Nordic cyanobacterial and algal lipids: TAG accumulation, chemotaxonomy and bioindustrial potential

Fatty liver is closely associated with elevated concentrations of nonesterified fatty acids (NEFA) and a low level of very low-density lipoproteins (VLDL) in blood of dairy cows. High NEFA inhibit the VLDL synthesis and assembly, and cause hepatic triacylglycerol (TAG) deposition. Sirtuin 3 (SIRT3), a mitochondrial deacetylase, antagonizes NEFA-induced TAG accumulation through modulating expressions of fatty acid synthesis and oxidation genes in cow hepatocytes. However, the role of SIRT3 in the VLDL synthesis and assembly was largely unknown. Here we aimed to test whether SIRT3 would recover the synthesis and assembly of VLDL in cow hepatocytes induced by high NEFA. Primary cow hepatocytes were isolated from 3 Holstein cows. Hepatocytes were infected with SIRT3 overexpression adenovirus (Ad-SIRT3), SIRT3-short interfering (si) RNA, or first infected with Ad-SIRT3 and then incubated with 1.0 mM NEFA (Ad-SIRT3 + NEFA). Expressions of key genes in VLDL synthesis and the VLDL contents in cell culture supernatants were measured. SIRT3 overexpression significantly increased the mRNA abundance of microsomal triglyceride transfer protein (MTP), apolipoprotein B100 (ApoB100) and ApoE (p < 0.01), and raised VLDL contents in the supernatants (p < 0.01). However, SIRT3 silencing displayed a reverse effect in comparison to SIRT3 overexpression. Compared with NEFA treatment alone, the Ad-SIRT3 + NEFA significantly upregulated the mRNA abundance of MTP, ApoB100 and ApoE (p < 0.01), and increased VLDL contents in the supernatants (p < 0.01). Our data demonstrated that SIRT3 restored the synthesis and assembly of VLDL in cow hepatocytes challenged with NEFA, providing an in vitro basis for further investigations testing its feasibility against hepatic TAG accumulation in dairy cows during the perinatal period.

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Plastidial ACP Δ9‐desaturase modulates EPA biosynthesis and TAG accumulation in Phaeodactylum tricornutum

The Plant Journal ◽

10.1111/tpj.15231 ◽

2021 ◽

Author(s):

Richard Smith ◽

Juliette Jouhet ◽

Chiara Gandini ◽

Vladimir Nekrasov ◽

Eric Marechal ◽

...

Keyword(s):

Phaeodactylum Tricornutum ◽

Δ9 Desaturase ◽

Tag Accumulation

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Engineering of Saccharomyces cerevisiae for the accumulation of high amounts of triacylglycerol

Microbial Cell Factories ◽

10.1186/s12934-021-01640-0 ◽

2021 ◽

Vol 20 (1) ◽

Author(s):

Simon Arhar ◽

Gabriela Gogg-Fassolter ◽

Mojca Ogrizović ◽

Klavdija Pačnik ◽

Katharina Schwaiger ◽

...

Keyword(s):

