Glutamate Family Research Articles

Comparative metabolomic analysis of the phloem sap of nine citrus relatives with different degrees of susceptibility to Huanglongbing disease

Citrus Huanglongbing (HLB) disease, also known as “citrus greening”, is currently considered the most devastating citrus disease due to its rapid spread, and high severity. Presently, research efforts are focused on searching for either curative treatments or resistant cultivars to combat the HLB-associated bacterium ‘Candidatus Liberibacter asiaticus’ (CLas).Metabolomics can help to unravel the mechanisms supporting the potential tolerance/resistance of citrus relatives. Herein, we carried out a metabolomic analysis to determine whether the level of resistance of nine citrus-related genotypes is influenced by their pre-existing metabolic background before infection. For this purpose, the healthy phloem of nine Citrinae genotypes previously categorized according to their different responses to HLB was analyzed. A total of 53 different metabolites were targeted, including amino acids, organic and inorganic acids, and sugars. Interestingly, we observed that resistant and partially resistant genotypes exhibited higher accumulations of organic acids such as quinic acid and citric acid. In contrast, the amount of total sugars showed a clear upward trend in the susceptible genotypes. Notably, within this last group of metabolites, sugar acids increase in both partially resistant and resistant accessions, being more evident in the resistant group.Alterations potentially linked to resistance levels were detected in specific amino acids belonging to the aspartate and glutamate families. Notably, only lysine levels exhibited a significant increase in the susceptible cultivars. The evaluation of five genes associated with lysine catabolism by RT-qPCR revealed differences in transcript abundance between resistant and susceptible samples suggesting a potential key role in plant defence. These findings open a new avenue to identify metabolites and/or substances that could aid in developing resistance strategies to this devastating disease.

Metabolomic Analysis Reveals the Effect of Insecticide Chlorpyrifos on Rice Plant Metabolism.

Pesticides as important agricultural inputs play a vital role in protecting crop plants from diseases and pests; however, the effect of pesticides on crop plant physiology and metabolism is still undefined. In this study, the effect of insecticide chlorpyrifos at three doses on rice plant physiology and metabolism was investigated. Our results revealed that chlorpyrifos cause oxidative stress in rice plants and even inhibit plant growth and the synthesis of protein and chlorophyll at high doses. The metabolomic results suggested that chlorpyrifos could affect the metabolic profiling of rice tissues and a total of 119 metabolites with significant changes were found, mainly including organic acids, amino acids, lipids, polyphenols, and flavonoids. Compared to the control, the content of glutamate family amino acids were significantly disturbed by chlorpyrifos, where defense-related proline and glutathione were significantly increased; however, glutamic acid, N-acetyl-glutamic acid and N-methyl-glutamic acid were significantly decreased. Many unsaturated fatty acids, such as linolenic acid and linoleic acid, and their derivatives lysophospholipids and phospholipids, were significantly accumulated in chlorpyrifos groups, which could act as osmolality substances to help rice cells relieve chlorpyrifos stress. Three organic acids, aminobenzoic acid, quinic acid, and phosphoenolpyruvic acid, involved in plant defenses, were significantly accumulated with the fold change ranging from 1.32 to 2.19. In addition, chlorpyrifos at middle- and high-doses caused the downregulation of most flavonoids. Our results not only revealed the effect of insecticide chlorpyrifos on rice metabolism, but also demonstrated the value of metabolomics in elucidating the mechanisms of plant responses to stresses.

Molecular Characterization and Expression Analysis of Putative Class C (Glutamate Family) G Protein-Coupled Receptors in Ascidian Styela clava.

