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Metabolism and functions of gamma-aminobutyric acid
... Mitochondrial GABA transaminase (GABA-T) enzymes, glutamate decarboxylase (GAD), and succinic semialdehyde dehydrogenase (SSADH) are all involved in the GABA shunt 16 . Many studies suggests that GABA may play a role in the regulation of cytosolic pH, the modi cation of nitrogen and carbon metabolism, and the provision of protection against oxidative stress 17,18 . Under oxidative stress and other abiotic conditions, it was expected that GABA breakdown would decrease the amount of reactive oxygen that accumulated 17 . ...
... Many studies suggests that GABA may play a role in the regulation of cytosolic pH, the modi cation of nitrogen and carbon metabolism, and the provision of protection against oxidative stress 17,18 . Under oxidative stress and other abiotic conditions, it was expected that GABA breakdown would decrease the amount of reactive oxygen that accumulated 17 . Plants that are subjected to biotic and abiotic stresses are rapidly accumulate GABA 17,19−21 . ...
... Under salt and osmotic stress, the transcription level of GAD signi cantly increased (P ≤ 0.05) in all treated barley genotypes tested in this investigation (Acsad 176, Athroh, and Rum) (Fig. 3). Decarboxylation of glutamate to CO2 and GABA is improved by the GAD 17 . Rum genotypes subjected to osmotic stress (mannitol) had signi cant levels of GAD expression (Fig. 4b), which was consistent with the high amount of GABA accumulation (mannitol). ...
GABA is a secondary metabolite that protects against oxidative damage, regulates nitrogen metabolism, and controls cytosolic pH in response to abiotic and biotic stress. This study examines the role of GABA shunt pathway response to salt and osmotic stress in three barley genotypes ("Acsad 176", "Athroh", and "Rum") interm of oxidative damage, seed germination and seedling growth as measured by glutamate decarboxylase gene (GAD) expression malondialdehyde accumulation (MDA), total proteins, total carbohydrates level, and GABA accumulation level. As Mannitol, NaCl, and sorbitol concentrations increased, seed germination decreased for all genotypes. All genotypes seedling MDA concentrations increased with increasing NaCl, mannitol, and sorbitol concentrations. Acsad 176 showed high GABA accumulation under NaCl treatment. Rum's GABA accumulation under mannitol testament increased significantly. All salt and osmotic treatments decreased chlorophyll a and b and carbohydrate content. At the same time it increased GAD transcription in all barley genotypes. Salt and osmotic stresses affected protein content in all genotypes. Acsad 176 may adapt to NaCl stress by accumulating carbohydrates more than Athroh and Rum. GABA shunt is a crucial signaling and metabolic pathway that facilitates barley's adaptation to salt and osmotic stress. In high salt and osmotic soil, Acsad 176 is the recommended genotype.
... The γ-aminobutyric acid (4-aminobutyric acid, GABA), is a non-protein amino acid, which possesses an amino group at γ-carbon rather than α-carbon, therefore does not integrate into proteins. It contributes significantly to the free amino acid pool and exists in unbound form (Shelp et al. 1999). It exhibits zwitterionic features at physiological pH, high solubility in water and can attain many conformations in solution, including cyclic structure like proline (Christensen et al. 1994). ...
... Though the contribution of GABA as a signalling molecule was considered quite early in animals, in plants, until recently, it was generally considered as a carbonnitrogen metabolite (Shelp et al. 1999;Bouche et al. 2003;Bouche and Fromm 2004). In plants, the recent outlook suggested it as an endogenous signalling molecule for plant growth regulation and plant development (Carillo 2018). ...
