Glycerol-3-phosphate Dehydrogenase Research Articles

Effects of stocking density on antioxidant status, metabolism and immune response in juvenile turbot (Scophthalmus maximus)

This study was designed to evaluate the physiological and immune responses of juvenile turbot to stocking density. Turbot (average weight 185.4g) were reared for 120days in a land based recirculating aquaculture system (RAS) under three stocking densities: low density (LD, ~9.3–26.1kg/m2, initial to final density), medium density (MD, ~13.6–38.2kg/m2) and high density (HD, ~19.1–52.3kg/m2). Fish were sampled at days 0, 40, 80 and 120 to obtain growth parameters and liver tissues. No significant difference was detected in growth, biochemical parameters and gene expression among the three densities until at the final sampling (day 120). At the end of this trial, fish reared in HD group showed lower specific growth rate (SGR) and mean weight than those reared in LD and MD groups. Similarly, oxidative stress and metabolism analyses represented that antioxidants (superoxide dismutase (SOD), catalase (CAT), glutathione (GSH)) and metabolic enzymes (glycerol-3-phosphate dehydrogenase (G3PDH) and glucose-6-phosphate dehydrogenase (G6PDH)) clearly reduced in the liver of turbot reared in HD group. The gene expression data showed that glutathione S-transferase (GST), cytochrome P450 1A (CYP1A), heat shock protein 70 (HSP 70) and metallothionein (MT) mRNA levels were significantly up-regulated, and lysozyme (LYS) and hepcidin (HAMP) mRNA levels were significantly down-regulated in HD group on day 120. Overall, our results indicate that overly high stocking density might block the activities of metabolic and antioxidant enzymes, and cause physiological stress and immunosuppression in turbot.

Ataxin-1 regulates the cerebellar bioenergetics proteome through the GSK3β-mTOR pathway which is altered in Spinocerebellar ataxia type 1 (SCA1).

A polyglutamine expansion within the ataxin-1 protein (ATXN1) underlies spinocerebellar ataxia type-1 (SCA1), a neurological disorder mainly characterized by ataxia and cerebellar deficits. In SCA1, both loss and gain of ATXN1 biological functions contribute to cerebellar pathogenesis. However, the critical ATXN1 functions and pathways involved remain unclear. To further investigate the early signalling pathways regulated by ATXN1, we performed an unbiased proteomic study of the Atxn1-KO 5-week-old mice cerebellum. Here, we show that lack of ATXN1 expression induces early alterations in proteins involved in glycolysis [pyruvate kinase, muscle, isoform 1 protein (PKM-i1), citrate synthase (CS), glycerol-3-phosphate dehydrogenase 2 (GPD2), glucose-6-phosphate isomerase (GPI), alpha -: enolase (ENO1)], ATP synthesis [CS, Succinate dehydrogenase complex,subunit A (SDHA), ATP synthase subunit d, mitochondrial (ATP5H)] and oxidative stress [peroxiredoxin-6 (PRDX6), aldehyde dehydrogenase family 1, subfamily A1, 10-formyltetrahydrofolate dehydrogenase]. In the SCA1 mice, several of these proteins (PKM-i1, ATP5H, PRDX6, proteome subunit A6) were down-regulated and ATP levels decreased. The underlying mechanism does not involve modulation of mitochondrial biogenesis, but dysregulation of the activity of the metabolic regulators glycogen synthase kinase 3B (GSK3β), decreased in Atxn1-KO and increased in SCA1 mice, and mechanistic target of rapamycin (serine/threonine kinase) (mTOR), unchanged in the Atxn1-KO and decreased in SCA1 mice cerebellum before the onset of ataxic symptoms. Pharmacological inhibition of GSK3β and activation of mTOR in a SCA1 cell model ameliorated identified ATXN1-regulated metabolic proteome and ATP alterations. Taken together, these results point to an early role of ATXN1 in the regulation of bioenergetics homeostasis in the mouse cerebellum. Moreover, data suggest GSK3β and mTOR pathways modulate this ATXN1 function in SCA1 pathogenesis that could be targeted therapeutically prior to the onset of disease symptoms in SCA1 and other pathologies involving dysregulation of ATXN1 functions.