Simple SummaryAscidians, known as the closest invertebrate relative to the vertebrate group, have a biphasic life cycle including the larval and sessile adult stages with strong adaptability to diverse environments. The nervous system of ascidians plays an essential role in its adaption to the external environment, but the molecular mechanisms underlying this process still need to be further clarified. The Class C G protein-coupled receptors (GPCRs) are a group of cell surface sensors for neurotransmitters and external chemicals, playing important functions in neurotransmission. We systematically characterized the putative Class C GPCRs in an important invasive ascidian species Styela clava and then analyzed their expression levels during different developmental stages and distribution in swimming larvae and multiple tissues of the adults. Our study suggests that S. clava Class C GPCRs potentially function as important molecules during neurotransmission related to physiological and morphogenetic changes in larvae and adults.In this study, we performed the genome-wide domain analysis and sequence alignment on the genome of Styela clava, and obtained a repertoire of 204 putative GPCRs, which exhibited a highly reduced gene number compared to vertebrates and cephalochordates. In this repertoire, six Class C GPCRs, including four metabotropic glutamate receptors (Sc-GRMs), one calcium-sensing receptor (Sc-CaSR), and one gamma-aminobutyric acid (GABA) type B receptor 2-like (Sc-GABABR2-like) were identified, with the absence of type 1 taste and vomeronasal receptors. All the Sc-GRMs and Sc-CaSR contained the typical “Venus flytrap” and cysteine-rich domains required for ligand binding and subsequent propagation of conformational changes. In swimming larvae, Sc-grm3 and Sc-casr were mainly expressed at the junction of the sensory vesicle and tail nerve cord while the transcripts of Sc-grm4, Sc-grm7a, and Sc-grm7b appeared at the anterior trunk, which suggested their important functions in neurotransmission. The high expression of these Class C receptors at tail-regression and metamorphic juvenile stages hinted at their potential involvement in regulating metamorphosis. In adults, the transcripts were highly expressed in several peripheral tissues, raising the possibility that S. clava Class C GPCRs might function as neurotransmission modulators peripherally after metamorphosis. Our study systematically characterized the ancestral chordate Class C GPCRs to provide insights into the origin and evolution of these receptors in chordates and their roles in regulating physiological and morphogenetic changes relevant to the development and environmental adaption.

Aspartate Transaminase AST2 Involved in Sporulation and Necrotrophic Pathogenesis in the Hemibiotrophs Magnaporthe oryzae and Colletotrichum graminicola.

Aspartate family includes five additional amino acids other than aspartate, among which most except aspartate have been reported for their action in pathogenesis by amino acid biosynthesis. However, how aspartate, the initial substrate of this family metabolic pathway, is involved in pathogenesis remains unknown. Here, we focused on aspartate transaminase (AST) that catalyzes transamination reaction between glutamate-aspartate in Magnaporthe oryzae. Three MoAST genes were bioinformatically analyzed, of which MoAST2 was uniquely upregulated when invasive hyphae switched to necrotrophic pathogenesis. MoAST2 deletion (ΔMoast2) caused a drastic reduction in conidiogenesis and appressorium formation. Particularly, ΔMoast2 was observed to be proliferated at the biotrophic phase but inhibited at the necrotrophic stage, and with invisible symptoms detected, suggesting a critical role in necrotrophic phase. Glutamate family restored the ΔMoast2 defects but aspartate family did not, inferring that transamination occurs from aspartate to glutamine. MoAST2 is cytosolic and possessed H2O2 stress tolerance. In parallel, Colletotrichum graminicola AST2, CgAST2 was proven to be a player in necrotrophic anthracnose development. Therefore, conserved AST2 is qualified to be a drug target for disease control.

Free amino acid regulation in fronds and roots of two Pteris cretica L. ferns under arsenic stress

In the present study, free amino acid (AA) regulation in the arsenic (As) hyperaccumulating ferns was evaluated in a pot experiment to determine the relationship between As stress and the characteristic change in metabolism of AAs. The ferns Pteris cretica cv. Albo-lineata (Pc-Al) and cv. Parkerii (Pc-Pa) were exposed to As treatments at 0, 20, 100, and 250 mg As/kg for 90 days. Greater As content, as well as higher biomass production, were identified in Pc-Al compared with Pc-Pa. Ferns showed changes in the stress metabolism of free AA homeostasis. These results indicate a disturbance in nitrogen metabolism and depletion of pool assimilated carbon metabolism. In the fronds and roots, Pc-Pa accumulated higher amounts of free AAs than Pc-Al. The total free AA content, as well as the ratio of the main AA family pathway (glutamate family), were increased by the accumulation of toxic As in the ferns. Results suggest that Pc-Al tolerates higher As doses better due to changes in AA biosynthesis; however, at higher As doses, Pc-Pa upregulated AA biosynthesis due to As toxicity. The most abundant free AAs of ferns was glutamine, which was enhanced by As. Furthermore, the ratios of selected individual free AAs revealed a characteristic phenotype difference between ferns.

