thaliana) has demonstrated the importance of isozymes of TAG synthesis from plants that evolved to store HFA and as a result have a high degree of specificity towards HFA substrates. Castor phospholipase A2α (RcPLA2) has specificity for HFA-containing phosphatidylcholine. However, expression of RcPLA2 in HFA-accumulating Arabidopsis line CL37-PLA2 reduced HFA content of TAG. This loss was interpreted as being due to poor ability of Arabidopsis longchain acyl-CoA synthetases (LACSs) to utilize HFAs substrates. LACS enzymes are essential to activate HFA to HFA-CoA for TAG synthesis. Physaria is a close relative of Arabidopsis in the Brassicaceae family. To test the hypothesis that this close relatedness would allow Physaria LACSs to interface successfully with Arabidopsis enzymes of seed lipid metabolism and thereby restore HFA accumulation, we transformed PfLACS4 and PfLACS8 constructs into the CL37-PLA2 line. However, HFA content was not recovered, and biochemical characterization of recombinant PfLACS4 and PfLACS8 indicated that these isozymes have substrate specificities and selectivities that are similar to their Arabidopsis orthologues. These and other results pose an important question about how HFA synthesized on phosphatidylcholine can be transferred into the acyl-CoA pool for TAG synthesis.

A proteomics approach reveals digestive and nutritional responses to food intake in anadromous Coilia nasus

The estuarine tapertail anchovy, Coilia nasus, is an anadromous fish that undertakes over a 600-km spawning migration along the Yangtze River of China. They generally cease feeding during this process, but we recently documented that a small proportion of them appear to feed. Research on proteomic responses is essential for understanding the phenomenon of C. nasus feeding. In this study, we used an iTRAQ-based proteomics approach to study the changes in protein expression in response to food intake in C. nasus following voluntary fasting. Coilia nasus in the feeding group (CSI) were fed shrimp or small fish, whereas those in the control group (CSN) were starved. We identified 3279 proteins in the gastric tissue/stomach, of which 279 were significantly differentially expressed. In all, 133 differentially expressed proteins (DEPs) were upregulated and 146 proteins were downregulated in CSI compared with those in CSN C. nasus. In addition to gastric acid secretion caused by gastric distention, a functional analysis suggested that a series of DEPs were involved mainly in the regulation of protein digestion (e.g., carboxypeptidase A1 and chymotrypsin A-like), immune response (e.g., lysozyme and alpha 2-macroglobulin), and nutrition metabolism (e.g., glyceraldehyde 3-phosphate dehydrogenase, glycogenin, long-chain acyl-CoA synthetase, and creatine kinase). Real-time PCR confirmed that the mRNA levels of the DEPs were similar those obtained using iTRAQ. These results indicate that the nutrients obtained through food were effectively utilized by C. nasus, thereby providing energy for swimming, gonadal maturation, primary metabolism, and an enhanced immune function to better resist pathogen interference. This research contributes to the elucidation of nutritional regulation mechanisms of C. nasus to better protect the wild population.

Fatty acid oxidation protects cancer cells from apoptosis by increasing mitochondrial membrane lipids.

Overcoming resistance to chemotherapies remains a major unmet need for cancers, such as triple-negative breast cancer (TNBC). Therefore, mechanistic studies to provide insight for drug development are urgently needed to overcome TNBC therapy resistance. Recently, an important role of fatty acid β-oxidation (FAO) in chemoresistance has been shown. But how FAO might mitigate tumor cell apoptosis by chemotherapy is unclear. Here, we show that elevated FAO activates STAT3 by acetylation via elevated acetyl-coenzyme A (CoA). Acetylated STAT3 upregulates expression of long-chain acyl-CoA synthetase 4 (ACSL4), resulting in increased phospholipid synthesis. Elevating phospholipids in mitochondrial membranes leads to heightened mitochondrial integrity, which in turn overcomes chemotherapy-induced tumor cell apoptosis. Conversely, in both cultured tumor cells and xenograft tumors, enhanced cancer cell apoptosis by inhibiting ASCL4 or specifically targeting acetylated-STAT3 is associated with a reduction in phospholipids within mitochondrial membranes. This study demonstrates a critical mechanism underlying tumor cell chemoresistance.

Explaining Unsaturated Fatty Acids (UFAs), Especially Polyunsaturated Fatty Acid (PUFA) Content in Subcutaneous Fat of Yaks of Different Sex by Differential Proteome Analysis.

