'Sugarcoating' 2-deoxyglucose: mechanisms that suppress its toxic effects.

Yeast and cancer cells are metabolically similar as they use fermentation of glucose as a primary means of generating energy. Reliance on glucose fermentation makes both of these cell types highly sensitive to the toxic glucose analog, 2-deoxyglucose. Here we review the cellular and metabolic pathways that play a role in 2-deoxyglucose sensitivity and discuss how the modifications to these pathways result in acquisition of 2-deoxyglucose resistance. Insights gained from genetic and proteomic studies in yeast provide new ideas for the design of combinatorial therapies for cancer treatment.

Convergent evolution of diverse Bacillus anthracis outbreak strains toward altered surface oligosaccharides that modulate anthrax pathogenesis.

Bacillus anthracis, a spore-forming gram-positive bacterium, causes anthrax. The external surface of the exosporium is coated with glycosylated proteins. The sugar additions are capped with the unique monosaccharide anthrose. The West African Group (WAG) B. anthracis have mutations rendering them anthrose deficient. Through genome sequencing, we identified 2 different large chromosomal deletions within the anthrose biosynthetic operon of B. anthracis strains from Chile and Poland. In silico analysis identified an anthrose-deficient strain in the anthrax outbreak among European heroin users. Anthrose-deficient strains are no longer restricted to West Africa so the role of anthrose in physiology and pathogenesis was investigated in B. anthracis Sterne. Loss of anthrose delayed spore germination and enhanced sporulation. Spores without anthrose were phagocytized at higher rates than spores with anthrose, indicating that anthrose may serve an antiphagocytic function on the spore surface. The anthrose mutant had half the LD50 and decreased time to death (TTD) of wild type and complement B. anthracis Sterne in the A/J mouse model. Following infection, anthrose mutant bacteria were more abundant in the spleen, indicating enhanced dissemination of Sterne anthrose mutant. At low sample sizes in the A/J mouse model, the mortality of ΔantC-infected mice challenged by intranasal or subcutaneous routes was 20% greater than wild type. Competitive index (CI) studies indicated that spores without anthrose disseminated to organs more extensively than a complemented mutant. Death process modeling using mouse mortality dynamics suggested that larger sample sizes would lead to significantly higher deaths in anthrose-negative infected animals. The model was tested by infecting Galleria mellonella with spores and confirmed the anthrose mutant was significantly more lethal. Vaccination studies in the A/J mouse model showed that the human vaccine protected against high-dose challenges of the nonencapsulated Sterne-based anthrose mutant. This work begins to identify the physiologic and pathogenic consequences of convergent anthrose mutations in B. anthracis.

Nucleotide polymorphism assay for the identification of west African group Bacillus anthracis: a lineage lacking anthrose.

BACKGROUND: The exosporium of the anthrax-causing Bacillus anthracis endospores display a tetrasaccharide composed of three rhamnose residues and an unusual sugar termed anthrose. Anthrose is a proposed potential target for immunotherapy and for specific detection of B. anthracis. Although originally thought to be ubiquitous in B. anthracis, previous work identified an anthrose negative strain from a West African lineage isolated from cattle that could represent a vaccine escape mutant. These strains carry genes required for expression of the anthrose operon but premature stop codons resulting from an 8-bp insertion in BAS3320 (an amino-transferase) and a C/T substitution at position 892 of the BAS3321 (a glycosyltransferase) gene prevent anthrose expression. Various other single nucleotide polymorphisms (SNPs) have been identified throughout the operon and could be the basis for detection of anthrose-deficient strains. RESULTS: In this study, we evaluated rhAmp genotypic assays based on SNPs at positions 892 and 1352 of BAS3321 for detection and differentiation of anthrose negative (Ant-) West African strains. Discrimination of anthrose negative West African isolates was achieved with as low as 100 fg of DNA, whereas consistent genotyping of Sterne necessitated at least 1 pg of DNA. CONCLUSIONS: Screening of a global panel of B. anthracis isolates showed anthrose-expressing alleles are prevalent worldwide whereas the anthrose-deficient phenotype is to date limited to West Africa. Our work also revealed a third, previously unreported anthrose genotype in which the operon is altogether missing from a Polish B. anthracis isolate.

