Your browser doesn't support javascript.
loading
Montrer: 20 | 50 | 100
Résultats 1 - 20 de 867
Filtrer
1.
PLoS Pathog ; 20(6): e1011979, 2024 Jun.
Article de Anglais | MEDLINE | ID: mdl-38900808

RÉSUMÉ

The cell surface of Toxoplasma gondii is rich in glycoconjugates which hold diverse and vital functions in the lytic cycle of this obligate intracellular parasite. Additionally, the cyst wall of bradyzoites, that shields the persistent form responsible for chronic infection from the immune system, is heavily glycosylated. Formation of glycoconjugates relies on activated sugar nucleotides, such as uridine diphosphate N-acetylglucosamine (UDP-GlcNAc). The glucosamine-phosphate-N-acetyltransferase (GNA1) generates N-acetylglucosamine-6-phosphate critical to produce UDP-GlcNAc. Here, we demonstrate that downregulation of T. gondii GNA1 results in a severe reduction of UDP-GlcNAc and a concomitant drop in glycosylphosphatidylinositols (GPIs), leading to impairment of the parasite's ability to invade and replicate in the host cell. Surprisingly, attempts to rescue this defect through exogenous GlcNAc supplementation fail to completely restore these vital functions. In depth metabolomic analyses elucidate diverse causes underlying the failed rescue: utilization of GlcNAc is inefficient under glucose-replete conditions and fails to restore UDP-GlcNAc levels in GNA1-depleted parasites. In contrast, GlcNAc-supplementation under glucose-deplete conditions fully restores UDP-GlcNAc levels but fails to rescue the defects associated with GNA1 depletion. Our results underscore the importance of glucosamine-6-phosphate acetylation in governing T. gondii replication and invasion and highlight the potential of the evolutionary divergent GNA1 in Apicomplexa as a target for the development of much-needed new therapeutic strategies.


Sujet(s)
Acétyl-glucosamine , Glucose-6-phosphate , Toxoplasma , Toxoplasma/métabolisme , Glucose-6-phosphate/métabolisme , Glucose-6-phosphate/analogues et dérivés , Acétyl-glucosamine/métabolisme , Acétylation , Animaux , Glucosamine 6-phosphate N-acetyltransferase/métabolisme , Humains , Glucosamine/métabolisme , Glucosamine/analogues et dérivés , Souris , Toxoplasmose/métabolisme , Toxoplasmose/parasitologie , Protéines de protozoaire/métabolisme , Protéines de protozoaire/génétique
2.
Biochem Biophys Res Commun ; 716: 150030, 2024 Jul 05.
Article de Anglais | MEDLINE | ID: mdl-38704889

RÉSUMÉ

Sugar phosphates are potential sources of carbon and phosphate for bacteria. Despite that the process of internalization of Glucose-6-Phosphate (G6P) through plasma membrane remained elusive in several bacteria. VCA0625-27, made of periplasmic ligand binding protein (PLBP) VCA0625, an atypical monomeric permease VCA0626, and a cytosolic ATPase VCA0627, recently emerged as hexose-6-phosphate uptake system of Vibrio cholerae. Here we report high resolution crystal structure of VCA0625 in G6P bound state that largely resembles AfuA of Actinobacillus pleuropneumoniae. MD simulations on VCA0625 in apo and G6P bound states unraveled an 'open to close' and swinging bi-lobal motions, which are diminished upon G6P binding. Mutagenesis followed by biochemical assays on VCA0625 underscored that R34 works as gateway to bind G6P. Although VCA0627 binds ATP, it is ATPase deficient in the absence of VCA0625 and VCA0626, which is a signature phenomenon of type-I ABC importer. Further, modeling, docking and systematic sequence analysis allowed us to envisage the existence of similar atypical type-I G6P importer with fused monomeric permease in 27 other gram-negative bacteria.


Sujet(s)
Protéines bactériennes , Glucose-6-phosphate , Vibrio cholerae , Vibrio cholerae/métabolisme , Vibrio cholerae/génétique , Protéines bactériennes/composition chimique , Protéines bactériennes/métabolisme , Protéines bactériennes/génétique , Cristallographie aux rayons X , Glucose-6-phosphate/métabolisme , Glucose-6-phosphate/composition chimique , Transporteurs ABC/métabolisme , Transporteurs ABC/composition chimique , Transporteurs ABC/génétique , Simulation de dynamique moléculaire , Conformation des protéines , Modèles moléculaires , Liaison aux protéines , Sites de fixation
3.
Plant Mol Biol ; 114(3): 60, 2024 May 17.
Article de Anglais | MEDLINE | ID: mdl-38758412