Energy and nitrogenous waste from glutamate/glutamine catabolism facilitates acute osmotic adjustment in non-neuroectodermal branchial cells

Maintenance of homeostasis is one of the most important physiological responses for animals upon osmotic perturbations. Ionocytes of branchial epithelia are the major cell types responsible for active ion transport, which is mediated by energy-consuming ion pumps (e.g., Na+-K+-ATPase, NKA) and secondary active transporters. Consequently, in addition to osmolyte adjustments, sufficient and immediate energy replenishment is essenttableial for acclimation to osmotic changes. In this study, we propose that glutamate/glutamine catabolism and trans-epithelial transport of nitrogenous waste may aid euryhaline teleosts Japanese medaka (Oryzias latipes) during acclimation to osmotic changes. Glutamate family amino acid contents in gills were increased by hyperosmotic challenge along an acclimation period of 72 hours. This change in amino acids was accompanied by a stimulation of putative glutamate/glutamine transporters (Eaats, Sat) and synthesis enzymes (Gls, Glul) that participate in regulating glutamate/glutamine cycling in branchial epithelia during acclimation to hyperosmotic conditions. In situ hybridization of glutaminase and glutamine synthetase in combination with immunocytochemistry demonstrate a partial colocalization of olgls1a and olgls2 but not olglul with Na+/K+-ATPase-rich ionocytes. Also for the glutamate and glutamine transporters colocalization with ionocytes was found for oleaat1, oleaat3, and olslc38a4, but not oleaat2. Morpholino knock-down of Sat decreased Na+ flux from the larval epithelium, demonstrating the importance of glutamate/glutamine transport in osmotic regulation. In addition to its role as an energy substrate, glutamate deamination produces NH4+, which may contribute to osmolyte production; genes encoding components of the urea production cycle, including carbamoyl phosphate synthetase (CPS) and ornithine transcarbamylase (OTC), were upregulated under hyperosmotic challenges. Based on these findings the present work demonstrates that the glutamate/glutamine cycle and subsequent transepithelial transport of nitrogenous waste in branchial epithelia represents an essential component for the maintenance of ionic homeostasis under a hyperosmotic challenge.

Insights on Salt Tolerance of Two Endemic Limonium Species from Spain.

We have analysed the salt tolerance of two endemic halophytes of the genus Limonium, with high conservation value. In the present study, seed germination and growth parameters as well as different biomarkers—photosynthetic pigments, mono and divalent ion contents—associated to salt stress were evaluated in response to high levels of NaCl. The study was completed with an untargeted metabolomics analysis of the primary compounds including carbohydrates, phosphoric and organic acids, and amino acids, identified by using a gas chromatography and mass spectrometry platform. Limonium albuferae proved to be more salt-tolerant than L. doufourii, both at the germination stage and during vegetative growth. The degradation of photosynthetic pigments and the increase of Na+/K+ ratio under salt stress were more accentuated in the less tolerant second species. The metabolomics analysis unravelled several differences between the two species. The higher salt tolerance of L. albuferae may rely on its specific accumulation of fructose and glucose under high salinity conditions, the first considered as a major osmolyte in this genus. In addition, L. albuferae showed steady levels of citric and malic acids, whereas the glutamate family pathway was strongly activated under stress in both species, leading to the accumulation of proline (Pro) and γ-aminobutyric acid (GABA).

The Klebsiella pneumoniae citrate synthase gene, gltA, influences site specific fitness during infection.