Glucose-mediated regulation of glycerol uptake in Rhodosporidium toruloides: Insights through transcriptomic analysis on dual substrate fermentation.

Rhodosporidium toruloides is a robust oleaginous yeast that can accumulate lipids to more than 70% of its dry cell mass. Even though it is extensively studied for its fermentation of substrates like glucose and glycerol, limited information is available about its metabolism of mixture of glucose and glycerol. During growth on mixture of glucose and glycerol a typical diauxic growth and higher lipid yields were observed. To understand this phenomenon, RNA-seq analysis was implemented to study the gene expression profiles during growth on mixtures mainly to elucidate regulation of glycerol metabolism. Insights into lipid biosynthesis on mixed substrates are provided at a systems level. Among others, transcriptional profiles showed that glycerol might be produced intracellularly and glycerol kinase (GUT1) and glycerol 3-phosphate dehydrogenase (GUT2) enzymes were not downregulated in the presence of glucose. Transcriptional analysis also showed that the regulation of glycerol uptake in the presence of glucose at transcriptional level is different from that observed in Saccharomyces cerevisiae.

Overexpression of ACC gene from oleaginous yeast Lipomyces starkeyi enhanced the lipid accumulation in Saccharomyces cerevisiae with increased levels of glycerol 3-phosphate substrates.

The conversion of acetyl-CoA to malonyl-CoA by acetyl-CoA carboxylase (ACC) is the rate-limiting step in fatty acid biosynthesis. In this study, a gene coding for ACC was isolated and characterized from an oleaginous yeast, Lipomyces starkeyi. Real-time quantitative PCR (qPCR) analysis of L. starkeyi acetyl-CoA carboxylase gene (LsACC1) showed that the expression levels were upregulated with the fast accumulation of lipids. The LsACC1 was co-overexpressed with the glycerol 3-phosphate dehydrogenase gene (GPD1), which regulates lipids biosynthesis by supplying another substrates glycerol 3-phosphate for storage lipid assembly, in the non-oleaginous yeast Saccharomyces cerevisiae. Further, the S. cerevisiae acetyl-CoA carboxylase (ScACC1) was transferred with GPD1 and its function was analyzed in comparison with LsACC1. The results showed that overexpressed LsACC1 and GPD1 resulted in a 63% increase in S. cerevisiae. This study gives new data in understanding of the molecular mechanisms underlying the regulation of fatty acids and lipid biosynthesis in yeasts.

Germinated waxy black rice extract inhibits lipid accumulation with regulation of multiple gene expression in 3T3-L1 adipocytes.

The aim of this study was to investigate the anti-obesity effects of germinated waxy black rice (GWBR) extract in 3T3-L1 adipocytes. The inhibitory effect of GWBR extract against adipocyte differentiation was evaluated using Oil Red O staining and glycerol-3-phosphate dehydrogenase (GPDH) assay. GWBR extract inhibited adipocyte differentiation, but was not found to elicit any cytotoxicity. The mRNA levels of adipogenic transcriptional factors such as C/EBP-α and -β, PPAR-γ, and SREBP-1c, as well as adipogenic enzymes, including aP2, LPL, and FAS were significantly downregulated by treatment with GWBR extract compared to untreated control cells. However, mRNA levels of lipolytic genes such as HSL and ATGL, β-oxidation related genes CPT1, and UCP2 involved in thermogenesis were significantly up-regulated by treatment with GWBR extract. These data suggest that GWBR extract may be a potential functional food, and may have pharmacological applications in both the prevention and treatment of obesity.

Efficient production of acetoin in Saccharomyces cerevisiae by disruption of 2,3-butanediol dehydrogenase and expression of NADH oxidase.