RÉSUMÉ

Pyruvate kinase (Pyk, EC 2.7.1.40) is a glycolytic enzyme that generates pyruvate and adenosine triphosphate (ATP) from phosphoenolpyruvate (PEP) and adenosine diphosphate (ADP), respectively. Pyk couples pyruvate and tricarboxylic acid metabolisms. Synechocystis sp. PCC 6803 possesses two pyk genes (encoded pyk1, sll0587 and pyk2, sll1275). A previous study suggested that pyk2 and not pyk1 is essential for cell viability; however, its biochemical analysis is yet to be performed. Herein, we biochemically analyzed Synechocystis Pyk2 (hereafter, SyPyk2). The optimum pH and temperature of SyPyk2 were 7.0 and 55 °C, respectively, and the Km values for PEP and ADP under optimal conditions were 1.5 and 0.053 mM, respectively. SyPyk2 is activated in the presence of glucose-6-phosphate (G6P) and ribose-5-phosphate (R5P); however, it remains unaltered in the presence of adenosine monophosphate (AMP) or fructose-1,6-bisphosphate. These results indicate that SyPyk2 is classified as PykA type rather than PykF, stimulated by sugar monophosphates, such as G6P and R5P, but not by AMP. SyPyk2, considering substrate affinity and effectors, can play pivotal roles in sugar catabolism under nonphotosynthetic conditions.


Sujet(s)
Glucose-6-phosphate , Phosphoénolpyruvate , Pyruvate kinase , Ribose monophosphate , Synechocystis , Synechocystis/métabolisme , Synechocystis/génétique , Pyruvate kinase/métabolisme , Pyruvate kinase/génétique , Phosphoénolpyruvate/métabolisme , Glucose-6-phosphate/métabolisme , Ribose monophosphate/métabolisme , Spécificité du substrat , Concentration en ions d'hydrogène , Protéines bactériennes/métabolisme , Protéines bactériennes/génétique , Cinétique , Température
4.
Sci Rep ; 14(1): 10682, 2024 05 09.
Article de Anglais | MEDLINE | ID: mdl-38724517

RÉSUMÉ

Choy Sum, a stalk vegetable highly valued in East and Southeast Asia, is characterized by its rich flavor and nutritional profile. Metabolite accumulation is a key factor in Choy Sum stalk development; however, no research has focused on metabolic changes during the development of Choy Sum, especially in shoot tip metabolites, and their effects on growth and flowering. Therefore, in the present study, we used a widely targeted metabolomic approach to analyze metabolites in Choy Sum stalks at the seedling (S1), bolting (S3), and flowering (S5) stages. In total, we identified 493 metabolites in 31 chemical categories across all three developmental stages. We found that the levels of most carbohydrates and amino acids increased during stalk development and peaked at S5. Moreover, the accumulation of amino acids and their metabolites was closely related to G6P, whereas the expression of flowering genes was closely related to the content of T6P, which may promote flowering by upregulating the expressions of BcSOC1, BcAP1, and BcSPL5. The results of this study contribute to our understanding of the relationship between the accumulation of stem tip substances during development and flowering and of the regulatory mechanisms of stalk development in Choy Sum and other related species.


Sujet(s)
Brassica , Fleurs , Régulation de l'expression des gènes végétaux , Brassica/composition chimique , Brassica/génétique , Brassica/croissance et développement , Brassica/métabolisme , Fleurs/croissance et développement , Fleurs/métabolisme , Métabolome , Tiges de plante/composition chimique , Tiges de plante/croissance et développement , Transcriptome , Glucides , Protéines végétales/génétique , Glucose-6-phosphate/métabolisme , Gènes de plante
5.
New Phytol ; 242(6): 2453-2463, 2024 Jun.
Article de Anglais | MEDLINE | ID: mdl-38567702

RÉSUMÉ

CO2 release in the light (RL) and its presumed source, oxidative pentose phosphate pathways, were found to be insensitive to CO2 concentration. The oxidative pentose phosphate pathways form glucose 6-phosphate (G6P) shunts that bypass the nonoxidative pentose phosphate reactions of the Calvin-Benson cycle. Using adenosine diphosphate glucose and uridine diphosphate glucose as proxies for labeling of G6P in the stroma and cytosol respectively, it was found that only the cytosolic shunt was active. Uridine diphosphate glucose, a proxy for cytosolic G6P, and 6-phosphogluconate (6PG) were significantly less labeled than Calvin-Benson cycle intermediates in the light. But ADP glucose, a proxy for stromal G6P, is labeled to the same degree as Calvin-Benson cycle intermediates and much greater than 6PG. A metabolically inert pool of sedoheptulose bisphosphate can slowly equilibrate keeping the label in sedoheptulose lower than in other stromal metabolites. Finally, phosphorylation of fructose 6-phosphate (F6P) in the cytosol can allow some unlabeled carbon in cytosolic F6P to dilute label in phosphenolpyruvate. The results clearly show that there is oxidative pentose phosphate pathway activity in the cytosol that provides a shunt around the nonoxidative pentose phosphate pathway reactions of the Calvin-Benson cycle and is not strongly CO2-sensitive.