... However, there are certain observations that backed the signalling feature of GABA as highlighted in a review (Ramesh et al. 2015), which include features like rapid increase in response to biotic and abiotic stress, gradient of GABA concentration across plant tissues, inter-and intracellular compartmentation of GABA metabolism, discovery of GABA-binding site in plant cell membrane, and GABA-regulated ion channels in plants. Under several abiotic (hypoxia, heat, cold, drought, mechanical wounding) or biotic stress (wounding due to herbivory and infection), the quick accumulation of GABA was noticed (Shelp et al. 1999;Shelp et al. 2012). The accumulation promotes plant growth and can alleviate stress by up-regulating the anti-oxidant defence systems. ...
The phenomenon of climate change has appeared in recent time as an unstoppable and unequivocal event due to the change of environmental condition, rising of sea level, ozone layer fluctuation, and most importantly, it has raised the global apprehension in lowering global food production. Abiotic stress is one of the major factors influencing overall performance of crop plants. The response of crop plant due to environmental alteration could be effective due to the accumulation of compatible solutes or osmoprotetctants, which contain amino acids, polyamines, sugar, and quaternary ammonium compounds. Compatible solutes play important role in mitigating the adverse effect of abiotic stresses in plants. Compatible solutes are low molecular weight compounds stabilizing cellular structure and scavenging reactive oxygen species (ROS) during abiotic stress conditions. The role of compatible solutes is very important in plant growth and development, i.e., glycinebetaine (GB) is helpful in maintaining osmotic potential, protecting proteins from denaturation and restoration of photosystem II complex (PSII) in plants. The biosynthesis of compatible solutes is important in plants especially adapting under stressful environment, in stabilizing the structure of PSII by protecting extrinsic proteins, and the introgression of genes associated with osmoprotactants from one plant to another by genetic engineering.
... γ-Aminobutyric acid (GABA) has shown a considerable role in regulating stress tolerance in many harvested fruit and vegetable products, which is tightly synchronized with polyamine metabolism (Shelp et al. 1999). Previous studies have shown that GABA treatments alleviated CI in cucumber fruit by increased contents of proline and GABA, as well as elevated antioxidant activities (Malekzadeh et al. 2017). ...
... GABA is a non-protein amino acid, which is a four carbon compound found in most living organisms (Shelp et al. 1999). GABA biosynthesis is stimulated with a coordinated manner of enzymes via the GABA shunt pathway (Deewatthanawong et al. 2010b;Hyun et al. 2013). ...
... GABA plays a significant role in CI in most vegetables and fruits in post-harvest conditions (Palma et al. 2014). It was indicated that GABA is likely to accumulate in the response defense system to biotic and abiotic stresses in plants (Shelp et al. 1999). The increased GABA shunt and GABA-T activity, provide carbon and nitrogen of NADH as a reduction molecule and of ATP as the energy currency, but also serves as a H 2 O 2 scavenger (Aghdam et al. 2016). ...
The mechanism of melatonin (MT) induced chilling tolerance in harvested cucumber fruit was investigated at commercial maturity. In this study, cucumber fruits were treated with 100 μmol L-1 MT at 4°C and 90% relative humidity for 15 d of storage. In comparison with the control, cucumber treatment with MT resulted in reduced chilling injury (CI), decreased electrolyte leakage and enhanced firmness. The fruits treated with MT showed higher chlorophyll contents in storage conditions with suppressed chlorophyllase enzyme activity. MT treatment increased arginine decarboxylase (ADC) and ornithine decarboxylase (ODC) enzyme activities. Moreover, enhanced expression of the Cucumis sativus ADC (CsADC) and C. sativus ODC (CsODC) genes resulted in the accumulation of polyamine contents. Similarly, proline levels exhibited higher levels among treated fruits. Meanwhile, the proline synthesizing enzymes △ 1-pyrroline-5-carboxylate syntheses (P5CS) and ornithine aminotransferase (OAT) were significantly increased, while a catabolic enzyme of proline dehydrogenase (PDH) activity was inhibited by treatment. In addition, MT induced expression of C. sativus OAT (CsOAT) and C. sativus P5CS (CsP5CS) genes. Cucumber fruits treated with MT also exhibited higher γ-aminobutyric acid (GABA) content by enhanced GABA transaminase (GABA-T) and glutamate decarboxylase (GAD) enzyme activities and a higher C. sativus GAD (CsGAD) gene expression. To sum up, the results show that MT treatment enhanced chilling tolerance, which was associated with the regulation of polyamines, as well as proline and γ-aminobutyric acid.
