Glycerol-3-phosphate Dehydrogenase Research Articles

Mitochondrial glycerol 3-phosphate facilitates bumblebee pre-flight thermogenesis

Bumblebees (Bombus terrestris) fly at low ambient temperatures where other insects cannot, and to do so they must pre-warm their flight muscles. While some have proposed mechanisms, none fully explain how pre-flight thermogenesis occurs. Here, we present a novel hypothesis based on the less studied mitochondrial glycerol 3-phosphate dehydrogenase pathway (mGPDH). Using calorimetry, and high resolution respirometry coupled with fluorimetry, we report substrate oxidation by mGPDH in permeabilised flight muscles operates, in vitro, at a high flux, even in the absence of ADP. This may be facilitated by an endogenous, mGPDH-mediated uncoupling of mitochondria. This uncoupling increases ETS activity, which results in increased heat release. Furthermore, passive regulation of this mechanism is achieved via dampened temperature sensitivity of mGPDH relative to other respiratory pathways, and subsequent consumption of its substrate, glycerol 3-phosphate (G3P), at low temperatures. Mitochondrial GPDH may therefore facilitate pre-flight thermogenesis through poor mitochondrial coupling. We calculate this can occur at a sufficient rate to warm flight muscles until shivering commences, and until flight muscle function is adequate for bumblebees to fly in the cold.

Acute renal metabolic effect of metformin assessed with hyperpolarised MRI in rats.

Metformin inhibits hepatic mitochondrial glycerol phosphate dehydrogenase, thereby increasing cytosolic lactate and suppressing gluconeogenesis flux in the liver. This inhibition alters cytosolic and mitochondrial reduction-oxidation (redox) potential, which has been reported to protect organ function in several disease states including diabetes. In this study, we investigated the acute metabolic and functional changes induced by metformin in the kidneys of both healthy and insulinopenic Wistar rats used as a model of diabetes. Diabetes was induced by intravenous injection of streptozotocin, and kidney metabolism in healthy and diabetic animals was investigated 4weeks thereafter using hyperpolarised 13C-MRI, Clark-type electrodes and biochemical analysis. Metformin increased renal blood flow, but did not change total kidney oxygen consumption. In healthy rat kidneys, metformin increased [1-13C]lactate production and reduced mitochondrial [1-13C]pyruvate oxidation (decreased the 13C-bicarbonate/[1-13C]pyruvate ratio) within 30min of administration. Corresponding alterations to indices of mitochondrial, cytosolic and whole-cell redox potential were observed. Pyruvate oxidation was maintained in the diabetic rats, suggesting that the diabetic state abrogates metabolic reprogramming caused by metformin. This study demonstrates that metformin-induced acute metabolic alterations in healthy kidneys favoured anaerobic metabolism at the expense of aerobic metabolism. The results suggest that metformin directly alters the renal redox state, with elevated renal cytosolic redox states as well as decreased mitochondrial redox state. These findings suggest redox biology as a novel target to eliminate the renal complications associated with metformin treatment in individuals with impaired renal function.

Peripheral neuropeptide Y differentially influences adipogenesis and lipolysis in chicks from lines selected for low or high body weight

Neuropeptide Y (NPY) stimulates appetite and promotes lipid deposition. We demonstrated a differential sensitivity in the food intake response to central NPY in chicks from lines selected for low (LWS) or high (HWS) body weight, but have not reported whether such differences exist in the periphery. At 5days, LWS and HWS chicks were intraperitoneally injected with 0 (vehicle), 60, or 120μg/kg BW NPY and subcutaneous adipose tissue and plasma were collected at 1, 3, 6, 12, and 24h (n=12). NPY injection increased glycerol-3-phosphate dehydrogenase (G3PDH) activity at 1 and 3h and reduced plasma non-esterified fatty acids (NEFAs) at 1 and 12h. G3PDH activity was greater in HWS than LWS while NEFAs were greater in LWS. At 1h, peroxisome proliferator-activated receptor gamma (PPARγ), CCAAT/enhancer binding protein (C/EBP)α, and microsomal triglyceride transfer protein (MTTP) mRNAs were reduced in NPY-injected chicks whereas NPY receptor 1 (NPYR1) was increased. Expression of stearoyl-CoA desaturase (SCD1) was increased by NPY at 1h in HWS but not LWS. PPARγ (3 and 6h), C/EBPβ (3h), C/EBPα (6h) and NPYR1 and 2 (24h) mRNAs were greater in NPY- than vehicle-injected chicks. At several times, adipose triglyceride lipase, MTTP, perilipin 1, NPYR1, and NPYR2 mRNAs were greater in LWS than HWS, while expression of SCD1, glycerol-3-phosphate acyltransferase 3 and lipoprotein lipase was greater in HWS than LWS. Thus, NPY promotes fat deposition and inhibits lipolysis in chicks, with line differences indicative of greater rates of lipolysis in LWS and adipogenesis in HWS.