Pyrraline Formation Modulated by Sodium Chloride and Controlled by Encapsulation with Different Coating Materials in the Maillard Reaction.

Advanced glycation end products (AGEs), which are present in heat-processed foods, have been associated with several chronic diseases. Sodium chloride (NaCl) modulates the formation of furfurals and acrylamide in the Maillard reaction; however, the effects of NaCl on AGE formation are inconsistent. In this study, we investigated the effects of NaCl on pyrraline formation using glucose-lysine model systems. NaCl, especially at 0.50%, promoted Maillard browning and pyrraline formation, with a simultaneous increase in the 3-deoxyglucosone concentration. To reduce the rate of pyrraline formation, NaCl coated with different gums and starches were used. The results showed that NaCl encapsulation is an effective approach to mitigate pyrraline and 3-deoxyglucosone formation. The content of NaCl in the microparticles were 284 ± 12, 269 ± 6, 258 ± 8, 247 ± 10, 273 ± 16, and 288 ± 15 mg/g (coated with waxy maize starch, normal maize starch, HYLON VII high amylose maize starch, gelatinized resistant starch, xanthan gum, and gum arabic, respectively). The heat resistance of the coating material was negatively correlated with the pyrraline and 3-deoxyglucosone formation, whereas the solubility of the coating material had the opposite results. Coating the material with gum had little effects on the reduction of pyrraline and 3-deoxyglucosone.

Immobilization rapidly induces thioredoxin-interacting protein gene expression together with insulin resistance in rat skeletal muscle.

Acute short duration of disuse induces the development of insulin resistance for glucose uptake in rodent skeletal muscle. Because thioredoxin-interacting protein (TXNIP) has been implicated in the downregulation of insulin signaling and glucose uptake, we examined the possibility that muscle disuse rapidly induces insulin resistance via increased TXNIP mRNA and protein expression. Male Wistar rats were subjected to unilateral 6-h hindlimb immobilization by plaster cast. At the end of this period, the soleus muscles from both immobilized and contralateral nonimmobilized hindlimbs were excised and examined. The 6-h immobilization resulted in an increase in TXNIP mRNA and protein expressions together with a decrease in insulin-stimulated 2-deoxyglucose uptake in the rat soleus muscle. Additionally, in the rats euthanized 6 h after the plaster cast removal, TXNIP protein expression and insulin-stimulated glucose uptake in the immobilized muscle had both been restored to a normal level. Various interventions (pretreatment with transcription inhibitor actinomycin D or AMP-dependent protein kinase activator 5-aminoimidazole-4-carboxamide ribonucleotide) also suppressed the increase in TXNIP protein expression in 6-h-immobilized muscle together with partial prevention of insulin resistance for glucose uptake. These results suggested the possibility that increased TXNIP protein expression in immobilized rat soleus muscles was associated with the rapid induction of insulin resistance for glucose uptake in that tissue. NEW & NOTEWORTHY The cellular mechanism by which disuse rapidly induces muscle insulin resistance for glucose uptake remains to be identified. Using a rat hindlimb immobilization model, our findings suggest the possibility that transcriptional upregulation of thioredoxin-interacting protein is associated with the immobilization-induced rapid development of insulin resistance in skeletal muscle.

Genome-wide association study of 1,5-anhydroglucitol identifies novel genetic loci linked to glucose metabolism.