Acetoin is widely used in food and cosmetic industry as taste and fragrance enhancer. For acetoin production in this study, Saccharomyces cerevisiae JHY605 was used as a host strain, where the production of ethanol and glycerol was largely eliminated by deleting five alcohol dehydrogenase genes (ADH1, ADH2, ADH3, ADH4, and ADH5) and two glycerol 3-phosphate dehydrogenase genes (GPD1 and GPD2). To improve acetoin production, acetoin biosynthetic genes from Bacillus subtilis encoding α-acetolactate synthase (AlsS) and α-acetolactate decarboxylase (AlsD) were overexpressed, and BDH1 encoding butanediol dehydrogenase, which converts acetoin to 2,3-butanediol, was deleted. Furthermore, by NAD+ regeneration through overexpression of water-forming NADH oxidase (NoxE) from Lactococcus lactis, the cofactor imbalance generated during the acetoin production from glucose was successfully relieved. As a result, in fed-batch fermentation, the engineered strain JHY617-SDN produced 100.1 g/L acetoin with a yield of 0.44 g/g glucose.

Metabolic engineering of enhanced glycerol-3-phosphate synthesis to increase lipid production in Synechocystis sp. PCC 6803.

With the growing attention to global warming and energy sustainability, biosynthesis of lipids by photosynthetic microorganisms has attracted more interest for the production of renewable transportation fuels. Recently, the cyanobacterium Synechocystis sp. PCC 6803 has been widely used for biofuel production through metabolic engineering because of its efficient photosynthesis and well-developed genetic tools. In lipid biosynthesis, glycerol-3-phosphate (G3P) is a key node for both CO2 fixation and lipid metabolism in cyanobacteria. However, few studies have explored the use of G3P synthesis to improve photosynthetic lipid production. In this study, metabolic engineering combined with flux balance analysis (FBA) was conducted to reveal the effect of G3P synthesis on lipid production. Heterologous genes that encoded glycerol-3-phosphate dehydrogenase (GPD) and diacylglycerol acyltransferase (DGAT) were engineered into Synechocystis sp. PCC 6803 to enhance G3P supply and lipid production. The resultant recombinant Synechocystis produced higher levels of lipids without a significant reduction in cell growth. Compared with the wild-type strain, lipid content and productivity of the engineered cyanobacteria increased by up to 36 and 31%, respectively, under autotrophic conditions. Lipid production under mixotrophic conditions of the engineered cyanobacteria was also investigated. This work demonstrated that enhanced G3P synthesis was an important factor in photosynthetic lipid production and that introducing heterologous GPD and DGAT genes was an effective strategy to increase lipid production in Synechocystis sp. PCC 6803.

Glycerol outflow in Symbiodinium under osmotic and nitrogen stress

In marine nutrient-poor environments, the success of Symbiodinium–cnidarian symbioses depends on the translocation of photosynthetically fixed carbon to the host. However, the mechanisms involved in this translocation are not well understood. Glycerol has been identified in the literature as one of the main photosynthates translocated from symbiont to host. Considering that glycerol is a preferred regulatory osmolyte in many unicellular eukaryotes and exposure of isolated symbionts to host homogenate results in the liberation of glycerol, we investigated the potential role of glycerol and its release in Symbiodinium. We studied the response to high osmolarity in two cultured species of Symbiodinium, examining glycerol production, specific activity of the enzyme glycerol 3-phosphate dehydrogenase and expression of the coding gene. We also assessed the production of glycerol and glucose under osmotic stress and nitrogen depletion. Results showed that cells exposed to high osmolarity conditions induced the synthesis of glycerol, although it was not retained effectively inside cells. In addition, nitrogen depletion also induced the synthesis of glycerol, although values were an order of magnitude lower. Interestingly, a significant decrease in glucose levels was detected in osmotically stressed cultures and under low nitrogen, possibly associated with the storage of carbon. While glycerol is a metabolite induced by osmotic stress, our results do not support its role as osmolyte. We conclude that glycerol may have a role as a sink for reducing power to prevent feedback inhibition of photosynthesis, particularly under low nitrogen, conditions that may be prevalent in the symbiosis.

