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FEMS Microbiology Letters Jul 2014Although DNA is the ultimate repository of biological information, deployment of its instructions is constrained by the metabolic and physiological status of the cell.... (Review)
Review
Although DNA is the ultimate repository of biological information, deployment of its instructions is constrained by the metabolic and physiological status of the cell. To this end, bacteria have evolved intricate devices that connect exogenous signals (e.g. nutrients, physicochemical conditions) with endogenous conditions (metabolic fluxes, biochemical networks) that coordinately influence expression or performance of a large number of cellular functions. The phosphoenolpyruvate:carbohydrate-phosphotransferase system (PTS) is a bacterial multi-protein phosphorylation chain which computes extracellular (e.g. sugars) and intracellular (e.g. phosphoenolpyruvate, nitrogen) signals and translates them into post-translational regulation of target activities through protein-protein interactions. The PTS of Pseudomonas putida KT2440 encompasses one complete sugar (fructose)-related system and the three enzymes that form the so-called nitrogen-related PTS (PTS(N) (tr) ), which lacks connection to transport of substrates. These two PTS branches cross-talk to each other, as the product of the fruB gene (a polyprotein EI-HPr-EIIA) can phosphorylate PtsN (EIIA(N) (tr) ) in vivo. This gives rise to a complex actuator device where diverse physiological inputs are ultimately translated into phosphorylation or not of PtsN (EIIA(N) (tr) ) which, in turn, checks the activity of key metabolic and regulatory proteins. Such a control of bacterial physiology highlights the prominence of biochemical homeostasis over genetic ruling -and not vice versa.
Topics: Adaptation, Physiological; Gene Expression Regulation, Bacterial; Metabolic Networks and Pathways; Phosphoenolpyruvate Sugar Phosphotransferase System; Pseudomonas putida; Signal Transduction
PubMed: 24801646
DOI: 10.1111/1574-6968.12459 -
Cell Proliferation Oct 2023
Topics: Humans; Liver Neoplasms; Mitochondria; Neoplasm Proteins; Cation Transport Proteins; Cyclic AMP Response Element-Binding Protein; Phosphoenolpyruvate Carboxykinase (GTP); Intracellular Signaling Peptides and Proteins
PubMed: 37435980
DOI: 10.1111/cpr.13527 -
European Journal of Nutrition Dec 2022Vitamin A is an essential nutrient with vital biological functions. The present study investigated the effect of different doses of vitamin A palmitate at different...
PURPOSE
Vitamin A is an essential nutrient with vital biological functions. The present study investigated the effect of different doses of vitamin A palmitate at different time intervals on thyroid hormones and glycemic markers.
METHODS
Male rats were administrated vitamin A palmitate at different doses (0, 0.7, 1.5, 3, 6, and 12 mg/kg, oral) and samples were collected at different time intervals of 2, 4, and 6 weeks. The levels of vitamin A, thyroid hormones (T3, T4, and TSH), deiodinases (Dio1 and Dio3), glycemic markers (blood insulin and fasting glucose levels, HOMA IR and HOMA β), retinol-binding protein 4 (RBP4) and the gluconeogenic enzyme phosphoenolpyruvate carboxykinase (PEPCK) were measured.
RESULTS
The findings demonstrated that long-term supplementation with high doses of vitamin A palmitate resulted in hypothyroidism (lower T3 and T4 levels and elevated TSH levels) as well as upregulation of Dio1 and Dio3 expression levels. This effect was associated with elevated glucose and insulin levels, enhanced HOMA IR, and decreased HOMA B index. In addition, prolonged vitamin A supplementation significantly increased RBP4 levels that upregulated the expression of PEPCK.
CONCLUSION
High doses of vitamin A supplementation increased the risk of hypothyroidism, modulated insulin sensitivity, and over a long period, increased the incidence of type 2 diabetes mellitus associated with oxidative stress and hepatitis.