Klebsiella pneumoniae (Kp), one of the most common causes of healthcare-associated infections, increases patient morbidity, mortality, and hospitalization costs. Kp must acquire nutrients from the host for successful infection; however, the host is able to prevent bacterial nutrient acquisition through multiple systems. This includes the innate immune protein lipocalin 2 (Lcn2), which prevents Kp iron acquisition. To identify novel Lcn2-dependent Kp factors that mediate evasion of nutritional immunity during lung infection, we undertook an InSeq study using a pool of >20,000 transposon mutants administered to Lcn2+/+ and Lcn2-/- mice. Comparing transposon mutant frequencies between mouse genotypes, we identified the Kp citrate synthase, GltA, as potentially interacting with Lcn2, and this novel finding was independently validated. Interestingly, in vitro studies suggest that this interaction is not direct. Given that GltA is involved in oxidative metabolism, we screened the ability of this mutant to use a variety of carbon and nitrogen sources. The results indicated that the gltA mutant has a distinct amino acid auxotrophy rendering it reliant upon glutamate family amino acids for growth. Deletion of Lcn2 from the host leads to increased amino acid levels in bronchioloalveolar lavage fluid, corresponding to increased fitness of the gltA mutant in vivo and ex vivo. Accordingly, addition of glutamate family amino acids to Lcn2+/+ bronchioloalveolar lavage fluid rescued growth of the gltA mutant. Using a variety of mouse models of infection, we show that GltA is an organ-specific fitness factor required for complete fitness in the spleen, liver, and gut, but dispensable in the bloodstream. Similar to bronchioloalveolar lavage fluid, addition of glutamate family amino acids to Lcn2+/+ organ lysates was sufficient to rescue the loss of gltA. Together, this study describes a critical role for GltA in Kp infection and provides unique insight into how metabolic flexibility impacts bacterial fitness during infection.

Combining metabolic flux analysis and adaptive evolution to enhance lipase production in Bacillus subtilis.

Metabolic fluxes during lipase production by Bacillus subtilis CICC 20034 in synthetic medium were studied using metabolic flux analysis (MFA). The MFA showed that lipase production was dependent on, and coupled to the tributyrin uptake rate, formation of biomass, lactate, ATP, as well as amino acids from the aspartate and glutamate family. Using tributyrin as the sole carbon source, an adaptive evolution strategy was applied to increase the tributyrin uptake rate. B. subtilis SPZ1 was obtained from CICC 20034 by adaptive evolution over 1000 generations of growth-based selection. The tributyrin consumption rate of strain SPZ1 reached 0.89g/(L·h) which was 1.9-fold higher than that of the original strain. The MFA indicated that the 212% increase of tributyrin uptake flux contributed to the 556% increase of lipase flux. Consequently, the lipase activity (0.65U/mL) of strain SPZ1 was 1.9-fold higher than that of the original strain. This was the highest lipase activity obtained by fermentation in synthetic medium reported for Bacillus strains. In complex culture medium, lipase activity of SPZ1 reached 3.3U/mL.

Metabolite Profiles of Raw Peanut Seeds Reveal Differences between Market‐Types

Peanuts (Arachis hypogaea L.) are prized for their flavor and popular worldwide as food or as food ingredients. The raw peanut seed contains the precursor compounds to roasted peanut flavor and has the potential to be manipulated through traditional breeding methods. However, little is known about the metabolome of the raw seeds. Comprehensive metabolite profiles of both raw runner and Virginia-type peanuts were determined. Using a system incorporating several methodologies including (RP)/UPLC-MS/MS and HILIC/UPLC-MS/MS, along with quantitation of fatty acids, free amino acids, and tocopherols, 365 metabolites were identified and of these, 52 were significantly different between market types (P<0.05). Higher levels of gamma-glutamylalanine, oxylipins, purine metabolites, and alpha-ketoglutarate derived members of the glutamate family of amino acids defined the Virginia-type, while runner-type peanuts were differentiated by their ethylmalonate and eicosenoate content. This study presents a comprehensive analysis of the raw peanut seed, providing knowledge of the range of small molecules present in peanuts. The new information presented here will enable future research for peanut quality improvement. PRACTICAL APPLICATION: Peanuts are widely used as snack foods and as food ingredients. Knowledge of the secondary metabolite compounds in raw peanuts is needed to determine their importance in peanut flavor and nutritional quality. This report used a nontargeted analytical approach for the identification of these types of compounds in peanuts for the first time. These data were supplemented with quantitative analysis of free amino acids and tocopherols and discussed as potential flavor precursors and health promoting compounds.

Contrasting amino acid profiles among permissive and non-permissive hosts of Candidatus Liberibacter asiaticus, putative causal agent of Huanglongbing.