1,5-anhydroglucitol (1,5-AG) is a biomarker of hyperglycemic excursions associated with diabetic complications. Because of its structural similarity to glucose, genetic studies of 1,5-AG can deliver complementary insights into glucose metabolism. We conducted genome-wide association studies of serum 1,5-AG concentrations in 7,550 European ancestry (EA) and 2,030 African American participants (AA) free of diagnosed diabetes from the ARIC Study. Seven loci in/near EFNA1/SLC50A1, MCM6/LCT, SI, MGAM, MGAM2, SLC5A10, and SLC5A1 showed genome-wide significant associations (P < 5 × 10-8) among EA participants, five of which were novel. Six of the seven loci were successfully replicated in 8,790 independent EA individuals, and MCM6/LCT and SLC5A10 were also associated among AA. Most of 1,5-AG-associated index SNPs were not associated with the clinical glycemic markers fasting glucose or the  HbA1c, and vice versa. Only the index variant in SLC5A1 showed a significant association with fasting glucose in the expected opposing direction. Products of genes in all 1,5-AG-associated loci have known roles in carbohydrate digestion and enteral or renal glucose transport, suggesting that genetic variants associated with 1,5-AG influence its concentration via effects on glucose metabolism and handling.

A Kluyveromyces marxianus 2-deoxyglucoseresistant mutant with enhanced activity of xylose utilization

Thermotolerant ethanologenic yeast Kluyveromyces marxianus is capable of fermenting various sugars including xylose but glucose represses to hamper the utilization of other sugars. To acquire glucose repression-defective strains, 33 isolates as 2-deoxyglucose (2-DOG)-resistant mutants were acquired from about 100 colonies grown on plates containing 2-DOG, which were derived from an efficient strain DMKU 3-1042. According to the characteristics of sugar consumption abilities and cell growth and ethanol accumulation along with cultivation time, they were classified into three groups. The first group (3 isolates) utilized glucose and xylose in similar patterns along with cultivation to those of the parental strain, presumably due to reduction of the uptake of 2-DOG or enhancement of its export. The second group (29 isolates) showed greatly delayed utilization of glucose, presumably by reduction of the uptake or initial catabolism of glucose. The last group, only one isolate, showed enhanced utilization ability of xylose in the presence of glucose. Further analysis revealed that the isolate had a single nucleotide mutation to cause amino acid substitution (G270S) in RAG5 encoding hexokinase and exhibited very low activity of the enzyme. The possible mechanism of defectiveness of glucose repression in the mutant is discussed in this paper (AU)

No disponible

The wonders of 2-deoxy-D-glucose.

Through the eons of time, out of all possible configurations, nature has selected glucose not only as a vital source of energy to sustain life but also as the molecule who's structure supplies the appropriate elements required for a cell to grow and multiply. This understanding, at least in part, explains the profound effects that the analog of glucose, 2-deoxy-d-glucose, has been shown to have on as common and widespread diseases as cancer, viral infection, aging-related morbidity, epilepsy, and others. This review is confined to summarizing some of the salient findings of this remarkable compound as they relate mainly to cancer.

The O-specific polysaccharide structure and gene cluster of serotype O:12 of the Yersinia pseudotuberculosis complex, and the identification of a novel L-quinovose biosynthesis gene.

A major virulence factor for Yersinia pseudotuberculosis is lipopolysaccharide, including O-polysaccharide (OPS). Currently, the OPS based serotyping scheme for Y. pseudotuberculosis includes 21 known O-serotypes, with genetic and structural data available for 17 of them. The completion of the OPS structures and genetics of this species will enable the visualization of relationships between O-serotypes and allow for analysis of the evolutionary processes within the species that give rise to new serotypes. Here we present the OPS structure and gene cluster of serotype O:12, thus adding one more to the set of completed serotypes, and show that this serotype is present in both Y. pseudotuberculosis and the newly identified Y. similis species. The O:12 structure is shown to include two rares ugars: 4-C[(R)-1-hydroxyethyl]-3,6-dideoxy-D-xylo-hexose(D-yersiniose) and 6-deoxy-L-glucopyranose (L-quinovose).We have identified a novel putative guanine diphosphate(GDP)-L-fucose 4-epimerase gene and propose a pathway for the synthesis of GDP-L-quinovose, which extends the known GDP-L-fucose pathway.

Oral advanced glycation endproducts (AGEs) promote insulin resistance and diabetes by depleting the antioxidant defenses AGE receptor-1 and sirtuin 1.