The Complex Role of Asn270 in Dianion Activation of Glycerol 3‐Phosphate Dehydrogenase‐Catalyzed Hydride Transfer

We are working to understand the mechanism for the strong phosphite dianion (HPi) activation of proton transfer reactions catalyzed by triosephosphate isomerase (TIM), decarboxylation reactions catalyzed by orotidine monophosphate decarboxylase (OMPDC), and hydride transfer reactions catalyzed by glycerol 3‐phosphate dehydrogenase (GPDH). The phosphodianion of substrate dihydroxyacetone phosphate (DHAP) for GPDH provides 11 kcal/mol stabilization of the transition state for hydride transfer. Eight kcal/mol of this stabilization is observed as HPi activation of GPDH for catalysis of the reduction of glycolaldehyde (GA). The X‐ray crystal structure of the nonproductive complex between GPDH, DHAP and NAD+ shows interactions of Arg269 and Asn270 with the substrate phosphodianion. The R269A mutation of GPDH results in a large 9.1 kcal/mol destabilization of the transition state for enzyme‐catalyzed reduction of DHAP from elimination of the enzyme‐substrate ion‐pair; and 6.7 kcal/mol of this stabilization is recovered when the reaction is carried out in the presence of 1.0 M guanidinium cation.1 Similar effects were reported for K12G and R235A mutations of TIM2 and OMPDC,3 respectively, consistent with a similar architecture of the dianion activation sites for these enzymes. However, the following effects of N270A and R269A/N270A mutations require an unexpected greater mechanistic complexity for dianion activation of GPDH, compared with OMPDC and TIM. 1.) The 5.6 kcal/mol effect from N270A mutation on transition state stability is more than twice as large as that predicted for the deleted phosphodianion‐amide interaction. 2.) The effects of consecutive R269A and N270A mutations depend strongly on the order of these mutations. 3.) The N270A mutation results in a surprising 40‐fold increase in the second‐order rate constant for GPDH‐catalyzed reduction of GA, and causes HPi to change from a strong activator to a weak inhibitor of this GPDH‐catalyzed reaction. These large and complex changes in the activation barriers for hydride transfer show that the amide side chain of Asn270 functions as a “linchpin” in maintaining the structural integrity of the dianion activation site for GPDH.

The Roles of GPD1 in Maintaining Blood Glucose Levels under Fasting Conditions

Glucose is used as one of the energy sources in many organs. Under feeding conditions, glucose can be obtained from dietary carbohydrate. On the other hand, when external supply of energy is interrupted, such as fasting, carbohydrate preserved in liver and glycogenic precursor derived from other organs are used to maintain blood glucose levels. Glycerol and glycogenic amino acid derived from adipocyte and skeletal muscle, respectively, are utilized as glycogenic precursor. But, it hasn't become clear which glycogenic precursor is more important in gluconeogenesis. The recent study revealed that Metformin, therapeutics for treating type 2 diabetes, depressed gluconeogenesis from glycerol by inhibition of glycerol‐3‐phosphate dehydrogenase (GPD) 2. This result suggests that glycerol have a potent impact on gluconeogenesis. In this study, we focused attention on GPD1, which is the enzyme that related gluconeogenesis using glycerol, and examined the roles of GPD1 in gluconeogenesis and importance of glycerol as glycogenic precursor. Using GPD1 null mutant model BALB/cHeA mice (HeA mice), we measured the change of blood glucose levels under fasting conditions and gene expressions related to gluconeogenesis in liver. BALB/cBy mice (By mice) were used as a control. In HeA mice, blood glucose levels at 1 to 4 hour after fasting were significantly higher than the By mice. The hepatic gene expressions involved in gluconeogenesis and aminotransferase such as glucose‐6‐phosphatase and alanine amino transferase were also higher than the By mice. Moreover, we examined availability of the glycogenic amino acid by alanine tolerance test. At 30 min after alanine administration, blood glucose level was significantly higher in the HeA mice. In short, these data suggests that lacking of GPD1 maintained blood glucose levels under fasting state by enhancing gluconeogenesis from glycogenic amino acid and release of glucose into the blood stream.Support or Funding InformationThis study was supported by the Council for Science, Technology and Innovation (CSTI), Cross‐ministerial Strategic Innovation Promotion Program (SIP, No.14533567), and “Technologies for creating next‐generation agriculture, forestry and fisheries” (funding agency: Bio‐oriented Technology Research Advancement Institution, NARO), Grants‐in‐Aid for Scientific Research (KAKENHI, No. 15J10165, 15K00827) from the Japanese Ministry of Education, Culture, Sports, Science and Technology (MEXT, Tokyo), and University of Shizuoka Grant for Scientific and Educational Research.