Huanglongbing is a devastating disease of citrus. In this study, a comprehensive profile of phloem sap amino acids (AA) in four permissive host plants of Candidatus Liberibacter asiaticus (CLas) and three non-permissive Rutaceae plants was conducted to gain a better understanding of host factors that may promote or suppress the bacterium. The AA profiles of Diaphorina citri nymphs and adults were similarly analyzed. A total of 38 unique AAs were detected in phloem sap of the various plants and D. citri samples, with phloem sap of young shoots containing more AAs and at higher concentrations than their mature counterparts. All AAs detected in phloem sap of non-permissive plants were also present in CLas -permissive hosts plus additional AAs in the latter class of plants. However, the relative composition of 18 commonly shared AAs varied between CLas -permissive hosts and non-permissive plants. Multivariate analysis with a partial least square discriminant methodology revealed a total of 12 AAs as major factors affecting CLas host status, of which seven were positively related to CLas tolerance/resistance and five positively associated with CLas susceptibility. Most of the AAs positively associated with CLas susceptibility were predominantly of the glutamate family, notably stressed-induced AAs such as arginine, GABA and proline. In contrast, AAs positively correlated with CLas tolerance/resistance were mainly of the serine family. Further analysis revealed that whereas the relative proportions of AAs positively associated with CLas susceptibility did not vary with host developmental stages, those associated with CLas tolerance/resistance increased with flush shoot maturity. Significantly, the proline-to-glycine ratio was determined to be an important discriminating factor for CLas permissivity with higher values characteristic of CLas -permissive hosts. This ratio could be exploited as a biomarker in HLB-resistance breeding programs.

Effects of salinity on the cellular physiological responses of Natrinema sp. J7-2.

The halophilic archaea (haloarchaea) live in hyersaline environments such as salt lakes, salt ponds and marine salterns. To cope with the salt stress conditions, haloarchaea have developed two fundamentally different strategies: the "salt-in" strategy and the "compatible-solute" strategy. Although investigation of the molecular mechanisms underlying the tolerance to high salt concentrations has made outstanding achievements, experimental study from the aspect of transcription is rare. In the present study, we monitored cellular physiology of Natrinema sp. J7-2 cells incubated in different salinity media (15%, 25% and 30% NaCl) from several aspects, such as cellular morphology, growth, global transcriptome and the content of intracellular free amino acids. The results showed that the cells were polymorphic and fragile at a low salt concentration (15% NaCl) but had a long, slender rod shape at high salt concentrations (25% and 30% NaCl). The cells grew best in 25% NaCl, mediocre in 30% NaCl and struggled in 15% NaCl. An RNA-seq analysis revealed differentially expressed genes (DEGs) in various salinity media. A total of 1,148 genes were differentially expressed, consisting of 719 DEGs (348 up-regulated and 371 down-regulated genes) between cells in 15% vs 25% NaCl, and 733 DEGs (521 up-regulated and 212 down-regulated genes) between cells in 25% vs 30% NaCl. Moreover, 304 genes were commonly differentially expressed in both 15% vs 25% and 25% vs30% NaCl. The DEGs were enriched in different KEGG metabolic pathways, such as amino acids, glycerolipid, ribosome, nitrogen, protoporphyrin, porphyrin and porhiniods. The intracellular predominant free amino acids consisted of the glutamate family (Glu, Arg and Pro), aspartate family (Asp) and aromatic amino acids (Phe and Trp), especially Glu and Asp.

Functional amino acids stimulate muscle development and improve fillet texture of Atlantic salmon

Sufficient firmness is essential for consumer appreciation and the suitability for processing of fish fillets. The objective of this study was to investigate the effect of functional amino acids (AA) on fillet texture and muscle development of Atlantic salmon. Triplicate net pens of 105 g salmon were fed a standard diet, or the same diet with added 15 g/kg arginine or 15 g/kg glutamate during a 5-month rearing period. The growth rate and FCR (0.91–0.92) showed no significant dietary effects (body weight 864–887 g). Glutamate supplementation resulted in delayed postmortem glycogen degradation (pH drop) and rigour development, along with improved fillet firmness and intercellular myofibre integrity. An in vitro study with salmon myosatellite cells showed that exogenous glutamine or arginine increased the expression of muscle growth markers (myog, tnnl2, myl) at both 8 and 16°C culture temperature. The expression of a marker for proteolysis (ctsb), myl and myog were highest for the glutamine treatment at 16°C. Significant interaction between exogenous AA and temperature indicated elevated AA requirement when growth is accelerated. It is concluded that AA from the glutamate family are vital for fillet firmness. The dispensable glutamine and glutamate appear more critical compared to arginine, particularly during high-performance periods.