The Inhibitory Effect of Tartary Buckwheat Extracts on Adipogenesis and Inflammatory Response.

Tartary buckwheat (Fagopyrum tataricum) has been established globally as a nutritionally important food item, particularly owing to high levels of bioactive compounds such as rutin. This study investigated the effect of tartary buckwheat extracts (TBEs) on adipogenesis and inflammatory response in 3T3-L1 cells. TBEs inhibited lipid accumulation, triglyceride content, and glycerol-3-phosphate dehydrogenase (GPDH) activity during adipocyte differentiation of 3T3 L1 cells. The mRNA levels of genes involved in fatty acid synthesis, such as peroxisome proliferator-activated receptor-γ (PPAR-γ), CCAAT/enhancer binding protein-α (CEBP-α), adipocyte protein 2 (aP2), acetyl-CoA carboxylase (ACC), fatty acid synthase (FAS), and stearoylcoenzyme A desaturase-1 (SCD-1), were suppressed by TBEs. They also reduced the mRNA levels of inflammatory mediators such as tumor necrosis factor-α (TNF-α), interleukin-6 (IL-6), monocyte chemoattractant protein 1 (MCP-1), and inducible nitric oxide synthase (iNOS). In addition, TBEs were decreased nitric oxide (NO) production. These results suggest that TBEs may inhibit adipogenesis and inflammatory response; therefore, they seem to be beneficial as a food ingredient to prevent obesity-associated inflammation.

Two Glycerol-3-Phosphate Dehydrogenases from Chlamydomonas Have Distinct Roles in Lipid Metabolism.

The metabolism of glycerol-3-phosphate (G3P) is important for environmental stress responses by eukaryotic microalgae. G3P is an essential precursor for glycerolipid synthesis and the accumulation of triacylglycerol (TAG) in response to nutrient starvation. G3P dehydrogenase (GPDH) mediates G3P synthesis, but the roles of specific GPDH isoforms are currently poorly understood. Of the five GPDH enzymes in the model alga Chlamydomonas reinhardtii, GPD2 and GPD3 were shown to be induced by nutrient starvation and/or salt stress. Heterologous expression of GPD2, a putative chloroplastic GPDH, and GPD3, a putative cytosolic GPDH, in a yeast gpd1Δ mutant demonstrated the functionality of both enzymes. C. reinhardtii knockdown mutants for GPD2 and GPD3 showed no difference in growth but displayed significant reduction in TAG concentration compared with the wild type in response to phosphorus or nitrogen starvation. Overexpression of GPD2 and GPD3 in C. reinhardtii gave distinct phenotypes. GPD2 overexpression lines showed only subtle metabolic phenotypes and no significant alteration in growth. In contrast, GPD3 overexpression lines displayed significantly inhibited growth and chlorophyll concentration, reduced glycerol concentration, and changes to lipid composition compared with the wild type, including increased abundance of phosphatidic acids but reduced abundance of diglycerides, triglycerides, and phosphatidylglycerol lipids. This may indicate a block in the downstream glycerolipid metabolism pathway in GPD3 overexpression lines. Thus, lipid engineering by GPDH modification may depend on the activities of other downstream enzyme steps. These results also suggest that GPD2 and GPD3 GPDH isoforms are important for nutrient starvation-induced TAG accumulation but have distinct metabolic functions.

Fatty Acids Have Different Adipogenic Differentiation Potentials in Stromal Vascular Cells Isolated from Abdominal Fat in Laying Hens.