Topics: Male; Rats; Animals; Insulin Resistance; Rats, Wistar; Vitamin A; Iodide Peroxidase; Phosphoenolpyruvate; Diabetes Mellitus, Type 2; Blood Glucose; Hypothyroidism; Glucose; Thyroid Hormones; Thyrotropin; Dietary Supplements; Insulins; Insulin
PubMed: 35804266
DOI: 10.1007/s00394-022-02945-5 -
Journal of Molecular Microbiology and... 2015The phosphoenolpyruvate:sugar phosphotransferase system (PTS) has been shown to control transport, cell metabolism and gene expression. We here present results... (Review)
Review
The phosphoenolpyruvate:sugar phosphotransferase system (PTS) has been shown to control transport, cell metabolism and gene expression. We here present results supporting the novel suggestion that in certain instances it also regulates the mutation rate. Directed mutations are defined as mutations that occur at higher frequencies when beneficial than when neutral or detrimental. To date, the occurrence of directed point mutations has not been documented and confirmed, but a few examples of transposon-mediated directed mutations have been reported. Here we focus on the first and best-studied example of directed mutation, which involves the regulation of insertion sequence-5 hopping into a specific site upstream of the glpFK glycerol utilization operon in Escherichia coli. This insertional event specifically activates expression of the glpFK operon, allowing the growth of wild-type cells with glycerol as a carbon source in the presence of nonmetabolizable glucose analogues which normally block glycerol utilization. The sugar-transporting PTS controls this process by regulating levels of cytoplasmic glycerol-3-phosphate and cyclic (c)AMP as established in previous publications. Direct involvement of the glycerol repressor, GlpR, and the cAMP receptor protein, Crp, in the regulation of transposon-mediated directed mutation has been demonstrated.
Topics: Cyclic AMP Receptor Protein; DNA Transposable Elements; Escherichia coli; Escherichia coli Proteins; Gene Expression Regulation, Bacterial; Glucose; Glycerol; Mutagenesis, Insertional; Mutation; Operon; Phosphoenolpyruvate; Phosphoenolpyruvate Sugar Phosphotransferase System; Promoter Regions, Genetic; Repressor Proteins
PubMed: 26159081
DOI: 10.1159/000375375 -
Frontiers in Bioengineering and... 2020Wild-type ATCC 13032 is known to possess two enzymes with anaplerotic (C4-directed) carboxylation activity, namely phosphoenolpyruvate carboxylase (PEPCx) and pyruvate...
Wild-type ATCC 13032 is known to possess two enzymes with anaplerotic (C4-directed) carboxylation activity, namely phosphoenolpyruvate carboxylase (PEPCx) and pyruvate carboxylase (PCx). On the other hand, C3-directed decarboxylation can be catalyzed by the three enzymes phosphoenolpyruvate carboxykinase (PEPCk), oxaloacetate decarboxylase (ODx), and malic enzyme (ME). The resulting high metabolic flexibility at the anaplerotic node compromises the unambigous determination of its carbon and energy flux in wild type. To circumvent this problem we performed a comprehensive analysis of selected single or double deletion mutants in the anaplerosis of wild-type under defined d-glucose conditions. By applying well-controlled lab-scale bioreactor experiments in combination with untargeted proteomics, quantitative metabolomics and whole-genome sequencing hitherto unknown, and sometimes counter-intuitive, genotype-phenotype relationships in these mutants could be unraveled. In comparison to the wild type the four mutants Δ Δ Δ Δ Δ, and Δ showed lowered specific growth rates and d-glucose uptake rates, underlining the importance of PCx and PEPCk activity for a balanced carbon and energy flux at the anaplerotic node. Most interestingly, the strain Δ Δ could be evolved to grow on d-glucose as the only source of carbon and energy, whereas this combination was previously considered lethal. The prevented anaplerotic carboxylation activity of PEPCx and PCx was found in the evolved strain to be compensated by an up-regulation of the glyoxylate shunt, potentially in combination with the 2-methylcitrate cycle.
PubMed: 33553115
DOI: 10.3389/fbioe.2020.602936 -
Cells Dec 2019Changes in phosphoenolpyruvate (PEP) concentrations secondary to variations in glucose availability can regulate calcium signaling in T cells as this metabolite potently...
Changes in phosphoenolpyruvate (PEP) concentrations secondary to variations in glucose availability can regulate calcium signaling in T cells as this metabolite potently inhibits the sarcoplasmic reticulum Ca/ATPase pump (SERCA). This regulation is critical to assert immune activation in the tumor as T cells and cancer cells compete for available nutrients. We examined here whether cytosolic calcium and the activation of downstream effector pathways important for tumor biology are influenced by the presence of glucose and/or cataplerosis through the phosphoenolpyruvate carboxykinase (PEPCK) pathway, as both are hypothesized to feed the PEP pool. Our data demonstrate that cellular PEP parallels extracellular glucose in two human colon carcinoma cell lines, HCT-116 and SW480. PEP correlated with cytosolic calcium and NFAT activity, together with transcriptional up-regulation of canonical targets PTGS2 and IL6 that was fully prevented by CsA pre-treatment. Similarly, loading the metabolite directly into the cell increased cytosolic calcium and NFAT activity. PEP-stirred cytosolic calcium was also responsible for the calmodulin (CaM) dependent phosphorylation of c-Myc at Ser62, resulting in increased activity, probably through enhanced stabilization of the protein. Protein expression of several c-Myc targets also correlated with PEP levels. Finally, the participation of PEPCK in this axis was interrogated as it should directly contribute to PEP through cataplerosis from TCA cycle intermediates, especially in glucose starvation conditions. Inhibition of PEPCK activity showed the expected regulation of PEP and calcium levels and consequential downstream modulation of NFAT and c-Myc activities. Collectively, these results suggest that glucose and PEPCK can regulate NFAT and c-Myc activities through their influence on the PEP/Ca axis, advancing a role for PEP as a second messenger communicating metabolism, calcium cell signaling, and tumor biology.