Sujet(s)
Dioxyde de carbone , Oxydoréduction , Voie des pentoses phosphates , Photosynthèse , Dioxyde de carbone/métabolisme , Glucose-6-phosphate/métabolisme , Cytosol/métabolisme , Lumière , Arabidopsis/métabolisme , Arabidopsis/physiologie
6.
Chemistry ; 30(28): e202400690, 2024 May 17.
Article de Anglais | MEDLINE | ID: mdl-38471074

RÉSUMÉ

Droplet formation via liquid-liquid phase separation is thought to be involved in the regulation of various biological processes, including enzymatic reactions. We investigated a glycolytic enzymatic reaction, the conversion of glucose-6-phosphate to 6-phospho-D-glucono-1,5-lactone with concomitant reduction of NADP+ to NADPH both in the absence and presence of dynamically controlled liquid droplet formation. Here, the nucleotide serves as substrate as well as the scaffold required for the formation of liquid droplets. To further expand the process parameter space, temperature and pressure dependent measurements were performed. Incorporation of the reactants in the liquid droplet phase led to a boost in enzymatic activity, which was most pronounced at medium-high pressures. The crowded environment of the droplet phase induced a marked increase of the affinity of the enzyme and substrate. An increase in turnover number in the droplet phase at high pressure contributed to a further strong increase in catalytic efficiency. Enzyme systems that are dynamically coupled to liquid condensate formation may be the key to deciphering many biochemical reactions. Expanding the process parameter space by adjusting temperature and pressure conditions can be a means to further increase the efficiency of industrial enzyme utilization and help uncover regulatory mechanisms adopted by extremophiles.


Sujet(s)
Glucose 6-phosphate dehydrogenase , NADP , Pression , Température , Glucose 6-phosphate dehydrogenase/métabolisme , Glucose 6-phosphate dehydrogenase/composition chimique , NADP/métabolisme , NADP/composition chimique , Glucose-6-phosphate/métabolisme , Glucose-6-phosphate/composition chimique , Gluconates/métabolisme , Gluconates/composition chimique , Lactones/composition chimique , Lactones/métabolisme , Cinétique , Activation enzymatique
7.
Mol Metab ; 79: 101838, 2024 Jan.
Article de Anglais | MEDLINE | ID: mdl-37995884

RÉSUMÉ

OBJECTIVE: Carbohydrate Response Element Binding Protein (ChREBP) is a glucose 6-phosphate (G6P)-sensitive transcription factor that acts as a metabolic switch to maintain intracellular glucose and phosphate homeostasis. Hepatic ChREBP is well-known for its regulatory role in glycolysis, the pentose phosphate pathway, and de novo lipogenesis. The physiological role of ChREBP in hepatic glycogen metabolism and blood glucose regulation has not been assessed in detail, and ChREBP's contribution to carbohydrate flux adaptations in hepatic Glycogen Storage Disease type 1 (GSD I) requires further investigation. METHODS: The current study aimed to investigate the role of ChREBP as a regulator of glycogen metabolism in response to hepatic G6P accumulation, using a model for acute hepatic GSD type Ib. The immediate biochemical and regulatory responses to hepatic G6P accumulation were evaluated upon G6P transporter inhibition by the chlorogenic acid S4048 in mice that were either treated with a short hairpin RNA (shRNA) directed against ChREBP (shChREBP) or a scrambled shRNA (shSCR). Complementary stable isotope experiments were performed to quantify hepatic carbohydrate fluxes in vivo. RESULTS: ShChREBP treatment normalized the S4048-mediated induction of hepatic ChREBP target genes to levels observed in vehicle- and shSCR-treated controls. In parallel, hepatic shChREBP treatment in S4048-infused mice resulted in a more pronounced accumulation of hepatic glycogen and further reduction of blood glucose levels compared to shSCR treatment. Hepatic ChREBP knockdown modestly increased glucokinase (GCK) flux in S4048-treated mice while it enhanced UDP-glucose turnover as well as glycogen synthase and phosphorylase fluxes. Hepatic GCK mRNA and protein levels were induced by shChREBP treatment in both vehicle- and S4048-treated mice, while glycogen synthase 2 (GYS2) and glycogen phosphorylase (PYGL) mRNA and protein levels were reduced. Finally, knockdown of hepatic ChREBP expression reduced starch domain binding protein 1 (STBD1) mRNA and protein levels while it inhibited acid alpha-glucosidase (GAA) activity, suggesting reduced capacity for lysosomal glycogen breakdown. CONCLUSIONS: Our data show that ChREBP activation controls hepatic glycogen and blood glucose levels in acute hepatic GSD Ib through concomitant regulation of glucose phosphorylation, glycogenesis, and glycogenolysis. ChREBP-mediated control of GCK enzyme levels aligns with corresponding adaptations in GCK flux. In contrast, ChREBP activation in response to acute hepatic GSD Ib exerts opposite effects on GYS2/PYGL enzyme levels and their corresponding fluxes, indicating that GYS2/PYGL expression levels are not limiting to their respective fluxes under these conditions.