Determinants of Cofactor Specificity for the Glucose-6-Phosphate Dehydrogenase from Escherichia coli: Simulation, Kinetics and Evolutionary Studies.

Glucose 6-Phosphate Dehydrogenases (G6PDHs) from different sources show varying specificities towards NAD+ and NADP+ as cofactors. However, it is not known to what extent structural determinants of cofactor preference are conserved in the G6PDH family. In this work, molecular simulations, kinetic characterization of site-directed mutants and phylogenetic analyses were used to study the structural basis for the strong preference towards NADP+ shown by the G6PDH from Escherichia coli. Molecular Dynamics trajectories of homology models showed a highly favorable binding energy for residues K18 and R50 when interacting with the 2'-phosphate of NADP+, but the same residues formed no observable interactions in the case of NAD+. Alanine mutants of both residues were kinetically characterized and analyzed with respect to the binding energy of the transition state, according to the kcat/KM value determined for each cofactor. Whereas both residues contribute to the binding energy of NADP+, only R50 makes a contribution (about -1 kcal/mol) to NAD+ binding. In the absence of both positive charges the enzyme was unable to discriminate NADP+ from NAD+. Although kinetic data is sparse, the observed distribution of cofactor preferences within the phylogenetic tree is sufficient to rule out the possibility that the known NADP+-specific G6PDHs form a monophyletic group. While the β1-α1 loop shows no strict conservation of K18, (rather, S and T seem to be more frequent), in the case of the β2-α2 loop, different degrees of conservation are observed for R50. Noteworthy is the fact that a K18T mutant is indistinguishable from K18A in terms of cofactor preference. We conclude that the structural determinants for the strict discrimination against NAD+ in the case of the NADP+-specific enzymes have evolved independently through different means during the evolution of the G6PDH family. We further suggest that other regions in the cofactor binding pocket, besides the β1-α1 and β2-α2 loops, play a role in determining cofactor preference.

Enzyme Architecture: A Startling Role for Asn270 in Glycerol 3-Phosphate Dehydrogenase-Catalyzed Hydride Transfer.

The side chains of R269 and N270 interact with the phosphodianion of dihydroxyacetone phosphate (DHAP) bound to glycerol 3-phosphate dehydrogenase (GPDH). The R269A, N270A, and R269A/N270A mutations of GPDH result in 9.1, 5.6, and 11.5 kcal/mol destabilization, respectively, of the transition state for GPDH-catalyzed reduction of DHAP by the reduced form of nicotinamide adenine dinucleotide. The N270A mutation results in a 7.7 kcal/mol decrease in the intrinsic phosphodianion binding energy, which is larger than the 5.6 kcal/mol effect of the mutation on the stability of the transition state for reduction of DHAP; a 2.2 kcal/mol stabilization of the transition state for unactivated hydride transfer to the truncated substrate glycolaldehyde (GA); and a change in the effect of phosphite dianion on GPDH-catalyzed reduction of GA, from strongly activating to inhibiting. The N270A mutation breaks the network of hydrogen bonding side chains, Asn270, Thr264, Asn205, Lys204, Asp260, and Lys120, which connect the dianion activation and catalytic sites of GPDH. We propose that this disruption dramatically alters the performance of GPDH at these sites.