Residents on the Tibetan Plateau intake a lot of yak subcutaneous fat by diet. Modern healthy diet ideas demand higher unsaturated fatty acids (UFAs), especially polyunsaturated fatty acid (PUFA) content in meat. Here, the gas chromatography (GC) and tandem mass tag (TMT) proteomic approaches were applied to explore the relationship between the proteomic differences and UFA and PUFA content in the subcutaneous fat of yaks with different sex. Compared with male yaks (MYs), the absolute contents of UFAs, monounsaturated fatty acids (MUFAs) and PUFAs in the subcutaneous fat of female yaks (FYs) were all higher (p < 0.01); the relative content of MUFAs and PUFAs in MY subcutaneous fat was higher, and the value of PUFAs/SFAs was above 0.4, so the MY subcutaneous fat is more healthy for consumers. Further studies showed the transcriptional regulation by peroxisome proliferator-activated receptor delta (PPARD) played a key role in the regulation of UFAs, especially PUFA content in yaks of different sex. In FY subcutaneous fat, the higher abundance of the downstream effector proteins in PPAR signal, including acyl-CoA desaturase (SCD), elongation of very-long-chain fatty acids protein 6 (ELOVL6), lipoprotein lipase (LPL), fatty acid-binding protein (FABP1), very-long-chain (3R)-3-hydroxyacyl-CoA dehydratase 3 (HACD3), long-chain fatty acid CoA ligase 5 (ACSL5) and acyl-CoA-binding protein 2 (ACBP2), promoted the UFAs’ transport and synthesis. The final result was the higher absolute content of c9-C14:1, c9-C18:1, c9,c12-C18:2n-6, c9, c12, c15-C18:3n-3, c5, c8, c11, c14, c17-C20:5n-3, c4, c7, c10, c13, -c16, c19-C22:6n-3, UFAs, MUFAs and PUFAs in FY subcutaneous fat. Further, LPL, FABP1, HACD3, ACSL1 and ACBP2 were the potential biomarkers for PUFA contents in yak subcutaneous fat. This study provides new insights into the molecular mechanisms associated with UFA contents in yak subcutaneous fat.

Inhibition of Fatty Acid Translocase (FAT/CD36) Palmitoylation Enhances Hepatic Fatty Acid β-Oxidation by Increasing Its Localization to Mitochondria and Interaction with Long-Chain Acyl-CoA Synthetase 1.

Aims: Impaired fatty acid oxidation (FAO) in mitochondria of hepatocytes causes lipid accumulation and excessive production of reactive oxygen species (ROS) and oxidative damage, leading to nonalcoholic fatty liver disease (NAFLD). Fatty acid translocase (FAT/cluster of differentiation 36 [CD36]), a transmembrane protein that facilitates the uptake of long-chain fatty acids (LCFAs), is recently found to be involved in FAO. The function of FAT/CD36 is associated with its subcellular localization. Palmitoylation, one of the most common lipid modifications, is generally thought to regulate FAT/CD36 subcellular localization. Here, we aimed to investigate the role of palmitoylation in FAT/CD36 localization to mitochondria and its influence on FAO in hepatocytes. Results: We demonstrated that FAT/CD36 exists on the mitochondria of hepatocytes. Palmitoylation of FAT/CD36 was significantly upregulated in NAFLD. Inhibition of FAT/CD36 palmitoylation resulted in an obvious increase in the distribution of FAT/CD36 to mitochondria of hepatocytes. Depalmitoylated FAT/CD36 on the mitochondrial membrane continues functioning by facilitating fatty acid trafficking to mitochondria. Abundant mitochondrial FAT/CD36 interacted with long-chain acyl-CoA synthetase 1 (ACSL1), and thus, more LCFAs were transported to ACSL1. This led to an increase in the generation of long-chain acyl-CoA, contributing to the enhancement of FAO and alleviating NAFLD. Innovation and Conclusion: This work revealed that inhibiting FAT/CD36 palmitoylation alleviates NAFLD by promoting FAT/CD36 localization to the mitochondria of hepatocytes. Mitochondrial FAT/CD36 functions as a molecular bridge between LCFAs and ACSL1 to increase the production of long-chain acyl-CoA, thus promoting FAO, thereby avoiding lipid accumulation and overproduction of ROS in hepatocytes. Antioxid. Redox Signal. 36, 1081-1100.