Sujet(s)
Glycémie , Glycogénose de type I , Animaux , Souris , Métabolisme glucidique , Modèles animaux de maladie humaine , Glucose/métabolisme , Glucose-6-phosphate/métabolisme , Glycogène/métabolisme , Glycogen synthase/métabolisme , Glycogène hépatique/métabolisme , Phosphates , ARN messager/métabolisme , Petit ARN interférent/métabolisme , Facteurs de transcription/génétique , Facteurs de transcription/métabolisme
8.
Protein Expr Purif ; 215: 106408, 2024 Mar.
Article de Anglais | MEDLINE | ID: mdl-38008389

RÉSUMÉ

Hexokinases (HKs) play a vital role in glucose metabolism, which controls the first committed step catalyzing the production of glucose-6-phosphate from glucose. Two HKs (CGIHK1 and CGIHK2) from the Pacific oyster Crassostrea giga were cloned and characterized. CGIHK1 and CGIHK2 were recombinantly expressed in Escherichia coli and successfully purified by the Ni-NTA column. The optimum pH of the two enzymes was pH 8.0 and 8.5, respectively. The optimum temperature of the two enzymes was 42 °C and 50 °C, respectively. Both enzymes showed a clear requirement for divalent magnesium and were strongly inhibited by SDS. CGIHK1 exhibited highly strict substrate specificity to glucose, while CGIHK2 could also catalyze other 11 monosaccharide substrates. This is the first report on the in vitro biosynthesis of glucose-6-phosphate by the hexokinases from Crassostrea gigas. The facile expression and purification procedures combined with different substrate specificities make CGIHK1 and CGIHK2 candidates for the biosynthesis of glucose-6-phosphate and other sugar-phosphates.


Sujet(s)
Crassostrea , Hexokinase , Animaux , Hexokinase/métabolisme , Crassostrea/génétique , Glucose-6-phosphate/métabolisme , Température , Glucose/métabolisme
9.
ACS Chem Biol ; 18(10): 2324-2334, 2023 10 20.
Article de Anglais | MEDLINE | ID: mdl-37793187

RÉSUMÉ

The glmS riboswitch is a motif found in 5'-untranslated regions of bacterial mRNA that controls the synthesis of glucosamine-6-phosphate (GlcN6P), an essential building block for the bacterial cell wall, by a feedback mechanism. Activation of the glmS riboswitch by GlcN6P mimics interferes with the ability of bacteria to synthesize its cell wall. Accordingly, GlcN6P mimics acting as glmS activators are promising candidates for future antibiotic drugs that may overcome emerging bacterial resistance against established antibiotics. We describe the synthesis of a series of phosphonate mimics of GlcN6P as well as the thiasugar analogue of GlcN6P. The phosphonate mimics differ in their pKa value to answer the question of whether derivatives with a pKa matching that of GlcN6P would be efficient glmS activators. We found that all derivatives activate the riboswitch, however, less efficiently than GlcN6P. This observation can be explained by the missing hydrogen bonds in the case of phosphonates and is valuable information for the design of future GlcN6P mimics. The thiasugar analogue of GlcN6P on the other hand turned out to be a glmS riboswitch activator with the same activity as the natural metabolite GlcN6P. The nonphosphorylated thiasugar displayed antimicrobial activity against certain bacilli. Therefore, the compound is a promising lead structure for the development of future antibiotics with a potentially novel mode of action.


Sujet(s)
Phosphonates , ARN catalytique , Riborégulateur , Protéines bactériennes/métabolisme , Phosphonates/pharmacologie , Antibactériens/pharmacologie , Bactéries/métabolisme , Glucosamine , Glucose-6-phosphate/métabolisme , Phosphates , ARN catalytique/composition chimique
10.
J Mol Endocrinol ; 71(4)2023 11 01.
Article de Anglais | MEDLINE | ID: mdl-37855366

RÉSUMÉ

In the endoplasmic reticulum (ER) lumen, glucose-6-phosphatase catalytic subunit 1 and 2 (G6PC1; G6PC2) hydrolyze glucose-6-phosphate (G6P) to glucose and inorganic phosphate whereas hexose-6-phosphate dehydrogenase (H6PD) hydrolyzes G6P to 6-phosphogluconate (6PG) in a reaction that generates NADPH. 11ß-hydroxysteroid dehydrogenase type 1 (HSD11B1) utilizes this NADPH to convert inactive cortisone to cortisol. HSD11B1 inhibitors improve insulin sensitivity whereas G6PC inhibitors are predicted to lower fasting blood glucose (FBG). This study investigated whether G6PC1 and G6PC2 influence G6P flux through H6PD and vice versa. Using a novel transcriptional assay that utilizes separate fusion genes to quantitate glucocorticoid and glucose signaling, we show that overexpression of H6PD and HSD11B1 in the islet-derived 832/13 cell line activated glucocorticoid-stimulated fusion gene expression. Overexpression of HSD11B1 blunted glucose-stimulated fusion gene expression independently of altered G6P flux. While overexpression of G6PC1 and G6PC2 blunted glucose-stimulated fusion gene expression, it had minimal effect on glucocorticoid-stimulated fusion gene expression. In the liver-derived HepG2 cell line, overexpression of H6PD and HSD11B1 activated glucocorticoid-stimulated fusion gene expression but overexpression of G6PC1 and G6PC2 had no effect. In rodents, HSD11B1 converts 11-dehydrocorticosterone (11-DHC) to corticosterone. Studies in wild-type and G6pc2 knockout mice treated with 11-DHC for 5 weeks reveal metabolic changes unaffected by the absence of G6PC2. These data suggest that HSD11B1 activity is not significantly affected by the presence or absence of G6PC1 or G6PC2. As such, G6PC1 and G6PC2 inhibitors are predicted to have beneficial effects by reducing FBG without causing a deleterious increase in glucocorticoid signaling.