This study was conducted to examine the effects of fatty acids (FA) with/without chicken serum (CS) on the expression of adipogenic transcripts and adipogenesis in chicken stromal vascular cells (SVC). In experiment 1, SVC were grown in DMEM containing 10% FBS (Control) and treated with 300µM oleic acid (OLA)+FBS, linoleic acid (LNA)+FBS, palmitic acid (PAM)+FBS, or stearic acid (STA)+FBS for 48h. In experiment 2, cells were grown in DMEM containing 5% CS and treated with 300µM OLA (CS+OLA), PAM (CS+PAM), STA (CS+STA) or 200µM LNA (CS+LNA) for 48h. Adipogenesis was determined using Oil Red O staining and glycerol-3-phosphate dehydrogenase (GPDH) activity. The proportion of OLA, PAM, or STA was increased (P<0.05) in SVC grown in either FBS or CS with OLA, PAM or STA. Adipogenesis was induced in FBS+OLA, FBS+LNA, FBS+PAM, FBS+STA, CS+OLA, CS+LNA, CS+PAM, or CS+SAT compared to FBS. GPDH activity was significantly higher in FBS+OLA and FBS+LNA than one in FBS. Compared to FBS, the expression of FABP4 mRNA increased (P<0.05) in FBS+OLA, FBS+LNA, or FBS+PAM, whereas that of C/EBPα, C/EBPβ, and ATGL increased (P<0.05) in FBS+OLA or FBS+LNA cells. Expression of FABP4 and C/EBPβ mRNA was higher in CS, CS+OLA, CS+LNA, CS+PAM, or CS+SAT compared with (FBS, whereas the expression of ATGL and C/EBPα was higher in CS, CS+OLA, or CS+LNA than FBS cells. In conclusion, these results showed that FA have different potentials to induce adipogenesis, LNA is the most potent among the tested FA, and these potentials can be improved in the presence of CS.

Preventive Effect of blueberries on Diethylnitrosamine-Induced Hepatocarcinogenesis in Rats

Nitrosamine compounds are known hepatic carcinogens. This study was designed to study the efficacy of dietary supplementation with blueberries (BB) on diethylnitrosamine (DEN)-initiated hepatocarcinogenesis in male wistar rats. Rats were divided into three groups. The first group served as normal control group, the second group received DEN at a dose of 10 mg/kg body weight five times a week for 15 weeks. The third one received DEN as in DEN-treated group simultaneously with 4% BB-supplemented diet. The results showed that BB caused significant decrease in the elevated serum levels of α-fetoprotein (AFP), homocysteine (Hcy) along with levels of glutathione(GSH), deoxyribonucleic acid (DNA), ribonucleic acid (RNA)and activity of glutathione reductase (GR) in liver. Normalization of elevated 2-macroglobulin (2M) and total antioxidant capacity (TAC) levels in serum, hepatic Glutathione-S-transferase (GST), glutathione peroxidase (GPx) activities and liver weight was achieved whereas body weight was significantly decreased. Moreover, no significant change was observed in elevated relative liver weight, hepatic glucose-6-P-dehydrogenase (G6PD), lactate dehydrogenase (LDH) along with serum aminotransferases, alkaline phosphatase (ALP) and ɤ -glutamyltransferase (ɤ-GT) activities. Significant increase in reduced hepatic activity of xanthine oxidase (XO) was achieved and histopathological damage was minimized in BB-treated group. It is suggested that BB suppress DEN-induced hepatocarcinogenesis and could be developed as a promising chemoprotective natural supplement for liver cancer.

Metabolic engineering strategies for optimizing acetate reduction, ethanol yield and osmotolerance in Saccharomyces cerevisiae