The Chlamydomonas transcription factor MYB1 mediates lipid accumulation under nitrogen depletion.

Microalgae accumulate high levels of oil under stress, but the underlying biosynthetic pathways are not fully understood. We sought to identify key regulators of lipid metabolism under stress conditions. We found that the Chlamydomonas reinhardtii gene encoding the MYB-type transcription factor MYB1 is highly induced under stress conditions. Two myb1 mutants accumulated less total fatty acids and storage lipids than their parental strain upon nitrogen (N) depletion. Transcriptome analysis revealed that genes involved in lipid metabolism are highly enriched in the wild-type but not in the myb1-1 mutant after 4 h of N depletion. Among these genes were several involved in the transport of fatty acids from the chloroplast to the endoplasmic reticulum (ER): acyl-ACP thioesterase (FAT1), Fatty Acid EXporters (FAX1, FAX2), and long-chain acyl-CoA synthetase1 (LACS1). Furthermore, overexpression of FAT1 in the chloroplast increased lipid production. These results suggest that, upon N depletion, MYB1 promotes lipid accumulation by facilitating fatty acid transport from the chloroplast to the ER. This study identifies MYB1 as an important positive regulator of lipid accumulation in C. reinhardtii upon N depletion, adding another player to the established regulators of this process, including NITROGEN RESPONSE REGULATOR 1 (NRR1) and TRIACYLGLYCEROL ACCUMULATION REGULATOR 1 (TAR1).

Marine bacteria-mediated abiotic-biotic coupling degradation mechanism of ibuprofen

Knowledge on the behavior and fate of pharmaceuticals and personal care products (PPCPs) is poorly explored in marine aphotic environment. In this study, the degradation mechanism of a typical PPCPs—ibuprofen (IBP) by a ubiquitous marine Pseudoalteromonas sp. was investigated based on transcriptome and key enzymes analysis. More importantly, a novel enzymatic-nonenzymatic coupling degradation mechanism was uncovered for the first time, namely, the degradation of IBP was firstly initiated by extracellular reactive oxygen species (ROS), then the intermediate (e.g.4-ethylresorcinol) was further degraded by intracellular enzymes. It was showed that biogenic •OH, O2•−and H2O2 were responsible for extracellular nonenzymatic degradation, in which IBP was degraded to 4-ethylresorcinol through hydrogenation, isobutyl moiety cleavage, oxidation and decarboxylation. 4-Hydroxyphenylpyruvate dioxygenase, homogentisate 1,2-dioxygenase, long-chain acyl-CoA synthetase, acetyl-CoA acyltransferase and enoyl-CoA hydratase were identified to be involved in intracellular degradation, leading 4-ethylresorcinol cracked and eventually mineralized. Ultimately, this novel degradation mechanism was demonstrated to be amino acids-driven through KEGG enrichment analysis and experimental data. Overall, our work uncovered a yet undiscovered abiotic-biotic coupling degradation mechanism in PPCPs biotransformation, thereby updating the conventional concept that contaminants transformation is solely accomplished by enzymes or non-enzymes, which can also provide new insights into PPCPs environmental behavior and fate.

Glucose Starvation or Pyruvate Dehydrogenase Activation Induce a Broad, ERK5-Mediated, Metabolic Remodeling Leading to Fatty Acid Oxidation.

Cells have metabolic flexibility that allows them to adapt to changes in substrate availability. Two highly relevant metabolites are glucose and fatty acids (FA), and hence, glycolysis and fatty acid oxidation (FAO) are key metabolic pathways leading to energy production. Both pathways affect each other, and in the absence of one substrate, metabolic flexibility allows cells to maintain sufficient energy production. Here, we show that glucose starvation or sustained pyruvate dehydrogenase (PDH) activation by dichloroacetate (DCA) induce large genetic remodeling to propel FAO. The extracellular signal-regulated kinase 5 (ERK5) is a key effector of this multistep metabolic remodeling. First, there is an increase in the lipid transport by expression of low-density lipoprotein receptor-related proteins (LRP), e.g., CD36, LRP1 and others. Second, an increase in the expression of members of the acyl-CoA synthetase long-chain (ACSL) family activates FA. Finally, the expression of the enzymes that catalyze the initial step in each cycle of FAO, i.e., the acyl-CoA dehydrogenases (ACADs), is induced. All of these pathways lead to enhanced cellular FAO. In summary, we show here that different families of enzymes, which are essential to perform FAO, are regulated by the signaling pathway, i.e., MEK5/ERK5, which transduces changes from the environment to genetic adaptations.