... Le GABA interviendrait dans la régulation du pH du cytosol, car il peut s'accumuler dans les tissus en réponse à une acidification de ce dernier. Le GABA joue également un rôle dans la protection contre le stress oxydatif et dans la défense contre les insectes (Shelp et al. 1999;Bouché and Fromm 2004). ...
... Arabidopsis, la glutamate décarboxylase (GAD, EC 4.1.1.15) convertit le glutamate en GABA (Shelp et al. 1999;Bouché and Fromm 2004). Au contraire de KO8-2 badh, les mutants gad sont très fortement affectés dans la capacité à accumuler le GABA en réponse au stress (Bouché and Fromm 2004). ...
... D'après les analyses réalisées dans le cadre de cette thèse, les AtALDH10 pourraient être impliquées dans la résistance au stress salin. Leur capacité à oxyder l'ABAL in vitro suggère qu'elles interviennent dans la biosynthèse de GABA, qui est un osmoprotectant (Shelp et al. 1999 (Galston et al. 1997). La moindre capacité du mutant KO8-2 badh à détoxifier l'ABAL et l'APAL pourrait donc également expliquer sa sensibilité accrue au stress salin. ...
La carnitine est un acide aminé non protéinogène présent dans l’ensemble du vivant. Son rôle dans le transport intracellulaire des acides gras est bien caractérisé chez les animaux. Chez les plantes aussi, la carnitine intervient dans le métabolisme lipidique, mais son rôle précis demeure peu documenté. Chez les animaux et les levures, la carnitine est synthétisée à partir de la triméthyllysine (TML) via 4 réactions enzymatiques. L’identification chez les plantes de la TML et de la γ-butyrobétaïne, le précurseur direct de la carnitine, suggère que la voie de biosynthèse de la carnitine chez les plantes est similaire à celle des animaux. Une recherche d’homologues a permis d’identifier les deux aldéhyde déshydrogénases 10 (ALDH10) d’Arabidopsis comme potentiellement impliquées dans la synthèse de γ-butyrobétaïne. Dans le cadre de ce travail de thèse, l’implication des AtALDH10 dans la biosynthèse de carnitine et dans la physiologie de la plante a été étudiée. Dans un premier temps, les AtALDH10 recombinantes ont été caractérisées in vitro. Cette étude indique qu’elles peuvent synthétiser la γ-butyrobétaïne, ainsi que des osmoprotectants. Ensuite, un mutant perte de fonction pour les deux AtALDH10 a été généré et étudié. Il présente une sensibilité accrue au stress salin, une hampe florale plus haute, un nombre plus élevé de hampes florales secondaires, et des siliques de taille réduite contenant moins de graines viables. Les graines parvenant à maturité sont enrichies en protéines et en lipides, et le développement de la plante est plus rapide. Ces résultats suggèrent une intervention des AtALDH10 dans la réponse au stress hydrique, l’architecture de la plante, le développement précoce. La teneur en γ-butyrobétaïne dans les tissus est significativement réduite chez le mutant, confirmant le rôle des AtALDH10 dans la synthèse de γ-butyrobétaïne in planta. La teneur résiduelle en γ-butyrobétaïne semble suffisante pour maintenir une teneur en carnitine inchangée. Cela pose la question de la régulation de la voie de biosynthèse de la carnitine chez les plantes, et suggère un lien entre la γ-butyrobétaïne et la réponse au stress, les voies de signalisation et l’architecture de la plante. Outre ce travail, une recherche bioinformatique a permis d’identifier une protéine d’Arabidopsis dont la structure est proche de celle de l’enzyme humaine responsable de la conversion de la γ-butyrobétaïne en carnitine. L’étude de cette protéine n’a pas confirmé son implication dans la synthèse de carnitine. L’identification et la caractérisation de mutants affectés dans la synthèse de carnitine ou son intervention dans le transport des acides gras serait précieuse pour une meilleure compréhension du rôle de la carnitine chez les plantes.