Saccharomyces Cerevisiae ◽

Fatty Acid ◽

Lipid Level ◽

Pharmaceutical Products ◽

Acetyl Coa Carboxylase ◽

Oleaginous Yeasts ◽

Regulatory Subunits ◽

Gene Coding ◽

Tag Accumulation ◽

Competing Pathways

Abstract Background Fatty acid-based substances play an important role in many products, from food supplements to pharmaceutical products and biofuels. The production of fatty acids, mainly in their esterified form as triacylglycerol (TAG), has been intensively studied in oleaginous yeasts, whereas much less effort has been invested into non-oleaginous species. In the present work, we engineered the model yeast Saccharomyces cerevisiae, which is commonly regarded as non-oleaginous, for the storage of high amounts of TAG, comparable to the contents achieved in oleaginous yeasts. Results We investigated the effects of several mutations with regard to increased TAG accumulation and identified six of them as important for this phenotype: a point mutation in the acetyl-CoA carboxylase Acc1p, overexpression of the diacylglycerol acyltransferase Dga1p, deletions of genes coding for enzymes involved in the competing pathways glycogen and steryl ester synthesis and TAG hydrolysis, and a deletion of CKB1, the gene coding for one of the regulatory subunits of casein kinase 2. With the combination of these mutations in a S. cerevisiae strain with a relatively high neutral lipid level already in the non-engineered state, we achieved a TAG content of 65% in the dry biomass. High TAG levels were not only obtained under conditions that favor lipid accumulation, but also in defined standard carbon-limited media. Conclusions Baker's yeast, which is usually regarded as inefficient in the storage of TAG, can be converted into a highly oleaginous strain that could be useful in processes aiming at the synthesis of fatty acid-based products. This work emphasizes the importance of strain selection in combination with metabolic engineering to obtain high product levels.

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Alteration of serum amino acid profiles by dietary adenine supplementation inhibits fatty liver development in rats

Scientific Reports ◽

10.1038/s41598-020-79234-w ◽

2020 ◽

Vol 10 (1) ◽

Author(s):

Hiroki Nishi ◽

Daisuke Yamanaka ◽

Masato Masuda ◽

Yuki Goda ◽

Koichi Ito ◽

...

Keyword(s):

Amino Acid ◽

Fatty Liver ◽

Causative Factor ◽

Amino Acid Profile ◽

Liver Development ◽

Concentration Data ◽

Amino Acid Profiles ◽

Arginine Deficiency ◽

Branched Chain ◽

Tag Accumulation

AbstractStudies on animal models have demonstrated that feeding a low-arginine diet inhibits triacylglycerol (TAG) secretion from the liver, resulting in marked fatty liver development in rats. Here, we first showed that culturing hepatocytes in the medium mimicking the serum amino acid profile of low-arginine diet-fed rats induced TAG accumulation in the cells, indicating that the specific amino acid profile caused TAG accumulation in hepatocytes. Dietary adenine supplementation completely recovered hepatic TAG secretion and abolished hepatic TAG accumulation in rats. A comprehensive non-linear analysis revealed that inhibition of hepatic TAG accumulation by dietary adenine supplementation could be predicted using only serum amino acid concentration data. Comparison of serum amino acid concentrations indicated that histidine, methionine, and branched-chain amino acid (BCAA) concentrations were altered by adenine supplementation. Furthermore, when the serum amino acid profiles of low-arginine diet-fed rats were altered by modifying methionine or BCAA concentrations in their diets, their hepatic TAG accumulation was abolished. Altogether, these results suggest that an increase in methionine and BCAA levels in the serum in response to dietary arginine deficiency is a key causative factor for hepatic TAG accumulation, and dietary adenine supplementation could disrupt this phenomenon by altering serum amino acid profiles.

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Characterisation of DGAT1 and DGAT2 from Jatropha curcas and their functions in storage lipid biosynthesis

Functional Plant Biology ◽

10.1071/fp12388 ◽

2014 ◽

Vol 41 (3) ◽

pp. 321 ◽

Cited By ~ 11

Author(s):

Ronghua Xu ◽

Tianquan Yang ◽

Ruling Wang ◽

Aizhong Liu

Keyword(s):