Two glycerol 3-phosphate dehydrogenase isogenes from Candida versatilis SN-18 play an important role in glycerol biosynthesis under osmotic stress

Two isogenes of glycerol 3-phosphate dehydrogenase (GPD) from Candida versatilis SN-18 were cloned and sequenced. These intronless genes (Cagpd1 and Cagpd2) were both predicted to encode a 378 amino acid polypeptide, and the deduced amino acid sequences mutually showed 76% identity. Interestingly, Cagpd1 and Cagpd2 were located tandemly in a locus of genomic DNA within a 262 bp interval. To our knowledge, this represents a novel instance of isogenic genes relating to glucose metabolism. The stress response element (STRE) was found respectively at -93 to -89 bp upstream of the 5'end of Cagpd1 and -707 to -703 bp upstream of Cagpd2, indicating that these genes are involved in osmotic stress response. In heterologous expression using a gpd1Δgpd2Δ double deletion mutant of Saccharomyces cerevisiae, Cagpd1 and Cagpd2 transformants complemented the function of GPD, with Cagpd2 being much more effective than Cagpd1 in promoting growth and glycerol synthesis. Phylogenetic analysis of the amino acid sequences suggested that Cagpd1p and Cagpd2p are NADP(+)-dependent GPDs (EC 1.1.1.94). However, crude enzyme extract from Cagpd1 and Cagpd2 transformants showed GPD activity with only NAD(+) as cofactor. Hence, both Cagpd1p and Cagpd2p are likely NAD(+)-dependent GPDs (EC 1.1.1.8), similar to GPDs from S. cerevisiae and Candida magnoliae.

Cloning, expression, and characterization of a thermostable glucose-6-phosphate dehydrogenase from Thermoanaerobacter tengcongensis.

Glucose-6-phosphate dehydrogenases (G6PDs) are important enzymes widely used in bioassay and biocatalysis. In this study, we reported the cloning, expression, and enzymatic characterization of G6PDs from the thermophilic bacterium Thermoanaerobacter tengcongensis MB4 (TtG6PD). SDS-PAGE showed that purified recombinant enzyme had an apparent subunit molecular weight of 60kDa. Kinetics assay indicated that TtG6PD preferred NADP(+) (k cat/K m=2618mM(-1)s(-1), k cat=249s(-1), K m=0.10±0.01mM) as cofactor, although NAD(+) (k cat/K m=138mM(-1)s(-1), k cat=604s(-1), K m=4.37±0.56mM) could also be accepted. The K m values of glucose-6-phosphate were 0.27±0.07mM and 5.08±0.68mM with NADP(+) and NAD(+) as cofactors, respectively. The enzyme displayed its optimum activity at pH 6.8-9.0 for NADP(+) and at pH 7.0-8.6 for NAD(+) while the optimal temperature was 80°C for NADP(+) and 70°C for NAD(+). This was the first observation that the NADP(+)-linked optimal temperature of a dual coenzyme-specific G6PD was higher than the NAD(+)-linked and growth (75°C) optimal temperature, which suggested G6PD might contribute to the thermal resistance of a bacterium. The potential of TtG6PD to measure the activity of another thermophilic enzyme was demonstrated by the coupled assays for a thermophilic glucokinase.

Debaryomyces Hansenii: Adaptation Mechanisms to Different Carbon Sources and Oxygen Concentrations

Debaryomyces hansenii is an oleaginous, mostly aerobic yeast. Its mitochondrial respiratory chain contains complejxes I, II, III and IV, an alternative oxidase, aln alternative external NADH dehydrogenase an a glycerol-phosphate dehydrogenase (Cabrera-Orefice et al, 2014, Biochim Biophys Acta 1837(1): 73-84; 2014, Biochimie 102: 124-136). Alternative dehydrogenases, seem to physiologically uncouple mitochondria in order to prevent ROS production (Guerrero-Castillo et al, 2011, Biochim Biophys Acta 1817(2): 353-62). In addition, the opening of a unspecific channel releases NAD+, thus inactivating complex I and decreasing the proton-pumping stoichiometry in the system. In regard to the carbon source, lactate or succinate led to the expression of AOX at all growth phases. In contrast, in the presence of glucose or galactose, AOX was only expressed at the late exponential phase. In contrast, in glycerol AOX was never expressed and biomass growth was larger. In addition, large lipidic production was observed. It is suggested that D. hansenii is useful to study cell adaptations to different growth conditions. Also its lipid production mechanisms might be interesting from a biotechnological point of view. The regulation of redox enzyme expression maintains a high respiratory activity even when ATP consumption is low, probably preventing overproduction of ROS.