Redox regulation of free amino acid levels in Arabidopsis thaliana.

Abiotic stresses induce oxidative stress, which modifies the level of several metabolites including amino acids. The redox control of free amino acid profile was monitored in wild-type and ascorbate or glutathione deficient mutant Arabidopsis thaliana plants before and after hydroponic treatment with various redox agents. Both mutations and treatments modified the size and redox state of the ascorbate (AsA) and/or glutathione (GSH) pools. The total free amino acid content was increased by AsA, GSH and H2 O2 in all three genotypes and a very large (threefold) increase was observed in the GSH-deficient pad2-1 mutant after GSH treatment compared with the untreated wild-type plants. Addition of GSH reduced the ratio of amino acids belonging to the glutamate family on a large scale and increased the relative amount of non-proteinogenic amino acids. The latter change was because of the large increase in the content of alpha-aminoadipate, an inhibitor of glutamatic acid (Glu) transport. Most of the treatments increased the proline (Pro) content, which effect was due to the activation of genes involved in Pro synthesis. Although all studied redox compounds influenced the amount of free amino acids and a mostly positive, very close (r > 0.9) correlation exists between these parameters, a special regulatory role of GSH could be presumed due to its more powerful effect. This may originate from the thiol/disulphide conversion or (de)glutathionylation of enzymes participating in the amino acid metabolism.

Editorial: Amino Acids of the Glutamate Family: Functions beyond Primary Metabolism.

EDITORIAL article Front. Plant Sci., 21 March 2016Sec. Plant Physiology Volume 7 - 2016 | https://doi.org/10.3389/fpls.2016.00318

Anaerobic growth of Corynebacterium glutamicum via mixed-acid fermentation.

Corynebacterium glutamicum, a model organism in microbial biotechnology, is known to metabolize glucose under oxygen-deprived conditions to l-lactate, succinate, and acetate without significant growth. This property is exploited for efficient production of lactate and succinate. Our detailed analysis revealed that marginal growth takes place under anaerobic conditions with glucose, fructose, sucrose, or ribose as a carbon and energy source but not with gluconate, pyruvate, lactate, propionate, or acetate. Supplementation of glucose minimal medium with tryptone strongly enhanced growth up to a final optical density at 600 nm (OD600) of 12, whereas tryptone alone did not allow growth. Amino acids with a high ATP demand for biosynthesis and amino acids of the glutamate family were particularly important for growth stimulation, indicating ATP limitation and a restricted carbon flux into the oxidative tricarboxylic acid cycle toward 2-oxoglutarate. Anaerobic cultivation in a bioreactor with constant nitrogen flushing disclosed that CO2 is required to achieve maximal growth and that the pH tolerance is reduced compared to that under aerobic conditions, reflecting a decreased capability for pH homeostasis. Continued growth under anaerobic conditions indicated the absence of an oxygen-requiring reaction that is essential for biomass formation. The results provide an improved understanding of the physiology of C. glutamicum under anaerobic conditions.

Functional characterization and expression analysis of rice δ(1)-pyrroline-5-carboxylate dehydrogenase provide new insight into the regulation of proline and arginine catabolism.

While intracellular proline accumulation in response to various stress conditions has been investigated in great detail, the biochemistry and physiological relevance of proline degradation in plants is much less understood. Moreover, the second and last step in proline catabolism, the oxidation of δ1-pyrroline-5-carboxylic acid (P5C) to glutamate, is shared with arginine catabolism. Little information is available to date concerning the regulatory mechanisms coordinating these two pathways. Expression of the gene coding for P5C dehydrogenase was analyzed in rice by real-time PCR either following the exogenous supply of amino acids of the glutamate family, or under hyperosmotic stress conditions. The rice enzyme was heterologously expressed in E. coli, and the affinity-purified protein was thoroughly characterized with respect to structural and functional properties. A tetrameric oligomerization state was observed in size exclusion chromatography, which suggests a structure of the plant enzyme different from that shown for the bacterial P5C dehydrogenases structurally characterized to date. Kinetic analysis accounted for a preferential use of NAD+ as the electron acceptor. Cations were found to modulate enzyme activity, whereas anion effects were negligible. Several metal ions were inhibitory in the micromolar range. Interestingly, arginine also inhibited the enzyme at higher concentrations, with a mechanism of uncompetitive type with respect to P5C. This implies that millimolar levels of arginine would increase the affinity of P5C dehydrogenase toward its specific substrate. Results are discussed in view of the involvement of the enzyme in either proline or arginine catabolism.