Linoleic Acid ◽

Jatropha Curcas ◽

Molecular Mechanisms ◽

Functional Divergence ◽

Critical Role ◽

Linoleic Acid Content ◽

Transgenic Yeast ◽

Kennedy Pathway ◽

Tag Biosynthesis ◽

Tag Accumulation

Diacylglycerol acyltransferases (DGATs) catalyse the final step of triacylglycerol (TAG) biosynthesis of the Kennedy pathway, and play a critical role during TAG accumulation in developing oleaginous seeds. In this study, the molecular cloning and characterisation of two DGAT genes, JcDGAT1 and JcDGAT2, from jatropha (Jatropha curcas L., a potential biodiesel plant) is presented. Using heterogonous overexpression techniques, both JcDGAT1 and JcDGAT2 were able to restore TAG biosynthesis in a yeast mutant H1246 strain, and enhance the quantity of TAG biosynthesis by 16.6 and 14.3%, respectively, in strain INVSc1. In transgenic tobacco, overexpression of JcDGAT1 and JcDGAT2 resulted in an increase in seed oil content of, respectively, 32.8 and 31.8%. Further, the functional divergence of JcDGAT1 and JcDGAT2 in TAG biosynthesis was demonstrated by comparing the fatty acid compositions in both the transgenic yeast and tobacco systems. In particular, JcDGAT2 incorporated a 2.5-fold higher linoleic acid content into TAG than JcDGAT1 in transgenic yeast and exhibited a significant linoleic acid substrate preference in both yeast and tobacco. This study provides new insights in understanding the molecular mechanisms of DGAT genes underlying the biosynthesis of linoleic acids and TAG in plants.

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The Rhodococcus opacus PD630 Heparin-Binding Hemagglutinin Homolog TadA Mediates Lipid Body Formation

Applied and Environmental Microbiology ◽

10.1128/aem.00985-10 ◽

2010 ◽

Vol 76 (21) ◽

pp. 7217-7225 ◽

Cited By ~ 36

Author(s):

Daniel P. MacEachran ◽

M. E. Prophete ◽

A. J. Sinskey

Keyword(s):

Kinetic Studies ◽

Lipid Body ◽

Dry Weight ◽

Rhodococcus Opacus ◽

Lipid Bodies ◽

Heparin Binding ◽

Tag Biosynthesis ◽

Tag Accumulation

ABSTRACT Generally, prokaryotes store carbon as polyhydroxyalkanoate, starch, or glycogen. The Gram-positive actinomycete Rhodococcus opacus strain PD630 is noteworthy in that it stores carbon in the form of triacylglycerol (TAG). Several studies have demonstrated that R. opacus PD630 can accumulate up to 76% of its cell dry weight as TAG when grown under nitrogen-limiting conditions. While this process is well studied, the underlying molecular and biochemical mechanisms leading to TAG biosynthesis and subsequent storage are poorly understood. We designed a high-throughput genetic screening to identify genes and their products required for TAG biosynthesis and storage in R. opacus PD630. We identified a gene predicted to encode a putative heparin-binding hemagglutinin homolog, which we have termed tadA (triacylglycerol accumulation deficient), as being important for TAG accumulation. Kinetic studies of TAG accumulation in both the wild-type (WT) and mutant strains demonstrated that the tadA mutant accumulates 30 to 40% less TAG than the parental strain (WT). We observed that lipid bodies formed by the mutant strain were of a different size and shape than those of the WT. Characterization of TadA demonstrated that the protein is capable of binding heparin and of agglutinating purified lipid bodies. Finally, we observed that the TadA protein localizes to lipid bodies in R. opacus PD630 both in vivo and in vitro. Based on these data, we hypothesize that the TadA protein acts to aggregate small lipid bodies, found in cells during early stages of lipid storage, into larger lipid bodies and thus plays a key role in lipid body maturation in R. opacus PD630.

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Human GDPD3 overexpression promotes liver steatosis by increasing lysophosphatidic acid production and fatty acid uptake

The Journal of Lipid Research ◽

10.1194/jlr.ra120000760 ◽

2020 ◽

Vol 61 (7) ◽

pp. 1075-1086 ◽

Cited By ~ 1

Author(s):

Chia-Chi C. Key ◽

Andrew C. Bishop ◽

Xianfeng Wang ◽

Qingxia Zhao ◽

Guan-yuan Chen ◽

...