Over-expression of Arabidopsis thaliana SFD1/GLY1, the gene encoding plastid localized glycerol-3-phosphate dehydrogenase, increases plastidic lipid content in transgenic rice plants

Lipids are the major constituents of all membranous structures in plants. Plants possess two pathways for lipid biosynthesis: the prokaryotic pathway (i.e., plastidic pathway) and the eukaryotic pathway (i.e., endoplasmic-reticulum (ER) pathway). Whereas some plants synthesize galactolipids from diacylglycerol assembled in the plastid, others, including rice, derive their galactolipids from diacylglycerols assembled by the eukaryotic pathway. Arabidopsis thaliana glycerol-3-phosphate dehydrogenase (G3pDH), coded by SUPPRESSOR OF FATTY ACID DESATURASE 1 (SFD1; alias GLY1) gene, catalyzes the formation of glycerol 3-phosphate (G3p), the backbone of many membrane lipids. Here SFD1 was introduced to rice as a transgene. Arabidopsis SFD1 localizes in rice plastids and its over-expression increases plastidic membrane lipid content in transgenic rice plants without any major impact on ER lipids. The results suggest that over-expression of plastidic G3pDH enhances biosynthesis of plastid-localized lipids in rice. Lipid composition in the transgenic plants is consistent with increased phosphatidylglycerol synthesis in the plastid and increased galactolipid synthesis from diacylglycerol produced via the ER pathway. The transgenic plants show a higher photosynthetic assimilation rate, suggesting a possible application of this finding in crop improvement.

Birth of Archaeal Cells: Molecular Phylogenetic Analyses of G1P Dehydrogenase, G3P Dehydrogenases, and Glycerol Kinase Suggest Derived Features of Archaeal Membranes Having G1P Polar Lipids.

Bacteria and Eukarya have cell membranes with sn-glycerol-3-phosphate (G3P), whereas archaeal membranes contain sn-glycerol-1-phosphate (G1P). Determining the time at which cells with either G3P-lipid membranes or G1P-lipid membranes appeared is important for understanding the early evolution of terrestrial life. To clarify this issue, we reconstructed molecular phylogenetic trees of G1PDH (G1P dehydrogenase; EgsA/AraM) which is responsible for G1P synthesis and G3PDHs (G3P dehydrogenase; GpsA and GlpA/GlpD) and glycerol kinase (GlpK) which is responsible for G3P synthesis. Together with the distribution of these protein-encoding genes among archaeal and bacterial groups, our phylogenetic analyses suggested that GlpA/GlpD in the Commonote (the last universal common ancestor of all extant life with a cellular form, Commonote commonote) acquired EgsA (G1PDH) from the archaeal common ancestor (Commonote archaea) and acquired GpsA and GlpK from a bacterial common ancestor (Commonote bacteria). In our scenario based on this study, the Commonote probably possessed a G3P-lipid membrane synthesized enzymatically, after which the archaeal lineage acquired G1PDH followed by the replacement of a G3P-lipid membrane with a G1P-lipid membrane.

Specific features of the lymphocyte enzyme status in newborn infants with the hypotrophic type of intrauterine growth retardation

Objective: to reveal the specific features of the lymphocyte enzyme status in newborn infants with hypotrophic type of intrauterine growth retardation under the conditions of Yakutia. Subjects and methods. A total of 137 full-term newborns (including 28 babies with intrauterine growth retardation) whose parents were Yakutsk newcomers were examined on 1, 3 and 5 days of life. The cytochemical activity of lymphocyte enzymes, by determining the level of and time course of changes in succinate dehydrogenase (SDH), glycerol-3-phosphate dehydrogenase (GPDH), lactate dehydrogenase (LDH) and its aerobic isoenzyme (LDH-H), NADFH2-diaphorase, monoamine oxidase (MAO), and acid phosphatase (AP), were analyzed. Results. It was ascertained that the activity of SDH, GPDH, and LDH was higher on 5 day of life and that of NADFH2-diaphorase was lower in infants with intrauterine growth retardation than in apparently healthy neonates. Desynchronosis of the metabolic pathways was established in the dynamics of early ontogenesis. Conclusion. It is necessary to rationally correct standard treatment regimens, by taking into account the found metabolic features in infants with intrauterine growth retardation under the conditions of Yakutia.