Roles of ERF2 in apple fruit cuticular wax synthesis

Ethylene response factors 2 (ERF2) belongs to the AP2/ERF transcription factor family and plays key roles in ethylene signal transduction. The roles of MdERF2 in apple (Malus × domestica) cuticular wax synthesis regulated by ethylene were studied in this work. Result shows that MdERF2 overexpression (ERF2-OE) up-regulated the expression levels of wax-related synthetic genes long-chain acyl-CoA synthetase 2 (MdLACS2), Wax synthase1 (MdWSD1), Eceriferum 4 (MdCER4) and Eceriferum 6 (MdCER6), while has no effect on the transcription of Eceriferum 1 (MdCER1), and shows different regulation effects on the expression levels of mid-chain alkane hydroxylase 1 (MdMAH1) between calli and fruit. Furthermore, different MdERF2 expression levels changed the apparent structure of the apple cuticular wax and positively regulated the total wax accumulation. ERF2-OE also increased alkane and alcohol production but decreased acid and ester contents. Thus, ethylene regulated the synthesis of apple cuticular wax at the transcriptional level through MdERF2, thereby altering the wax structure and composition.

Fatty acid transport protein 2 interacts with ceramide synthase 2 to promote ceramide synthesis

Dihydroceramide is a lipid molecule generated via the action of (dihydro)ceramide synthases (CerSs), which use two substrates, namely sphinganine and fatty acyl-CoAs. Sphinganine is generated via the sequential activity of two integral membrane proteins located in the endoplasmic reticulum. Less is known about the source of the fatty acyl-CoAs, although a number of cytosolic proteins in the pathways of acyl-CoA generation modulate ceramide synthesis via direct or indirect interaction with the CerSs. In this study, we demonstrate, by proteomic analysis of immunoprecipitated proteins, that fatty acid transporter protein 2 (FATP2) (also known as very long-chain acyl-CoA synthetase) directly interacts with CerS2 in mouse liver. Studies in cultured cells demonstrated that other members of the FATP family can also interact with CerS2, with the interaction dependent on both proteins being catalytically active. In addition, transfection of cells with FATP1, FATP2, or FATP4 increased ceramide levels although only FATP2 and 4 increased dihydroceramide levels, consistent with their known intracellular locations. Finally, we show that lipofermata, an FATP2 inhibitor which is believed to directly impact tumor cell growth via modulation of FATP2, decreased de novo dihydroceramide synthesis, suggesting that some of the proposed therapeutic effects of lipofermata may be mediated via (dihydro)ceramide rather than directly via acyl-CoA generation. In summary, our study reinforces the idea that manipulating the pathway of fatty acyl-CoA generation will impact a wide variety of down-stream lipids, not least the sphingolipids, which utilize two acyl-CoA moieties in the initial steps of their synthesis.

Adoptive transfer of metabolically reprogrammed macrophages for atherosclerosis treatment in diabetic ApoE−/- mice

Atherosclerosis is characterized by inflammation in the arterial wall, which is known to be exacerbated by diabetes. Therapeutic repression of inflammation is a promising strategy for treating atherosclerosis. In this study, we showed that diabetes aggravated atherosclerosis in apolipoproteinE knockout (ApoE−/-) mice, in which increased expression of long-chain acyl-CoA synthetase 1 (Acsl1) in macrophages played an important role. Knockdown of Acsl1 in macrophages (MφshAcsl1) reprogrammed macrophages to an anti-inflammatory phenotype, especially under hyperglycemic conditions. Injection of MφshAcsl1 reprogrammed macrophages into streptozotocin (STZ)-induced diabetic ApoE−/- mice (ApoE−/-+ STZ) alleviated inflammation locally in the plaque, liver and spleen. Consistent with the reduction in inflammation, plaques became smaller and more stable after the adoptive transfer of reprogrammed macrophages. Taken together, our findings indicate that increased Acsl1 expression in macrophages play a key role in aggravated atherosclerosis of diabetic mice, possibly by promoting inflammation. Adoptive transfer of Acsl1 silenced macrophages may serve as a potential therapeutic strategy for atherosclerosis.