Keyword(s):

Hepatic Steatosis ◽

Liver Steatosis ◽

Fatty Acid Uptake ◽

Western Diet ◽

Acid Uptake ◽

Mrna Levels ◽

Glycerol Phosphate ◽

Adeno Associated Virus ◽

Mouse Hepatocytes ◽

Tag Accumulation

The glycerol phosphate pathway produces more than 90% of the liver triacylglycerol (TAG). LysoPA, an intermediate in this pathway, is produced by glycerol-3-phosphate acyltransferase. Glycerophosphodiester phosphodiesterase domain containing 3 (GDPD3), whose gene was recently cloned, contains lysophospholipase D activity, which produces LysoPA from lysophospholipids. Whether human GDPD3 plays a role in hepatic TAG homeostasis is unknown. We hypothesized that human GDPD3 increases LysoPA production and availability in the glycerol phosphate pathway, promoting TAG biosynthesis. To test our hypothesis, we infected C57BL/6J mice with adeno-associated virus encoding a hepatocyte-specific albumin promoter that drives GFP (control) or FLAG-tagged human GDPD3 overexpression and fed the mice chow or a Western diet to induce hepatosteatosis. Hepatic human GDPD3 overexpression induced LysoPA production and increased FA uptake and incorporation into TAG in mouse hepatocytes and livers, ultimately exacerbating Western diet-induced liver steatosis. Our results also showed that individuals with hepatic steatosis have increased GDPD3 mRNA levels compared with individuals without steatosis. Collectively, these findings indicate that upregulation of GDPD3 expression may play a key role in hepatic TAG accumulation and may represent a molecular target for managing hepatic steatosis.

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Sirtuin 1 is involved in oleic acid-induced calf hepatocyte steatosis via alterations in lipid metabolism-related proteins

Journal of Animal Science ◽

10.1093/jas/skab250 ◽

2021 ◽

Vol 99 (10) ◽

Author(s):

Hongyan Ding ◽

Yu Li ◽

Leihong Liu ◽

Ning Hao ◽

Suping Zou ◽

...

Keyword(s):

Lipid Metabolism ◽

Mrna Abundance ◽

Sirtuin 1 ◽

Peroxisome Proliferator ◽

Acetyl Coa Carboxylase ◽

Peroxisome Proliferator Activated Receptor ◽

Lower Protein ◽

Tag Accumulation

Abstract Sirtuin 1 (SIRT1), an NAD-dependent protein deacetylase, plays a central role in the control of lipid metabolism in nonruminants. However, the role of SIRT1 in hepatic lipid metabolism in dairy cows with fatty liver is not well known. Thus, we used isolated primary bovine hepatocytes to determine the role of SIRT1 in protecting cells against oleic acid (OA)-induced steatosis. Recombinant adenoviruses to overexpress (AD-GFP-SIRT1-E) or knockdown (AD-GFP-SIRT1-N) SIRT1 were used for transduction of hepatocytes. Calf hepatocytes isolated from five female calves (1 d old, 30 to 40 kg) were used to determine both time required and the lowest dose of OA that could induce triacylglycerol (TAG) accumulation. Analyses indicated that 0.25 mM OA for 24 h was suitable to induce TAG accumulation. In addition, OA not only led to an increase in TAG, but also upregulated mRNA and protein abundance of sterol regulatory element-binding transcription factor 1 (SREBF1) and downregulated SIRT1 and peroxisome proliferator-activated receptor-gamma coactivator 1 alpha (PPARGC1A). Thus, these in vitro conditions were deemed optimal for subsequent experiments. Calf hepatocytes were cultured and incubated with OA (0.25 mM) for 24 h, followed by adenoviral AD-GFP-SIRT1-E or AD-GFP-SIRT1-N transduction for 48 h. Overexpression of SIRT1 led to greater protein and mRNA abundance of SIRT1 along with fatty acid oxidation-related genes including PPARGC1A, peroxisome proliferator-activated receptor alpha (PPARA), retinoid X receptor α (RXRA), and ratio of phospho-acetyl-CoA carboxylase alpha (p-ACACA)/total acetyl-CoA carboxylase alpha (ACACA). In contrast, it resulted in lower protein and mRNA abundance of genes related to lipid synthesis including SREBF1, fatty acid synthase (FASN), apolipoprotein E (APOE), and low-density lipoprotein receptor (LDLR). The concentration of TAG decreased due to SIRT1 overexpression. In contrast, silencing SIRT1 led to lower protein and mRNA abundance of SIRT1, PPARGC1A, PPARA, RXRA, and greater protein and mRNA abundance of SREBF1, FASN, APOE, and LDLR. Further, those responses were accompanied by greater content of cellular TAG and total cholesterol (TC). Overall, data from these in vitro studies indicated that SIRT1 is involved in the regulation of lipid metabolism in calf hepatocytes subjected to an increase in the supply of OA. Thus, it is possible that alterations in SIRT1 abundance and activity in vivo contribute to development of fatty liver in dairy cows.