Sujet(s)
Glucocorticoïdes , Glucose-6-phosphate , Animaux , Souris , 11-beta-Hydroxysteroid dehydrogenase type 1/génétique , 11-beta-Hydroxysteroid dehydrogenase type 1/métabolisme , Lignée cellulaire , Glucocorticoïdes/pharmacologie , Glucocorticoïdes/métabolisme , Glucose/métabolisme , Glucose-6-phosphate/métabolisme , NADP/métabolisme , Humains
11.
Nat Commun ; 14(1): 3835, 2023 06 28.
Article de Anglais | MEDLINE | ID: mdl-37380648

RÉSUMÉ

Takotsubo cardiomyopathy is a stress-induced cardiovascular disease with symptoms comparable to those of an acute coronary syndrome but without coronary obstruction. Takotsubo was initially considered spontaneously reversible, but epidemiological studies revealed significant long-term morbidity and mortality, the reason for which is unknown. Here, we show in a female rodent model that a single pharmacological challenge creates a stress-induced cardiomyopathy similar to Takotsubo. The acute response involves changes in blood and tissue biomarkers and in cardiac in vivo imaging acquired with ultrasound, magnetic resonance and positron emission tomography. Longitudinal follow up using in vivo imaging, histochemistry, protein and proteomics analyses evidences a continued metabolic reprogramming of the heart towards metabolic malfunction, eventually leading to irreversible damage in cardiac function and structure. The results combat the supposed reversibility of Takotsubo, point to dysregulation of glucose metabolic pathways as a main cause of long-term cardiac disease and support early therapeutic management of Takotsubo.


Sujet(s)
Modèles animaux de maladie humaine , Coeur , Stress psychologique , Syndrome de tako-tsubo , Humains , Femelle , Animaux , Rats , Syndrome de tako-tsubo/métabolisme , Syndrome de tako-tsubo/anatomopathologie , Rat Wistar , Coeur/physiopathologie , Myocytes cardiaques/métabolisme , Myocytes cardiaques/anatomopathologie , Glucose-6-phosphate/métabolisme , Glycolyse , Stress psychologique/complications
12.
Nat Metab ; 4(10): 1287-1305, 2022 10.
Article de Anglais | MEDLINE | ID: mdl-36203054

RÉSUMÉ

Microglial cells consume adenosine triphosphate (ATP) during phagocytosis to clear neurotoxic ß-amyloid in Alzheimer's disease (AD). However, the contribution of energy metabolism to microglial function in AD remains unclear. Here, we demonstrate that hexokinase 2 (HK2) is elevated in microglia from an AD mouse model (5xFAD) and AD patients. Genetic deletion or pharmacological inhibition of HK2 significantly promotes microglial phagocytosis, lowers the amyloid plaque burden and attenuates cognitive impairment in male AD mice. Notably, the ATP level is dramatically increased in HK2-deficient or inactive microglia, which can be attributed to a marked upregulation in lipoprotein lipase (LPL) expression and subsequent increase in lipid metabolism. We further show that two downstream metabolites of HK2, glucose-6-phosphate and fructose-6-phosphate, can reverse HK2-deficiency-induced upregulation of LPL, thus supporting ATP production and microglial phagocytosis. Our findings uncover a crucial role for HK2 in phagocytosis through regulation of microglial energy metabolism, suggesting a potential therapeutic strategy for AD by targeting HK2.


Sujet(s)
Maladie d'Alzheimer , Microglie , Animaux , Souris , Mâle , Microglie/métabolisme , Lipoprotein lipase/métabolisme , Lipoprotein lipase/usage thérapeutique , Hexokinase/génétique , Hexokinase/métabolisme , Hexokinase/usage thérapeutique , Métabolisme lipidique , Adénosine triphosphate/métabolisme , Glucose-6-phosphate/métabolisme , Glucose-6-phosphate/usage thérapeutique , Maladie d'Alzheimer/traitement médicamenteux , Maladie d'Alzheimer/génétique , Maladie d'Alzheimer/métabolisme
13.
Int J Mol Sci ; 23(20)2022 Oct 17.
Article de Anglais | MEDLINE | ID: mdl-36293272

RÉSUMÉ

The reconfiguration of the primary metabolism is essential in plant-pathogen interactions. We compared the local metabolic responses of cucumber leaves inoculated with Pseudomonas syringae pv lachrymans (Psl) with those in non-inoculated systemic leaves, by examining the changes in the nicotinamide adenine dinucleotides pools, the concentration of soluble carbohydrates and activities/gene expression of carbohydrate metabolism-related enzymes, the expression of photosynthesis-related genes, and the tricarboxylic acid cycle-linked metabolite contents and enzyme activities. In the infected leaves, Psl induced a metabolic signature with an altered [NAD(P)H]/[NAD(P)+] ratio; decreased glucose and sucrose contents, along with a changed invertase gene expression; and increased glucose turnover and accumulation of raffinose, trehalose, and myo-inositol. The accumulation of oxaloacetic and malic acids, enhanced activities, and gene expression of fumarase and l-malate dehydrogenase, as well as the increased respiration rate in the infected leaves, indicated that Psl induced the tricarboxylic acid cycle. The changes in gene expression of ribulose-l,5-bis-phosphate carboxylase/oxygenase large unit, phosphoenolpyruvate carboxylase and chloroplast glyceraldehyde-3-phosphate dehydrogenase were compatible with a net photosynthesis decline described earlier. Psl triggered metabolic changes common to the infected and non-infected leaves, the dynamics of which differed quantitatively (e.g., malic acid content and metabolism, glucose-6-phosphate accumulation, and glucose-6-phosphate dehydrogenase activity) and those specifically related to the local or systemic response (e.g., changes in the sugar content and turnover). Therefore, metabolic changes in the systemic leaves may be part of the global effects of local infection on the whole-plant metabolism and also represent a specific acclimation response contributing to balancing growth and defense.