Elevated thyroid hormones caused by high concentrate diets participate in hepatic metabolic disorders in dairy cows.

ObjectiveHigh concentrate diets are widely used to satisfy high-yielding dairy cows; however, long-term feeding of high concentrate diets can cause subacute ruminal acidosis (SARA). The endocrine disturbance is one of the important reasons for metabolic disorders caused by SARA. However, there is no current report about thyroid hormones involved in liver metabolic disorders induced by a high concentrate diet.MethodsIn this study, 12 mid-lactating dairy cows were randomly assigned to HC (high concentrate) group (60% concentrate of dry matter, n = 6) and LC (low concentrate) group (40% concentrate of dry matter, n = 6). All cows were slaughtered on the 21st day, and the samples of blood and liver were collected to analyze the blood biochemistry, histological changes, thyroid hormones, and the expression of genes and proteins.ResultsCompared with LC group, HC group showed decreased serum triglyceride, free fatty acid, total cholesterol, low-density lipoprotein cholesterol, increased hepatic glycogen, and glucose. For glucose metabolism, the gene and protein expression of glucose-6-phosphatase and phosphoenolpyruvate carboxykinase 1 in the liver were significantly up-regulated in HC group. For lipid metabolism, the expression of sterol regulatory element-binding protein 1, long-chain acyl-CoA synthetase 1, and fatty acid synthase in the liver was decreased in HC group, whereas carnitine palmitoyltransferase 1α and peroxisome proliferator activated receptor α were increased. Serum triiodothyronine, thyroxin, free triiodothyronine (FT3), and hepatic FT3 increased in HC group, accompanied by increased expression of thyroid hormone receptor (THR) in the liver.ConclusionTaken together, thyroid hormones may increase hepatic gluconeogenesis, β-oxidation and reduce fatty acid synthesis through the THR pathway to participate in the metabolic disorders caused by a high concentrate diet.

Comparative effects of dietary soybean oil and fish oil on the growth performance, fatty acid composition and lipid metabolic signaling of grass carp, Ctenopharyngodon idella

Lipid was an essential constituent of aquatic feeds, among which fish oil (FO) was the most excellent lipid source for its nutritive values. However, high price and limited production restricted its application, thus leading to the consideration of developing other alternative lipid sources, and soybean oil (SO) instead of FO was applied as the main lipid source in the commercial feed of grass carp. In the present study, two typical lipids sources FO and SO were used to prepare grass carp feed, and their comparative effects on the growth performance, nutrient composition and metabolic signaling were investigated. The growth performance of two groups exhibited no significant difference, while crude lipid was much higher in the muscle of SO group. Additionally, muscle fatty acid composition of both groups were highly correlated with that of diets, and the proportion of muscle n-3 polyunsaturated fatty acids was notably higher in the FO group, especially that of C20:5n-3 (EPA) and C22:6n-3 (DHA). Comparative digital gene expression profiles (DGE) was conducted and 1261 differentially expressed genes (DEGs) had been identified, then gene ontology (GO) enrichment analysis revealed the enriched functional subterms were primarily focused on cellular metabolic processes and regulation. KEGG pathway analysis elucidated the main metabolism related pathways, and DEGs involved in fatty acid elongation, fatty acid metabolism, and glycerolipid metabolism pathways were listed, including elongation of very long chain fatty acids protein 7 (el7), lysophospholipid acyltransferase 4 (lpcat4), long-chain acyl-CoA synthetase (acs), phosphatidate phosphatase LPIN (lpin), cathepsin L, and glucose-6-phosphate isomerase (gpi) et al. In conclusion, the nutritional composition rather than growth performance of grass carp was affected by different lipid sources, and the underlying regulatory signaling was predicted, which would provide theoretical basis for fish diet formulation.

Glycolytic and oxidative muscles under acute glucose supplementation differ in their metabolic responses to fatty acyl-CoA synthetase gene suppression

Glycolytic and oxidative muscles under acute glucose supplementation differ in their metabolic responses to fatty acyl-CoA synthetase gene suppression

Multiplexed CRISPR/Cas9 editing of the long-chain acyl-CoA synthetase family in the diatom Phaeodactylum tricornutum reveals that mitochondrial ptACSL3 is involved in the synthesis of storage lipids.