MAPK in Dunaliella tertiolecta regulates glycerol production in response to osmotic shock

Glycerol production is modulated in some halo-tolerant organisms in response to extracellular osmotic stress. Glycerol-3-phosphate dehydrogenase (GPDH) is a rate-limiting enzyme of glycerol synthesis in both yeast and the green microalgae Dunaliella. In yeast, a High-Osmolarity-Glycerol (HOG) pathway, which is a mitogen-activated protein kinase (MAPK) cascade, has been suggested to activate the expression of GPDH and thus the accumulation of glycerol under osmotic stress. In Dunaliella tertiolecta, however, the osmo-regulatory mechanisms for glycerol synthesis are not well understood. In this study, the homologues of MAPK and GPDH in D. tertiolecta were cloned using Rapid Amplification of cDNA Ends (RACE) to investigate the molecular basis of the osmo-regulatory mechanisms. The isolated cDNA sequences were named DtMAPK (GenBank: KJ930518) and DtGPDH (GenBank: KJ930370), respectively. It was found that after osmotic shock DtMAPK and DtGPDH expression increased within 0.5 h and 1 h, respectively. In addition, glycerol production and DtGPDH expression level paralleled the expression of DtMAPK under various osmotic stress conditions. This suggests a close correlation between the expression of these two genes and glycerol production in D. tertiolecta. Moreover, suppressed transcription of DtGPDH and delayed accumulation of intracellular glycerol were observed in DtMAPK knock-down cells upon hyper-osmotic shock, providing further evidence that DtMAPK is involved in the regulation of DtGPDH expression and thus glycerol synthesis. Our findings demonstrate that a MAPK-mediated signalling pathway similar to the yeast HOG pathway may exist in D. tertiolecta in response to osmotic stress.

Vitamin A deficiency suppresses high fructose-induced triglyceride synthesis and elevates resolvin D1 levels

Background/AimsVitamin A and its metabolites are known to regulate lipid metabolism. However so far, no study has assessed, whether vitamin A deficiency per se aggravates or attenuates the development of non-alcoholic fatty liver disease (NAFLD). Therefore, here, we tested the impact of vitamin A deficiency on the development of NAFLD. MethodsMale weanling Wistar rats were fed one of the following diets; control, vitamin A-deficient (VAD), high fructose (HFr) and VAD with HFr (VADHFr) of AIN93G composition, for 16weeks, except half of the VAD diet-fed rats were shifted to HFr diet (VAD(s)HFr), at the end of 8th week. ResultsAnimals fed on VAD diet with HFr displayed hypotriglyceridemia (33.5mg/dL) with attenuated hepatic triglyceride accumulation (8.2mg/g), compared with HFr diet (89.5mg/dL and 20.6mg/g respectively). These changes could be partly explained by the decreased activity of glycerol 3-phosphate dehydrogenase (GPDH) and the down-regulation of stearoyl CoA desaturase 1 (SCD1), both at gene and protein levels, the key determinants of triglyceride biosynthesis. On the other hand, n-3 long chain polyunsaturated fatty acid, docosahexaenoic acid and its active metabolite; resolvin D1 (RvD1) levels were elevated in the liver and plasma of VAD diet-fed groups, which was negatively associated with triglyceride levels. All these factors confer vitamin A deficiency-mediated protection against the development of hepatic steatosis, which was also evident from the group shifted from VAD to HFr diet. ConclusionsVitamin A deficiency attenuates high fructose-induced hepatic steatosis, by regulating triglyceride synthesis, possibly through GPDH, SCD1 and RvD1.