The epidemics of insulin resistance (IR) and type 2 diabetes (T2D) affect the first world as well as less-developed countries, and now affect children as well. Persistently elevated oxidative stress and inflammation (OS/Infl) precede these polygenic conditions. A hallmark of contemporary lifestyle is a preference for thermally processed nutrients, replete with pro-OS/Infl advanced glycation endproducts (AGEs), which enhance appetite and cause overnutrition. We propose that chronic ingestion of oral AGEs promotes IR and T2D. The mechanism(s) involved in these findings were assessed in four generations of C57BL6 mice fed isocaloric diets with or without AGEs [synthetic methyl-glyoxal-derivatives (MG(+))]. F3/MG(+) mice manifested increased adiposity and premature IR, marked by severe deficiency of anti-AGE advanced glycation receptor 1 (AGER1) and of survival factor sirtuin 1 (SIRT1) in white adipose tissue (WAT), skeletal muscle, and liver. Impaired 2-deoxy-glucose uptake was associated with marked changes in insulin receptor (InsR), IRS-1, IRS-2, Akt activation, and a macrophage and adipocyte shift to a pro-OS/inflammatory (M1) phenotype. These features were absent in F3/MG(-) mice. MG stimulation of 3T3-L1 adipocytes led to suppressed AGER1 and SIRT1, and altered InsR, IRS-1, IRS-2 phosphorylation, and nuclear factor kappa-light chain enhancer of activated B cells (Nf-κB) p65 acetylation. Gene modulation revealed these effects to be coregulated by AGER1 and SIRT1. Thus, prolonged oral exposure to MG-AGEs can deplete host-defenses AGER1 and SIRT1, raise basal OS/Infl, and increase susceptibility to dysmetabolic IR. Because exposure to AGEs can be decreased, these insights provide an important framework for alleviating a major lifestyle-linked disease epidemic.

Characterization of the enzymes encoded by the anthrose biosynthetic operon of Bacillus anthracis.

Bacillus anthracis spores, the etiological agents of anthrax, possess a loosely fitting outer layer called the exosporium that is composed of a basal layer and an external hairlike nap. The filaments of the nap are formed by trimers of the collagenlike glycoprotein BclA. Multiple pentasaccharide and trisaccharide side chains are O linked to BclA. The nonreducing terminal residue of the pentasaccharide side chain is the unusual sugar anthrose. A plausible biosynthetic pathway for anthrose biosynthesis has been proposed, and an antABCD operon encoding four putative anthrose biosynthetic enzymes has been identified. In this study, we genetically and biochemically characterized the activities of these enzymes. We also used mutant B. anthracis strains to determine the effects on BclA glycosylation of individually inactivating the genes of the anthrose operon. The inactivation of antA resulted in the appearance of BclA pentasaccharides containing anthrose analogs possessing shorter side chains linked to the amino group of the sugar. The inactivation of antB resulted in BclA being replaced with only trisaccharides, suggesting that the enzyme encoded by the gene is a dTDP-ß-L-rhamnose α-1,3-L-rhamnosyl transferase that attaches the fourth residue of the pentasaccharide side chain. The inactivation of antC and antD resulted in the disappearance of BclA pentasaccharides and the appearance of a tetrasaccharide lacking anthrose. These phenotypes are entirely consistent with the proposed roles for the antABCD-encoded enzymes in anthrose biosynthesis. Purified AntA was then shown to exhibit ß-methylcrotonyl-coenzyme A (CoA) hydratase activity, as we predicted. Similarly, we confirmed that purified AntC had aminotransferase activity and that purified AntD displayed N-acyltransferase activity.

Normotopic and heterotopic cortical representations of mystacial vibrissae in rats with subcortical band heterotopia.