BackgroundGlycerol, whose formation contributes to cellular redox balancing and osmoregulation in Saccharomyces cerevisiae, is an important by-product of yeast-based bioethanol production. Replacing the glycerol pathway by an engineered pathway for NAD+-dependent acetate reduction has been shown to improve ethanol yields and contribute to detoxification of acetate-containing media. However, the osmosensitivity of glycerol non-producing strains limits their applicability in high-osmolarity industrial processes. This study explores engineering strategies for minimizing glycerol production by acetate-reducing strains, while retaining osmotolerance.ResultsGPD2 encodes one of two S. cerevisiae isoenzymes of NAD+-dependent glycerol-3-phosphate dehydrogenase (G3PDH). Its deletion in an acetate-reducing strain yielded a fourfold lower glycerol production in anaerobic, low-osmolarity cultures but hardly affected glycerol production at high osmolarity. Replacement of both native G3PDHs by an archaeal NADP+-preferring enzyme, combined with deletion of ALD6, yielded an acetate-reducing strain the phenotype of which resembled that of a glycerol-negative gpd1Δ gpd2Δ strain in low-osmolarity cultures. This strain grew anaerobically at high osmolarity (1 mol L−1 glucose), while consuming acetate and producing virtually no extracellular glycerol. Its ethanol yield in high-osmolarity cultures was 13% higher than that of an acetate-reducing strain expressing the native glycerol pathway.ConclusionsDeletion of GPD2 provides an attractive strategy for improving product yields of acetate-reducing S. cerevisiae strains in low, but not in high-osmolarity media. Replacement of the native yeast G3PDHs by a heterologous NADP+-preferring enzyme, combined with deletion of ALD6, virtually eliminated glycerol production in high-osmolarity cultures while enabling efficient reduction of acetate to ethanol. After further optimization of growth kinetics, this strategy for uncoupling the roles of glycerol formation in redox homeostasis and osmotolerance can be applicable for improving performance of industrial strains in high-gravity acetate-containing processes.

Chick subcutaneous and abdominal adipose tissue depots respond differently in lipolytic and adipogenic activity to α-melanocyte stimulating hormone (α-MSH)

In birds, α-MSH is anorexigenic, but effects on adipose tissue are unknown. Four day-old chicks were intraperitoneally injected with 0 (vehicle), 5, 10, or 50μg of α-MSH and subcutaneous and abdominal adipose tissue collected at 60min for RNA isolation (n=10). Plasma was collected post-euthanasia at 60 and 180min for measuring non-esterified fatty acids (NEFA) and α-MSH (n=10). Relative to the vehicle, food intake was reduced in the 50μg-treated group. Plasma NEFAs were greater in 10μg than vehicle-treated chicks at 3h. Plasma α-MSH was 3.06±0.57ng/ml. In subcutaneous tissue, melanocortin receptor 5 (MC5R) mRNA was increased in 10μg, MC2R and CCAAT-enhancer-binding protein β (C/EBPβ) mRNAs increased in 50μg, peroxisome proliferator-activated receptor γ and C/EBPα decreased in 5, 10 and 50μg, and Ki67 mRNA decreased in 50μg α-MSH-injected chicks, compared to vehicle-injected chicks. In abdominal tissue, adipose triglyceride lipase mRNA was greater in 10μg α-MSH- than vehicle-treated chicks. Cells isolated from abdominal fat that were treated with 10 and 100nM α-MSH for 4h expressed more MC5R and perilipin-1 than control cells (n=6). Cells that received 100nM α-MSH expressed more fatty acid binding protein 4 and comparative gene identification-58 mRNA than control cells. Glycerol-3-phosphate dehydrogenase (G3PDH) activity was greater in cells at 9days post-differentiation that were treated with 1 and 100nM α-MSH for 4h than in control cells (n=3). Results suggest that α-MSH increases lipolysis and reduces adipogenesis in adipose tissue.

A multidomain enzyme, with glycerol-3-phosphate dehydrogenase and phosphatase activities, is involved in a chloroplastic pathway for glycerol synthesis in Chlamydomonas reinhardtii.

Understanding the unique features of algal metabolism may be necessary to realize the full potential of algae as feedstock for the production of biofuels and biomaterials. Under nitrogen deprivation, the green alga C.reinhardtii showed substantial triacylglycerol (TAG) accumulation and up-regulation of a gene, GPD2, encoding a multidomain enzyme with a putative phosphoserine phosphatase (PSP) motif fused to glycerol-3-phosphate dehydrogenase (GPD) domains. Canonical GPD enzymes catalyze the synthesis of glycerol-3-phosphate (G3P) by reduction of dihydroxyacetone phosphate (DHAP). G3P forms the backbone of TAGs and membrane glycerolipids and it can be dephosphorylated to yield glycerol, an osmotic stabilizer and compatible solute under hypertonic stress. Recombinant Chlamydomonas GPD2 showed both reductase and phosphatase activities invitro and it can work as a bifunctional enzyme capable of synthesizing glycerol directly from DHAP. In addition, GPD2 and a gene encoding glycerol kinase were up-regulated in Chlamydomonas cells exposed to high salinity. RNA-mediated silencing of GPD2 revealed that the multidomain enzyme was required for TAG accumulation under nitrogen deprivation and for glycerol synthesis under high salinity. Moreover, a GPD2-mCherry fusion protein was found to localize to the chloroplast, supporting the existence of a GPD2-dependent plastid pathway for the rapid synthesis of glycerol in response to hyperosmotic stress. We hypothesize that the reductase and phosphatase activities of PSP-GPD multidomain enzymes may be modulated by post-translational modifications/mechanisms, allowing them to synthesize primarily G3P or glycerol depending on environmental conditions and/or metabolic demands in algal species of the core Chlorophytes.