Long-chain acyl-CoA synthetases (LACS) play diverse and fundamentally important roles in lipid metabolism. While their functions have been well established in bacteria, yeast and plants, the mechanisms by which LACS isozymes regulate lipid metabolism in unicellular oil-producing microalgae, including the diatom Phaeodactylum tricornutum, remain largely unknown. In P. tricornutum, a family of five genes (ptACSL1-ptACSL5) encodes LACS activities. We generated single lacs knockout/knockdown mutants using multiplexed CRISPR/Cas9 method, and determined their substrate specificities towards different fatty acids (FAs) and subcellular localisations. ptACSL3 is localised in the mitochondria and its disruption led to compromised growth and reduced triacylglycerol (TAG) content when cells were bubbled with air. The ptACSL3 mutants showed altered FA profiles in two galactoglycerolipids and phosphatidylcholine (PC) with significantly reduced distribution of 16:0 and 16:1. ptACSL5 is localised in the peroxisome and its knockdown resulted in reduced growth rate and altered molecular species of PC and TAG, indicating a role in controlling the composition of acyl-CoAs for lipid synthesis. Our work demonstrates the potential of generating gene knockout mutants with the mutation of large fragment deletion using multiplexed CRISPR/Cas9 and provides insight into the functions of LACS isozymes in lipid metabolism in the oleaginous microalgae.

Combined berberine and probiotic treatment as an effective regimen for improving postprandial hyperlipidemia in type 2 diabetes patients: a double blinded placebo controlled randomized study

ABSTRACT Non-fasting lipidemia (nFL), mainly contributed by postprandial lipidemia (PL), has recently been recognized as an important cardiovascular disease (CVD) risk as fasting lipidemia (FL). PL serves as a common feature of dyslipidemia in Type 2 Diabetes (T2D), albeit effective therapies targeting on PL were limited. In this study, we aimed to evaluate whether the therapy combining probiotics (Prob) and berberine (BBR), a proven antidiabetic and hypolipidemic regimen via altering gut microbiome, could effectively reduce PL in T2D and to explore the underlying mechanism. Blood PL (120 min after taking 100 g standard carbohydrate meal) was examined in 365 participants with T2D from the Probiotics and BBR on the Efficacy and Change of Gut Microbiota in Patients with Newly Diagnosed Type 2 Diabetes (PREMOTE study), a random, placebo-controlled, and multicenter clinical trial. Prob+BBR was superior to BBR or Prob alone in improving postprandial total cholesterol (pTC) and low-density lipoprotein cholesterol (pLDLc) levels with decrement of multiple species of postprandial lipidomic metabolites after 3 months follow-up. This effect was linked to the changes of fecal Bifidobacterium breve level responding to BBR alone or Prob+BBR treatment. Four fadD genes encoding long-chain acyl-CoA synthetase were identified in the genome of this B. breve strain, and transcriptionally activated by BBR. In vitro BBR treatment further decreased the concentration of FFA in the culture medium of B. breve compared to vehicle. Thus, the activation of fadD by BBR could enhance FFA import and mobilization in B. breve and diliminish the intraluminal lipids for absorption to mediate the effect of Prob+BBR on PL. Our study confirmed that BBR and Prob (B. breve) could exert a synergistic hypolipidemic effect on PL, acting as a gut lipid sink to achieve better lipidemia and CVD risk control in T2D.

Interference With ACSL1 Gene in Bovine Adipocytes: Transcriptome Profiling of mRNA and lncRNA Related to Unsaturated Fatty Acid Synthesis.

The enzyme long-chain acyl-CoA synthetase 1 (ACSL1) is essential for lipid metabolism. The ACSL1 gene controls unsaturated fatty acid (UFA) synthesis as well as the formation of lipid droplets in bovine adipocytes. Here, we used RNA-Seq to determine lncRNA and mRNA that regulate UFA synthesis in bovine adipocytes using RNA interference and non-interference with ACSL1. The corresponding target genes of differentially expressed (DE) lncRNAs and the DE mRNAs were found to be enriched in lipid and FA metabolism-related pathways, according to GO and KEGG analyses. The differentially expressed lncRNA- differentially expressed mRNA (DEL-DEM) interaction network indicated that some DELs, such as TCONS_00069661, TCONS_00040771, TCONS_ 00035606, TCONS_00048301, TCONS_001309018, and TCONS_00122946, were critical for UFA synthesis. These findings assist our understanding of the regulation of UFA synthesis by lncRNAs and mRNAs in bovine adipocytes.