The tish rat is a neurological mutant exhibiting bilateral cortical heterotopia similar to those found in certain epileptic patients. Previous work has shown that thalamocortical fibers originating in the ventroposteromedial nucleus, which in normal animals segregate as 'barrel' representations for individual whiskers, terminate in both normotopic and heterotopic areas of the tish cortex (Schottler et al., 1998). Thalamocortical innervation terminates as barrels in layer IV and diffusely in layer VI of the normotopic area. Discrete patches of terminals are also observed in the underlying heterotopic area suggesting that representations of individual vibrissa may be present in the heterotopic somatosensory areas. The present study examines this issue by investigating the organization of the vibrissal somatosensory system in the tish cortex. Staining for cytochrome oxidase or Nissl substance reveals a normal complement of vibrissal barrels in the normotopic area of the tish cortex. Dense patches of cytochrome oxidase staining are also found in the underlying lateral portions of the heterotopic area (i.e. the same area that is innervated by the ventroposteromedial nucleus). Injections of retrograde tracers into vibrissal areas of either the normotopic or heterotopic area produce topographically organized labeling of neurons restricted to one or a small number of barreloids within the ventroposteromedial nucleus of the thalamus. Physical stimulation of a single whisker (D3 or E3) elicits enhanced uptake of [(14)C]2-deoxyglucose in restricted zones of both the normotopic and heterotopic areas, demonstrating that single whisker stimulation can increase functional activity in both normotopic and heterotopic neurons. These findings indicate that the barrels are intact in the normotopic area and are most consistent with the hypothesis that at least some of the individual vibrissae are 'dually' represented in normotopic and heterotopic positions in the primary somatosensory areas of the tish cortex.

Comparison of industrial yeast strains for fermentation of spent sulphite pulping liquor fortified with wood hydrolysate.

Ethanol production from spent sulphite pulping liquor (SSL) was compared for four different yeasts. A second strain of S. cerevisiae as well as a 2-deoxyglucose-resistant strain formed through protoplast fusions between S. uvarum and S. diastaticus produced up to 27% more ethanol from SSL fortified with hydrolysis sugars than was produced by S. cerevisiae. The incremental improvement in ethanol yield appeared to vary with the degree of fortification, ranging from 5.8% for unfortified SSL, to 27% for the highest level of fortification tested. Decreasing fermentation rates were observed for SSL fortified with glucose, mannose and galactose, respectively. Sugar uptake rates in SSL fortified with glucose, galactose and mannose were 6.8, 2.8 and 2.0 g L-1 h-1, respectively. However, when these sugars were fermented along with a glucose cosubstrate, the rate at which the combined glucose/mannose medium was fermented was nearly identical to that of the glucose control.

The expression of a specific 2-deoxyglucose-6P phosphatase prevents catabolite repression mediated by 2-deoxyglucose in yeast.

2-deoxyglucose (2-DOG), a non-metabolize analogue of glucose, is taken up by yeast using the same transporter(s) as glucose and is phosphorylated by hexokinases producing 2-deoxyglucose-6-P. We found that in DOGR yeasts, 2-DOG was not able to trigger glucose repression, even at concentrations of 0.5%. This result suggests that the specific 2-DOG-6P phosphatase, the enzyme responsible for the DOGR phenotype, may be involved in inhibiting the process of catabolite repression mediated by 2-DOG.

Effect of prolactin on 2-deoxyglucose uptake in mouse mammary gland explants.

These studies were carried out to explore the possible effect of prolactin (PRL) on glucose uptake into culture mammary gland explants derived from 12- to 14-day pregnant mice. PRL was found to stimulate an increased rate of uptake of a nonmetabolized glucose analogue, 2-[3H]deoxyglucose, into cultured mammary tissues. The onset of this response was 16 h after the addition of PRL, and the response persisted for at least 24 h. A similar temporal response was observed when the PRL stimulation of [14C]glucose oxidation to 14CO2 was determined. The lowest PRL concentration that elicited a stimulation of 2-deoxyglucose uptake was 20 ng/ml, and a maximum response occurred with PRL at a concentration of 250 ng/ml. Ongoing protein synthesis appears to be essential for PRL to express its effect on 2-deoxyglucose transport since cyclohexamide, puromycin, and actinomycin D abolished the PRL response. It is also apparent that the PRL stimulation of 2-deoxyglucose involves activation of a specific carrier-mediated uptake transport system, since the rate of uptake of L-glucose into mouse mammary gland explants was unaffected by PRL.