Sujet(s)
Carbon-nitrogen ligases , Cucumis sativus , Pseudomonas syringae/physiologie , Cucumis sativus/génétique , Cucumis sativus/métabolisme , Carbone/métabolisme , Phosphoenolpyruvate carboxylase/génétique , beta-Fructofuranosidase/métabolisme , Malate dehydrogenase/métabolisme , Raffinose/métabolisme , Tréhalose/métabolisme , NAD/métabolisme , Fumarate hydratase , Glucose-6-phosphate/métabolisme , Glucose 6-phosphate dehydrogenase/métabolisme , Feuilles de plante/métabolisme , Photosynthèse/physiologie , Métabolisme glucidique , Saccharose/métabolisme , Phosphates/métabolisme , Oxygénases/métabolisme , Inositol/métabolisme , Carbon-nitrogen ligases/métabolisme , Nicotinamide/métabolisme , Adénine/métabolisme , Glucose/métabolisme
14.
Plant Physiol ; 190(4): 2137-2154, 2022 11 28.
Article de Anglais | MEDLINE | ID: mdl-36111879

RÉSUMÉ

In Arabidopsis (Arabidopsis thaliana), the plastidial isoform of phosphoglucose isomerase (PGI1) mediates photosynthesis, metabolism, and development, probably due to its involvement in the synthesis of isoprenoid-derived signals in vascular tissues. Microbial volatile compounds (VCs) with molecular masses of <45 Da promote photosynthesis, growth, and starch overaccumulation in leaves through PGI1-independent mechanisms. Exposure to these compounds in leaves enhances the levels of GLUCOSE-6-PHOSPHATE/PHOSPHATE TRANSLOCATOR2 (GPT2) transcripts. We hypothesized that the PGI1-independent response to microbial volatile emissions involves GPT2 action. To test this hypothesis, we characterized the responses of wild-type (WT), GPT2-null gpt2-1, PGI1-null pgi1-2, and pgi1-2gpt2-1 plants to small fungal VCs. In addition, we characterized the responses of pgi1-2gpt2-1 plants expressing GPT2 under the control of a vascular tissue- and root tip-specific promoter to small fungal VCs. Fungal VCs promoted increases in growth, starch content, and photosynthesis in WT and gpt2-1 plants. These changes were substantially weaker in VC-exposed pgi1-2gpt2-1 plants but reverted to WT levels with vascular and root tip-specific GPT2 expression. Proteomic analyses did not detect enhanced levels of GPT2 protein in VC-exposed leaves and showed that knocking out GPT2 reduced the expression of photosynthesis-related proteins in pgi1-2 plants. Histochemical analyses of GUS activity in plants expressing GPT2-GUS under the control of the GPT2 promoter showed that GPT2 is mainly expressed in root tips and vascular tissues around hydathodes. Overall, the data indicated that the PGI1-independent response to microbial VCs involves resetting of the photosynthesis-related proteome in leaves through long-distance GPT2 action.


Sujet(s)
Protéines d'Arabidopsis , Arabidopsis , Protéines d'Arabidopsis/génétique , Protéines d'Arabidopsis/métabolisme , Glucose-6-phosphate/métabolisme , Protéomique , Arabidopsis/métabolisme , Glucose 6-phosphate isomerase/métabolisme , Amidon/métabolisme , Glucose/métabolisme , Phosphates/métabolisme
15.
Cell Rep ; 40(13): 111346, 2022 09 27.
Article de Anglais | MEDLINE | ID: mdl-36170813

RÉSUMÉ

Mast cells (MCs) are granulated cells implicated in inflammatory disorders because of their capacity to degranulate, releasing prestored proinflammatory mediators. As MCs have the unique capacity to reform granules following degranulation in vitro, their potential to regranulate in vivo is linked to their pathogenesis. It is not known what factors regulate regranulation, let alone if regranulation occurs in vivo. We report that mice can undergo multiple bouts of MC regranulation following successive anaphylactic reactions. mTORC1, a nutrient sensor that activates protein and lipid synthesis, is necessary for regranulation. mTORC1 activity is regulated by a glucose-6-phosphate transporter, Slc37a2, which increases intracellular glucose-6-phosphate and ATP during regranulation, two upstream signals of mTOR. Additionally, Slc37a2 concentrates extracellular metabolites within endosomes, which are trafficked into nascent granules. Thus, the metabolic switch associated with MC regranulation is mediated by the interactions of a cellular metabolic sensor and a transporter of extracellular metabolites into MC granules.