Evidence for synergistic control of glutamate biosynthesis by glutamate dehydrogenases and glutamate in Bacillus subtilis.

In the Gram-positive bacterium, Bacillus subtilis glutamate is synthesized by the glutamine synthetase and the glutamate synthase (GOGAT). During growth with carbon sources that exert carbon catabolite repression, the rocG glutamate dehydrogenase (GDH) gene is repressed and the transcription factor GltC activates the expression of the GOGAT encoding gltAB genes. In the presence of amino acids of the glutamate family, the GDH RocG is synthesized and the enzyme prevents GltC from binding to DNA. The dual control of glutamate biosynthesis allows the efficient utilization of the available nutrients. Here we provide genetic and biochemical evidence that, like RocG, also the paralogous GDH GudB can inhibit the transcription factor GltC, thereby controlling glutamate biosynthesis. Contradictory previous observations show that high level of GDH activity does not result in permanent inhibition of GltC. By controlling the intracellular levels of glutamate through feeding with exogenous arginine, we observed that the GDH-dependent control of GltC and thus expression of the gltAB genes inversely correlates with the glutamate pool. These results suggest that the B. subtilis GDHs RocG and GudB in fact act as glutamate sensors. In conclusion, the GDH-mediated control of glutamate biosynthesis seems to depend on the intracellular glutamate concentration.

Expression and functional role of orphan receptor GPR158 in prostate cancer growth and progression.

Prostate cancer (PCa) is the second-leading cause of cancer-related mortality, after lung cancer, in men from developed countries. In its early stages, primary tumor growth is dependent on androgens, thus generally can be controlled by androgen deprivation therapy (ADT). Eventually however, the disease progresses to castration-resistant prostate cancer (CRPC), a lethal form in need of more effective treatments. G-protein coupled receptors (GPCRs) comprise a large clan of cell surface proteins that have been implicated as therapeutic targets in PCa growth and progression. The findings reported here provide intriguing evidence of a role for the newly characterized glutamate family member GPR158 in PCa growth and progression. We found that GPR158 promotes PCa cell proliferation independent of androgen receptor (AR) functionality and that this requires its localization in the nucleus of the cell. This suggests that GPR158 acts by mechanisms different from other GPCRs. GPR158 expression is stimulated by androgens and GPR158 stimulates AR expression, implying a potential to sensitize tumors to low androgen conditions during ADT via a positive feedback loop. Further, we found GPR158 expression correlates with a neuroendocrine (NE) differentiation phenotype and promotes anchorage-independent colony formation implying a role for GPR158 in therapeutic progression and tumor formation. GPR158 expression was increased at the invading front of prostate tumors that formed in the genetically defined conditional Pten knockout mouse model, and co-localized with elevated AR expression in the cell nucleus. Kaplan-Meier analysis on a dataset from the Memorial Sloan Kettering cancer genome portal showed that increased GPR158 expression in tumors is associated with lower disease-free survival. Our findings strongly suggest that pharmaceuticals targeting GPR158 activities could represent a novel and innovative approach to the prevention and management of CRPC.

Latrepirdina: revisión sistemática de los estudios preclínicos

We conduct a systematic review of the preclinical studies published to date involving the use of latrepirdine (Dimebon ®). Latrepirdine is capable of modulating different targets, such as those related with mitochondria, acetylcholinesterase activity or intraneuronal calcium levels, perhaps thanks to its action upon the N-methyl-D-aspartate-type receptor, which belongs to the glutamate family. The findings published on the possible effect of latrepirdine in protein aggregation processes in disorders such as Alzheimer's disease and Parkinson's disease are quite controversial. Likewise, the possible neuroprotective effect of latrepirdine has been evaluated in animal models of neurodegenerative diseases, again with heterogeneous results. Consequently, it can be concluded that no preclinical scientific evidence has been found to justify carrying out clinical trials.