Topics: Calcium; Calcium Signaling; Cell Line, Tumor; Colonic Neoplasms; Cyclooxygenase 2; Cytosol; Gene Expression Regulation, Neoplastic; Glycolysis; HCT116 Cells; Humans; Interleukin-6; NFATC Transcription Factors; Phosphoenolpyruvate; Phosphoenolpyruvate Carboxykinase (ATP); Proto-Oncogene Proteins c-myc
PubMed: 31861674
DOI: 10.3390/cells9010018 -
Biochemistry Feb 2018
Topics: Anti-Bacterial Agents; Biosynthetic Pathways; Escherichia coli; Fosfomycin; Genes, Bacterial; Methylation; Methyltransferases; Phosphoenolpyruvate; S-Adenosylmethionine; Streptomyces; Substrate Specificity
PubMed: 29345912
DOI: 10.1021/acs.biochem.7b01281 -
Endocrinology and Metabolism (Seoul,... Feb 2024Sodium-dependent glucose cotransporter 2 (SGLT2) mediates glucose reabsorption in the renal proximal tubules, and SGLT2 inhibitors are used as therapeutic agents for...
BACKGRUOUND
Sodium-dependent glucose cotransporter 2 (SGLT2) mediates glucose reabsorption in the renal proximal tubules, and SGLT2 inhibitors are used as therapeutic agents for treating type 2 diabetes mellitus. This study aimed to elucidate the effects and mechanisms of SGLT2 inhibition on hepatic glucose metabolism in both serum deprivation and serum supplementation states.
METHODS
Huh7 cells were treated with the SGLT2 inhibitors empagliflozin and dapagliflozin to examine the effect of SGLT2 on hepatic glucose uptake. To examine the modulation of glucose metabolism by SGLT2 inhibition under serum deprivation and serum supplementation conditions, HepG2 cells were transfected with SGLT2 small interfering RNA (siRNA), cultured in serum-free Dulbecco's modified Eagle's medium for 16 hours, and then cultured in media supplemented with or without 10% fetal bovine serum for 8 hours.
RESULTS
SGLT2 inhibitors dose-dependently decreased hepatic glucose uptake. Serum deprivation increased the expression levels of the gluconeogenesis genes peroxisome proliferator-activated receptor gamma co-activator 1 alpha (PGC-1α), glucose 6-phosphatase (G6pase), and phosphoenolpyruvate carboxykinase (PEPCK), and their expression levels during serum deprivation were further increased in cells transfected with SGLT2 siRNA. SGLT2 inhibition by siRNA during serum deprivation induces nuclear localization of the transcription factor forkhead box class O 1 (FOXO1), decreases nuclear phosphorylated-AKT (p-AKT), and p-FOXO1 protein expression, and increases phosphorylated-adenosine monophosphate-activated protein kinase (p-AMPK) protein expression. However, treatment with the AMPK inhibitor, compound C, reversed the reduction in the protein expression levels of nuclear p- AKT and p-FOXO1 and decreased the protein expression levels of p-AMPK and PEPCK in cells transfected with SGLT2 siRNA during serum deprivation.
CONCLUSION
These data show that SGLT2 mediates glucose uptake in hepatocytes and that SGLT2 inhibition during serum deprivation increases gluconeogenesis via the AMPK/AKT/FOXO1 signaling pathway.
Topics: Humans; AMP-Activated Protein Kinases; Diabetes Mellitus, Type 2; Gluconeogenesis; Glucose; Phosphoenolpyruvate Carboxykinase (ATP); Proto-Oncogene Proteins c-akt; RNA, Small Interfering; Signal Transduction; Sodium; Sodium-Glucose Transporter 2; Sodium-Glucose Transporter 2 Inhibitors
PubMed: 38171209
DOI: 10.3803/EnM.2023.1786 -
Scientific Reports Feb 2020Phosphoenolpyruvate carboxylase (PEPc) is an essential enzyme in plants. A photosynthetic form is present both as dimer and tetramer in C4 and CAM metabolism....