Triosephosphate isomerase (TPI) facilitates the replication of WSSV in Exopalaemon carinicauda

Triosephosphate isomerase (TPI) is a vital enzyme in the glycolytic pathway, which can catalyze the interconversion of glyceraldehyde-3-phosphate (GAP) and dihydroxyacetone phosphate (DHAP). DHAP is involved in lipid metabolism and phospholipid synthesis. In order to know the role of TPI in WSSV infection to prawn, we cloned the full length cDNA of triosephosphate isomerase gene (EcTPI) from Exopalaemon carinicauda, and its function during WSSV infection was analyzed. EcTPI transcripts were widely distributed in all tissues, but showed relatively higher expression levels in the gill and epidermis. Its expression was apparently up-regulated after 24 h post WSSV injection (hpi), when the virus load began to rise. Furthermore, we detected the expressions of the key genes encoding the enzymes which catalyze the key steps in the glycolysis during WSSV infection. The data showed that genes encoding the enzymes which catalyzed upper steps of glycolysis to produce GAP, including hexokinase (HK), glucose-6-phosphate isomerase (GPI) and phosphofructokinase-1 (PFK-1), were significantly up-regulated at 24 and 27 hpi. Genes encoding the enzymes catalyzing down steps of glycolysis after GAP, including glyceraldehyde-3-phosphate dehydrogenase (GAPDH), enolase (ENO) and pyruvate kinase (PK), were apparent down-regulated at 24 and 27 hpi. Meanwhile, the gene encoding the enzyme glycerol-3-phosphate dehydrogenase (GPDH) catalyzing DHAP to glycerol-3-phosphate (G-3-P) showed down-regulation at 12–27 hpi, while the gene encoding dihydroxyacetone-phosphate acyltransferase (DHAPAT) catalyzing DHAP to further synthesis of phospholipids showed up-regulation at 12–24 hpi. These data suggested that WSSV infection could change the glycolysis pathway to make them produce more phospholipids which could be very helpful for virus replication. In order to further confirm the above speculation, dsRNA interference (RNAi) approach was used to knock down EcTPI gene and analyze its effect on WSSV load in prawn. The data showed that interference of EcTPI gene led to a significant decrease of WSSV loads in WSSV infected prawn. These data provided useful information to understand the infection mechanism of WSSV.

The Study of Physiological Reaction of GPDH in Maize Seedlings Under Abiotic Stress

The independent breeding maize variety He No. 344 seeds were used to study the physiological response of glycerol-3-phosphate dehydrogenase (GPDH) on maize under abiotic stresses, including salinity treatment (200 mM NaCl), alkali treatment (150 mM NaHCO3), dehydration treatment (20%-PEG) and low temperature (4°C). The results showed that the abiotic stress can significantly reduced the plant height, root length and dry weight of maize and the combined injury of saline alkali treatment was higher than that of drought and low temperature treatments; The abiotic stress also increased GPDH activity of maize leaf, and the activity of GPDH under saline alkali treatment was significantly higher than that of drought and low temperature treatments; The abiotic stress resulted in increased level of TBARS and H2O2 contents and increased NADH/ NAD+ of maize leaf; Moreover, the abiotic stress also increased SOD, APX, GR and GPX activities and ASA/DHA, GSH/GSSG; In addition, the GPDH activity induced by abiotic stress was significantly correlated with NADH/NAD+, APX, GPX, GR, ASA/DHA, GSH/GSSG, TBARS and H2O2 level, and these results indicate that the increased resistance of maize in the process of abiotic stresses was closely related with the metabolic level of GPDH.