Sujet(s)
Dégranulation cellulaire , Mastocytes , Adénosine triphosphate/métabolisme , Animaux , Antiports , Glucose/métabolisme , Glucose-6-phosphate/métabolisme , Lipides , Mastocytes/métabolisme , Complexe-1 cible mécanistique de la rapamycine/métabolisme , Souris , Protéines de transport du phosphate/métabolisme
16.
PLoS Pathog ; 18(9): e1010864, 2022 09.
Article de Anglais | MEDLINE | ID: mdl-36121870

RÉSUMÉ

Metabolic pathways underpin the growth and virulence of intracellular parasites and are therefore promising antiparasitic targets. The pentose phosphate pathway (PPP) is vital in most organisms, providing a reduced form of nicotinamide adenine dinucleotide phosphate (NADPH) and ribose sugar for nucleotide synthesis; however, it has not yet been studied in Toxoplasma gondii, a widespread intracellular pathogen and a model protozoan organism. Herein, we show that T. gondii has a functional PPP distributed in the cytoplasm and nucleus of its acutely-infectious tachyzoite stage. We produced eight parasite mutants disrupting seven enzymes of the PPP in T. gondii. Our data show that of the seven PPP proteins, the two glucose-6-phosphate dehydrogenases (TgG6PDH1, TgG6PDH2), one of the two 6-phosphogluconate dehydrogenases (Tg6PGDH1), ribulose-5-phosphate epimerase (TgRuPE) and transaldolase (TgTAL) are dispensable in vitro as well as in vivo, disclosing substantial metabolic plasticity in T. gondii. Among these, TgG6PDH2 plays a vital role in defense against oxidative stress by the pathogen. Further, we show that Tg6PGDH2 and ribulose-5-phosphate isomerase (TgRPI) are critical for tachyzoite growth. The depletion of TgRPI impairs the flux of glucose in central carbon pathways, and causes decreased expression of ribosomal, microneme and rhoptry proteins. In summary, our results demonstrate the physiological need of the PPP in T. gondii while unraveling metabolic flexibility and antiparasitic targets.


Sujet(s)
Voie des pentoses phosphates , Toxoplasma , Antiparasitaires , Carbone/métabolisme , Glucose/métabolisme , Glucose-6-phosphate/métabolisme , Isomerases/métabolisme , NADP/métabolisme , Voie des pentoses phosphates/physiologie , Phosphates/métabolisme , Racémases et épimérases/métabolisme , Ribose , Toxoplasma/métabolisme , Transaldolase/métabolisme
17.
Front Cell Infect Microbiol ; 12: 866729, 2022.
Article de Anglais | MEDLINE | ID: mdl-35795184

RÉSUMÉ

The obligate intracellular bacteria Chlamydia trachomatis store glycogen in the lumen of the vacuoles in which they grow. Glycogen catabolism generates glucose-1-phosphate (Glc1P), while the bacteria can take up only glucose-6-phosphate (Glc6P). We tested whether the conversion of Glc1P into Glc6P could be catalyzed by a phosphoglucomutase (PGM) of host or bacterial origin. We found no evidence for the presence of the host PGM in the vacuole. Two C. trachomatis proteins, CT295 and CT815, are potential PGMs. By reconstituting the reaction using purified proteins, and by complementing PGM deficient fibroblasts, we demonstrated that only CT295 displayed robust PGM activity. Intriguingly, we showed that glycogen accumulation in the lumen of the vacuole of a subset of Chlamydia species (C. trachomatis, C. muridarum, C. suis) correlated with the presence, in CT295 orthologs, of a secretion signal recognized by the type three secretion (T3S) machinery of Shigella. C. caviae and C. pneumoniae do not accumulate glycogen, and their CT295 orthologs lack T3S signals. In conclusion, we established that the conversion of Glc1P into Glc6P was accomplished by a bacterial PGM, through the acquisition of a T3S signal in a "housekeeping" protein. Acquisition of this signal likely contributed to shaping glycogen metabolism within Chlamydiaceae.


Sujet(s)
Chlamydia trachomatis , Phosphoglucomutase , Chlamydia trachomatis/génétique , Chlamydia trachomatis/métabolisme , Glucose-6-phosphate/métabolisme , Glycogène/métabolisme , Phosphoglucomutase/génétique , Phosphoglucomutase/métabolisme , Vacuoles/métabolisme
18.
Nat Struct Mol Biol ; 29(7): 628-638, 2022 07.
Article de Anglais | MEDLINE | ID: mdl-35835870

RÉSUMÉ

Glycogen synthase (GYS1) is the central enzyme in muscle glycogen biosynthesis. GYS1 activity is inhibited by phosphorylation of its amino (N) and carboxyl (C) termini, which is relieved by allosteric activation of glucose-6-phosphate (Glc6P). We present cryo-EM structures at 3.0-4.0 Å resolution of phosphorylated human GYS1, in complex with a minimal interacting region of glycogenin, in the inhibited, activated and catalytically competent states. Phosphorylations of specific terminal residues are sensed by different arginine clusters, locking the GYS1 tetramer in an inhibited state via intersubunit interactions. The Glc6P activator promotes conformational change by disrupting these interactions and increases the flexibility of GYS1, such that it is poised to adopt a catalytically competent state when the sugar donor UDP-glucose (UDP-glc) binds. We also identify an inhibited-like conformation that has not transitioned into the activated state, in which the locking interaction of phosphorylation with the arginine cluster impedes subsequent conformational changes due to Glc6P binding. Our results address longstanding questions regarding the mechanism of human GYS1 regulation.