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Effects of B vitamins and methyl group donors on milk production, milk composition and blood biochemistry in dairy cows

Proceedings of the British Society of Animal Science ◽

10.1017/s1752756200008528 ◽

2002 ◽

Vol 2002 ◽

pp. 196-196

Author(s):

S.E Richards ◽

S Hicklin ◽

T Lord ◽

A Nickson ◽

J Long ◽

...

Keyword(s):

Milk Yield ◽

Milk Composition ◽

Metabolic Fate ◽

Dry Period ◽

Very Low Density Lipoproteins ◽

Methyl Group ◽

Fat Reserves ◽

Esterified Fatty Acids ◽

Pregnancy And Lactation ◽

Tag Accumulation

Recent reviews highlight the importance of the liver in the coordination of nutrient fluxes in support of pregnancy and lactation (e.g. Drackley et al., 2001). Mobilisation of body fat reserves in the late dry period and early lactation leads to an increase in uptake of non-esterified fatty acids (NEFA) by the liver. Their metabolic fate is either oxidation or esterification into triacylglycerides (TAG) that are either exported in very low density lipoproteins (VLDL) or accumulated within liver cells. Recent evidence indicates that TAG accumulation impairs ureagenic and gluconeogenic capacity of the liver, with consequent reductions in feed intake and milk yield, increased incidence of disease and decreased reproductive performance (Overton and Piepenbrink, 1999).LiFTTM (NuTec Ltd.) is a proprietary mixture of B-group vitamins and methyl group donors (rumen protected choline, niacin, vitamin B12, biotin, folic acid and thiamine) designed to reduce the accumulation of TAG in the liver and accelerate VLDL export. The objective of this experiment was to evaluate the effect of LiFT on milk yield and composition and concentrations of metabolites in blood.

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Lipid Production in Nannochloropsis gaditana during Nitrogen Starvation

Biology ◽

10.3390/biology8010005 ◽

2019 ◽

Vol 8 (1) ◽

pp. 5 ◽

Cited By ~ 6

Author(s):

Jorijn H. Janssen ◽

René H. Wijffels ◽

Maria J. Barbosa

Keyword(s):

Nitrogen Starvation ◽

De Novo ◽

Lipid Production ◽

Omega 3 ◽

Supply Rate ◽

Nannochloropsis Gaditana ◽

Omega 3 Fatty Acid ◽

Light Regimes ◽

Lipid Fractions ◽

Tag Accumulation

The microalga Nannochloropsis gaditana is a natural producer of triacylglycerol (TAG) and the omega-3 fatty acid eicosapentaenoic acid (EPA). TAG accumulation is induced by nitrogen starvation. The biomass-specific photon supply rate used had an effect on EPA and TAG accumulation during nitrogen starvation as well as on the localization of EPA accumulation. Clear differences in TAG yield on light were found for different biomass-specific photon supply rates and light regimes during nitrogen starvation. De novo EPA synthesis or the translocation of EPA between lipid fractions might be limiting for EPA accumulation in TAG. Further studies are needed to fully understand EPA accumulation in TAG during nitrogen starvation. To elucidate the function of EPA in TAG nitrogen recovery, experiments are suggested. The overexpression of genes involved in de novo EPA synthesis and translocation is proposed to elucidate the exact metabolic routes involved in these processes during nitrogen starvation. This work addresses future opportunities to increase EPA accumulation.

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