Metabolic background for establishment of the subpopulation structure in early ontogenesis in the Atlantic salmon (the case of energy of carbohydrate metabolism)

The activity of some enzymes involved in energy and carbohydrate metabolism was studied in Atlantic salmon embryos at the eyed egg stage and in salmon fingerlings (0+) from two trophic–ecological groups: the Varzuga River bed and two tributaries, the Pyatka and Sobachii rivers (Kola Peninsula). It has been demonstrated that heterogeneity of embryos was most evident in the case of cytochrome c oxidase (CO), malate dehydrogenase (MDH), glycerol-1-phosphate dehydrogenase (G1PDH), and glucose-6-phosphate dehydrogenase (G6PDH), while the lowest level of heterogeneity was observed for lactate dehydrogenase (LDH) and aldolase. A positive correlation was revealed between the activities of CO, LDH, MDH, and G1PDH. It was noted that G6PDH showed a negative correlation with almost all enzymes under study. It was found that salmon juveniles inhabiting the tributaries were characterized by high LDH, aldolase, and G1PDH activity and lower activity of G6PDH compared to the juveniles inhabiting the main river bed. Notably, the differences in the activity of the enzymes involved in aerobic metabolism between the two groups of fingerlings under analysis were observed only in the autumn.

Highly active antiretroviral therapy dysregulates proliferation and differentiation of human pre-adipocytes.

AIMTo investigate the mechanism(s) by which potential effects of multi-drug highly-active antiretroviral therapy contributes to lipodystrophy syndrome.METHODSPreadipocytes from healthy donors were assessed for proliferation and differentiation in the presence of nucleoside reverse transcriptase inhibitors (NRTIs), nonnucleoside reverse transcriptase inhibitors (NNRTIs), and protease inhibitors (PIs) individually and in combination. Effects on proliferation were assessed with a 3-[4,5-dimethylthiazol-2-yl]-2,5-diphenyl tetrazolium bromide assay and effects on differentiation were assessed from glycerol-3-phosphate dehydrogenase (GP DH) activity and quantitation of Oil Red O staining for intracellular lipid. Data were analyzed with a randomized block ANOVA with post-hoc Fisher’s Least Significant Difference test.RESULTSPreadipocyte proliferation was inhibited by a combination of NNRTI + NRTI (14% at 48 h, P < 0.001) and PI + NRTI (19% at 48 h, P < 0.001) with additional suppression when ritonavir (RTV) was added (26% at 48 h). The drug combination of atazanavir (ATV) + RTV + emtricitabine (FTC) + tenofovir (TDF) had the greatest inhibitory effect on proliferation at 48 h. Preadipocyte differentiation was most significantly reduced by the efavirenz + FTC + TDF assessed either by GPDH activity (64%) or lipid accumulation (39%), P < 0.001. Combining NRTIs with a PI (ATV + FTC + TDF) significantly suppressed differentiation (GPDH activity reduced 29%, lipid accumulation reduced by 19%, P < 0.01). This effect was slightly greater when a boosting amount of RTV was added (ATV + FTC + TDF + RTV, P < 0.001).CONCLUSIONAlthough combination antiretroviral therapy is clinically more efficacious than single drug regimens, it also has a much greater inhibitory effect on preadipocyte proliferation and differentiation.

Experimentally Induced Bleaching in the Sea Anemone Exaiptasia Supports Glucose as a Main Metabolite Associated with Its Symbiosis

Our current understanding of carbon exchange between partners in the Symbiodinium-cnidarian symbioses is still limited, even though studies employing carbon isotopes have made us aware of the metabolic complexity of this exchange. We examined glycerol and glucose metabolism to better understand how photosynthates are exchanged between host and symbiont. The levels of these metabolites were compared between symbiotic and bleached Exaiptasia pallida anemones, assaying enzymes directly involved in their metabolism. We measured a significant decrease of glucose levels in bleached animals but a significant increase in glycerol and G3P pools, suggesting that bleached animals degrade lipids to compensate for the loss of symbionts and seem to rely on symbiotic glucose. The lower glycerol 3-phosphate dehydrogenase but higher glucose 6-phosphate dehydrogenase specific activities measured in bleached animals agree with a metabolic deficit mainly due to the loss of glucose from the ruptured symbiosis. These results corroborate previous observations on carbon translocation from symbiont to host in the sea anemone Exaiptasia, where glucose was proposed as a main translocated metabolite. To better understand photosynthate translocation and its regulation, additional research with other symbiotic cnidarians is needed, in particular, those with calcium carbonate skeletons.