Phosphoenolpyruvate carboxylase (PEPc) is an essential enzyme in plants. A photosynthetic form is present both as dimer and tetramer in C4 and CAM metabolism. Additionally, non-photosynthetic PEPcs are also present. The single, non-photosynthetic PEPc of the unicellular cyanobacterium Synechococcus PCC 7002 (Synechococcus), involved in the TCA cycle, was examined. Using size exclusion chromatography (SEC) and small angle X-ray scattering (SAXS), we observed that PEPc in Synechococcus exists as both a dimer and a tetramer. This is the first demonstration of two different oligomerization states of a non-photosynthetic PEPc. High concentration of Mg, the substrate PEP and a combination of low concentration of Mg and HCO induced the tetramer form of the carboxylase. Using SEC-SAXS analysis, we showed that the oligomerization state of the carboxylase is concentration dependent and that, among the available crystal structures of PEPc, the scattering profile of PEPc of Synechococcus agrees best with the structure of PEPc from Escherichia coli. In addition, the kinetics of the tetramer purified in presence of Mg using SEC, and of the mixed population purified in presence of Mg using a Strep-tagged column were examined. Moreover, the enzyme showed interesting allosteric regulation, being activated by succinate and inhibited by glutamine, and not affected by either malate, 2-oxoglutarate, aspartic acid or citric acid.
Topics: Allosteric Regulation; Bacterial Proteins; Chromatography, Gel; Crystallization; Crystallography, X-Ray; Dimerization; Escherichia coli; Glutamine; Magnesium; Phosphoenolpyruvate Carboxylase; Protein Conformation; Scattering, Small Angle; Synechococcus
PubMed: 32107404
DOI: 10.1038/s41598-020-60249-2 -
Journal of Bacteriology Apr 2015Xylulose 5-phosphate/fructose 6-phosphate phosphoketolase (Xfp), which catalyzes the conversion of xylulose 5-phosphate (X5P) or fructose 6-phosphate (F6P) to acetyl...
UNLABELLED
Xylulose 5-phosphate/fructose 6-phosphate phosphoketolase (Xfp), which catalyzes the conversion of xylulose 5-phosphate (X5P) or fructose 6-phosphate (F6P) to acetyl phosphate, plays a key role in carbohydrate metabolism in a number of bacteria. Recently, we demonstrated that the fungal Cryptococcus neoformans Xfp2 exhibits both substrate cooperativity for all substrates (X5P, F6P, and Pi) and allosteric regulation in the forms of inhibition by phosphoenolpyruvate (PEP), oxaloacetic acid (OAA), and ATP and activation by AMP (K. Glenn, C. Ingram-Smith, and K. S. Smith. Eukaryot Cell 13: 657-663, 2014). Allosteric regulation has not been reported previously for the characterized bacterial Xfps. Here, we report the discovery of substrate cooperativity and allosteric regulation among bacterial Xfps, specifically the Lactobacillus plantarum Xfp. L. plantarum Xfp is an allosteric enzyme inhibited by PEP, OAA, and glyoxylate but unaffected by the presence of ATP or AMP. Glyoxylate is an additional inhibitor to those previously reported for C. neoformans Xfp2. As with C. neoformans Xfp2, PEP and OAA share the same or possess overlapping sites on L. plantarum Xfp. Glyoxylate, which had the lowest half-maximal inhibitory concentration of the three inhibitors, binds at a separate site. This study demonstrates that substrate cooperativity and allosteric regulation may be common properties among bacterial and eukaryotic Xfp enzymes, yet important differences exist between the enzymes in these two domains.
IMPORTANCE
Xylulose 5-phosphate/fructose 6-phosphate phosphoketolase (Xfp) plays a key role in carbohydrate metabolism in a number of bacteria. Although we recently demonstrated that the fungal Cryptococcus Xfp is subject to substrate cooperativity and allosteric regulation, neither phenomenon has been reported for a bacterial Xfp. Here, we report that the Lactobacillus plantarum Xfp displays substrate cooperativity and is allosterically inhibited by phosphoenolpyruvate and oxaloacetate, as is the case for Cryptococcus Xfp. The bacterial enzyme is unaffected by the presence of AMP or ATP, which act as a potent activator and inhibitor of the fungal Xfp, respectively. Our results demonstrate that substrate cooperativity and allosteric regulation may be common properties among bacterial and eukaryotic Xfps, yet important differences exist between the enzymes in these two domains.
Topics: Adenosine Monophosphate; Adenosine Triphosphate; Aldehyde-Lyases; Gene Expression Regulation, Bacterial; Gene Expression Regulation, Enzymologic; Glyoxylates; Hydrogen-Ion Concentration; Lactobacillus plantarum; Oxaloacetic Acid; Pentosephosphates; Phosphoenolpyruvate
PubMed: 25605308
DOI: 10.1128/JB.02380-14