Sujet(s)
Glucose-6-phosphate , Glycogen synthase , Arginine/métabolisme , Glucose-6-phosphate/métabolisme , Glycogen synthase/composition chimique , Glycogen synthase/métabolisme , Humains , Phosphorylation , Uridine diphosphate/métabolisme
19.
mBio ; 13(4): e0146922, 2022 08 30.
Article de Anglais | MEDLINE | ID: mdl-35856562

RÉSUMÉ

The reactions of α-d-phosphohexomutases (αPHM) are ubiquitous, key to primary metabolism, and essential for several processes in all domains of life. The functionality of these enzymes relies on an initial phosphorylation step which requires the presence of α-d-glucose-1,6-bisphosphate (Glc-1,6-BP). While well investigated in vertebrates, the origin of this activator compound in bacteria is unknown. Here we show that the Slr1334 protein from the unicellular cyanobacterium Synechocysitis sp. PCC 6803 is a Glc-1,6-BP-synthase. Biochemical analysis revealed that Slr1334 efficiently converts fructose-1,6-bisphosphate (Frc-1,6-BP) and α-d-glucose-1-phosphate/α-d-glucose-6-phosphate into Glc-1,6-BP and also catalyzes the reverse reaction. As inferred from phylogenetic analysis, the slr1334 product belongs to a primordial subfamily of αPHMs that is present especially in deeply branching bacteria and also includes human commensals and pathogens. Remarkably, the homologue of Slr1334 in the human gut bacterium Bacteroides salyersiae catalyzes the same reaction, suggesting a conserved and essential role for the members of this αPHM subfamily. IMPORTANCE Glc-1,6-BP is known as an essential activator of phosphoglucomutase (PGM) and other members of the αPHM superfamily, making it a central regulator in glycogen metabolism, glycolysis, amino sugar formation as well as bacterial cell wall and capsule formation. Despite this essential role in carbon metabolism, its origin in prokaryotes has so far remained elusive. In this study we identify a member of a specific αPHM subfamily as the first bacterial Glc-1,6-BP synthase, forming free Glc-1,6-BP by using Frc-1,6-BP as phosphoryl-donor. PGMs of this subfamily are widely distributed among prokaryotes including human commensals and pathogens. By showing that a distinct subfamily member can also form Glc-1,6-BP, we provide evidence that Glc-1,6-BP synthase activity is a general feature of this group.


Sujet(s)
Glucose-6-phosphate , Phosphoglucomutase , Animaux , Glucose , Glucose-6-phosphate/analogues et dérivés , Glucose-6-phosphate/métabolisme , Humains , Phosphoglucomutase/composition chimique , Phosphoglucomutase/génétique , Phosphoglucomutase/métabolisme , Phylogenèse
20.
Plant J ; 111(6): 1643-1659, 2022 09.
Article de Anglais | MEDLINE | ID: mdl-35862290

RÉSUMÉ

Nitrate (NO3 - ) and phosphate (Pi) deficiencies are the major constraints for chickpea productivity, significantly impacting global food security. However, excessive fertilization is expensive and can also lead to environmental pollution. Therefore, there is an urgent need to develop chickpea cultivars that are able to grow on soils deficient in both NO3 - and Pi. This study focused on the identification of key NO3 - and/or Pi starvation-responsive metabolic pathways in the leaves and roots of chickpea grown under single and double nutrient deficiencies of NO3 - and Pi, in comparison with nutrient-sufficient conditions. A global metabolite analysis revealed organ-specific differences in the metabolic adaptation to nutrient deficiencies. Moreover, we found stronger adaptive responses in the roots and leaves to any single than combined nutrient-deficient stresses. For example, chickpea enhanced the allocation of carbon among nitrogen-rich amino acids (AAs) and increased the production of organic acids in roots under NO3 - deficiency, whereas this adaptive response was not found under double nutrient deficiency. Nitrogen remobilization through the transport of AAs from leaves to roots was greater under NO3 - deficiency than double nutrient deficiency conditions. Glucose-6-phosphate and fructose-6-phosphate accumulated in the roots under single nutrient deficiencies, but not under double nutrient deficiency, and higher glycolytic pathway activities were observed in both roots and leaves under single nutrient deficiency than double nutrient deficiency. Hence, the simultaneous deficiency generated a unique profile of metabolic changes that could not be simply described as the result of the combined deficiencies of the two nutrients.


Sujet(s)
Cicer , Acides aminés/métabolisme , Carbone/métabolisme , Cicer/métabolisme , Glucose-6-phosphate/métabolisme , Nitrates/métabolisme , Azote/métabolisme , Phosphates/métabolisme , Racines de plante/métabolisme , Sol
SÉLECTION CITATIONS
DÉTAIL DE RECHERCHE
...