Cocktail supplement with rosiglitazone: a novel inducer for chicken preadipocyte differentiation invitro.

The preadipocyte differentiation biological process involves a cascade of transcriptional events that culminates in the expression of peroxisome proliferator-activated receptor (PPAR) γ. The differentiation cocktail [insulin (INS), dexamethasone (DEX) and isobutylmethylxanthine (IBMX)] can induce preadipocyte differentiation in mammals, but it is insufficient for chicken (Gallus gallus) adipogenesis. Oleate can induce chicken preadipocyte differentiation, but these differentiated preadipocytes may not be fully functional. The objective of the current study was to evaluate whether chicken preadipocytes can be induced to mature adipocytes by a novel induction method using differentiation cocktail supplemented with PPARγ agonist(s). Chicken preadipocytes cultured in cocktail supplemented with rosiglitazone or troglitazone resulted in a marked increase in lipid droplet accumulation (P<0.05), glycerol-3-phosphate dehydrogenase (GPDH) activity (P<0.05), mRNA expression level of adipocyte fatty acid-binding protein (aP2; P<0.05), G0/G1 switch gene 2 (G0S2; P<0.05) and lipolysis (P<0.05). In addition, supplementation of the cocktail with rosiglitazone promoted PPARγ mRNA expression (P<0.05). In conclusion, our data indicated that chicken preadipocytes can be induced to mature adipocytes using differentiation cocktail supplemented with rosiglitazone. The results of the present study provide a novel induction method for invitro chicken preadipocyte differentiation.

Genomic, Transcriptomic, and Proteomic Analysis Provide Insights Into the Cold Adaptation Mechanism of the Obligate Psychrophilic Fungus Mrakia psychrophila.

Mrakia psychrophila is an obligate psychrophilic fungus. The cold adaptation mechanism of psychrophilic fungi remains unknown. Comparative genomics analysis indicated that M. psychrophila had a specific codon usage preference, especially for codons of Gly and Arg and its major facilitator superfamily (MFS) transporter gene family was expanded. Transcriptomic analysis revealed that genes involved in ribosome and energy metabolism were upregulated at 4°, while genes involved in unfolded protein binding, protein processing in the endoplasmic reticulum, proteasome, spliceosome, and mRNA surveillance were upregulated at 20°. In addition, genes related to unfolded protein binding were alternatively spliced. Consistent with other psychrophiles, desaturase and glycerol 3-phosphate dehydrogenase, which are involved in biosynthesis of unsaturated fatty acid and glycerol respectively, were upregulated at 4°. Cold adaptation of M. psychrophila is mediated by synthesizing unsaturated fatty acids to maintain membrane fluidity and accumulating glycerol as a cryoprotectant. The proteomic analysis indicated that the correlations between the dynamic patterns between transcript level changes and protein level changes for some pathways were positive at 4°, but negative at 20°. The death of M. psychrophila above 20° might be caused by an unfolded protein response.

Promotion of adipogenesis by neuropeptide Y during the later stages of chicken preadipocyte differentiation.

Neuropeptide Y (NPY) promotes adipogenesis in both birds and mammals, although mechanisms in avians remain unclear. The objective of this study was thus to evaluate effects of NPY on chick preadipocyte proliferation and differentiation. Preadipocytes were treated with 0, 1, 10, or 100 nmol/L NPY and gene expression and cellular proliferation were evaluated at 12, 24, and 48 h. At 12 h posttreatment, mRNA abundance of topoisomerase II alpha (TOP2A), and thioredoxin‐dependent peroxidase 2 was upregulated and NPY was downregulated in response to NPY (0 vs. 100 nmol/L) in preadipocytes. Cells were also treated with NPY during differentiation and harvested at 8, 10, and 12 days postinduction of differentiation. At day 8 postinduction of differentiation, there was increased lipid accumulation (0 vs. 10 and 100 nmol/L), expression of CCAAT/enhancer binding protein β and fatty acid binding protein 4 (FABP4) (0 vs. 100 nmol/L), and sterol regulatory element‐binding protein (0 vs. 10 and 100 nmol/L) mRNA in NPY‐treated cells. The number of proliferating cells decreased on day 8 in response to NPY (0 vs. 10 nmol/L). At day 10, FABP4 and Kruppel‐like factor 7 mRNAs were downregulated (0 vs. 10 and 100 nmol/L, and 100 nmol/L, respectively), and at day 12, TOP2A mRNA was down‐regulated (0 vs. 100 nmol/L) in response to NPY treatment. Activity of glycerol‐3‐phosphate dehydrogenase (G3PDH) was increased on days 10 and 12 in NPY‐treated cells (0 vs. 100 nmol/L). Increased gene expression of proliferation markers in preadipocytes, and during differentiation increased expression of transcription factors and a fatty acid transporter, increased lipid accumulation, and increased activity of G3PDH suggest that NPY may enhance preadipocyte activity, adipogenesis, and promotes lipid accumulation throughout chicken adipocyte differentiation.

Glycerol-3-phosphate dehydrogenase 1 deficiency induces compensatory amino acid metabolism during fasting in mice

BackgroundGlucose is used as an energy source in many organs and obtained from dietary carbohydrates. However, when the external energy supply is interrupted, e.g., during fasting, carbohydrates preserved in the liver and glycogenic precursors derived from other organs are used to maintain blood glucose levels. Glycerol and glycogenic amino acids derived from adipocytes and skeletal muscles are utilized as glycogenic precursors. Glycerol-3-phosphate dehydrogenase 1 (GPD1), an NAD+/NADH-dependent enzyme present in the cytosol, catalyzes the reversible conversion of glycerol-3-phosphate (G3P) to dihydroxyacetone phosphate (DHAP). Since G3P is one of the substrates utilized for gluconeogenesis in the liver, the conversion of G3P to DHAP by GPD1 is essential for maintaining blood glucose levels during fasting. We focused on GPD1 and examined its roles in gluconeogenesis during fasting. MethodsUsing GPD1 null model BALB/cHeA mice (HeA mice), we measured gluconeogenesis from glycerol and the change of blood glucose levels under fasting conditions. We also measured gene expression related to gluconeogenesis in the liver and protein metabolism in skeletal muscle. BALB/cBy mice (By mice) were used as a control. ResultsThe blood glucose levels in the HeA mice were lower than that in the By mice after glycerol administration. Although lack of GPD1 inhibited gluconeogenesis from glycerol, blood glucose levels in the HeA mice after 1–4h of fasting were significantly higher than that in the By mice. Muscle protein synthesis in HeA mice was significantly lower than that in the By mice. Moreover, blood alanine levels and usage of alanine for gluconeogenesis in the liver were significantly higher in the HeA mice than that in the By mice. ConclusionsAlthough these data indicate that a lack of GPD1 inhibits gluconeogenesis from glycerol, chronic GPD1 deficiency may induce an adaptation that enhances gluconeogenesis from glycogenic amino acids.

Plastidic P2 glucose-6P dehydrogenase from poplar is modulated by thioredoxin m-type: Distinct roles of cysteine residues in redox regulation and NADPH inhibition

A cDNA coding for a plastidic P2-type G6PDH isoform from poplar (Populus tremula x tremuloides) has been used to express and purify to homogeneity the mature recombinant protein with a N-terminus His-tag. The study of the kinetic properties of the recombinant enzyme showed an in vitro redox sensing modulation exerted by reduced DTT. The interaction with thioredoxins (TRXs) was then investigated.Five cysteine to serine variants (C145S – C175S – C183S – C195S – C242S) and a variant with a double substitution for Cys175 and Cys183 (C175S/C183S) have been generated, purified and biochemically characterized in order to investigate the specific role(s) of cysteines in terms of redox regulation and NADPH-dependent inhibition.Three cysteine residues (C145, C194, C242) are suggested to have a role in controlling the NADP+ access to the active site, and in stabilizing the NADPH regulatory binding site.Our results also indicate that the regulatory disulfide involves residues Cys175 and Cys183 in a position similar to those of chloroplastic P1-G6PDHs, but the modulation is exerted primarily by TRX m-type, in contrast to P1-G6PDH, which is regulated by TRX f.This unexpected specificity indicates differences in the mechanism of regulation, and redox sensing of plastidic P2-G6PDH compared to chloroplastic P1-G6PDH in higher plants.