... Phenylalanine (Phe) is the substrate of phenylalanine ammonia-lyase (PAL), which is the entry step of the penylpropanoid metabolism, which feeds the polyphenol biosynthesis pathway (Kataoka et al., 1983). γaminobutyric acid (GABA) is a non-proteinaceous, four carbon amino acid that is reported to be involved in multiple processes in plant, including the carbon flux into the TCA cycle, pH regulation, plant development and insect defence signalling (reviewed by Shelp et al., 1999, Hildebrandt et al., 2015. Besides, GABA might play a role in the regulation of malate metabolism via the connection with the TCA cycle (Han et al., 2018b). ...
... Amino acids profiles were influenced by the modulation of L/F, especially for some free amino acids associated with Glycolysis and TCA cycle process. GABA is a four-carbon amino acid that is reported to be involved in maintaining C/N balance by regulating carbon flux into the TCA cycle (reviewed by Shelp et al., 1999, Hildebrandt et al., 2015, and it also can be catabolised to Glu and subsequently Asp to maintain osmotic homeostasis during fruit ripening (Michaeli and Fromm, 2015). The changes in these intermediates linking primary and secondary metabolites are likely to be in response to changes in sugar concentration in different L/Fs. ...
... Among the 21 free amino acids that we detected; GABA (-aminobutyric acid) increased significantly in DGH containing medium, under different glucose concentrations (Fig. 2B). GABA is a nonproteinaceous, four-carbon amino acid that is reported to be involved in multiple processes in plant, including the carbon flux into the TCA cycle, pH regulation, plant development and insect-defence, signalling (reviewed by Shelp et al., 1999, Hildebrandt et al., 2015. The increase in GABA may be due to the effect of DGH on the activity of glucose-related ...
Ongoing climate change causes extreme weather events to increase both in frequency and intensity, significantly affecting vine physiology and grape berry composition at harvest. Elevated temperatures during the growing season increase must sugar content, while reducing organic acids and anthocyanins levels, modifying wine quality and typicality. Viticultural practices such as leaf-to-fruit ratio manipulations combined with applications of plant hormones can potentially be used to adapt to climate change and maintain the sustainability of wine production. To optimize adaptation strategies, we need to gain better insights into the links between sugars and anthocyanins accumulation during berry development and ripening.Fruiting-cuttings made up of one vertical shoot with one grape cluster of cv. Cabernet Sauvignon were grown between 2013 and 2018 in a naturally lighted and semi-controlled greenhouse. Various leaf-to-fruit ratios (2 to12 leaves per cluster) were applied, and combined with spraying abscisic acid (ABA, 400 mg.L-1) on berries at the pre-veraison stage. To explore the biological mechanisms underlying the effect of leaf-to-fruit ratio manipulation and application of exogenous ABA on berry composition, transcript abundance of genes related to sugar, anthocyanins and ABA metabolism were studied in this material. To further analyze the potential interplay between sugar and ABA signaling on anthocyanins biosynthesis, in vitro berry culture experiments were also conducted. Berries were cultured 2-3 weeks on solid medium supplemented with different sugar levels (0%, 2%, 8%) combined with ABA (200 µM), ABA biosynthesis inhibitor (NDGA), or hexokinase inhibitors (NAG, GDH).The results confirmed that reducing the leaf-to-fruit ratio had a significant effect on berry composition: reduction of sugar and anthocyanin content, slight increase in total organic acids and modification of the free amino acids composition. Exogenous ABA application increased sugars and anthocyanins concentrations, and partially restored the coupling between sugar and anthocyanins accumulation under low leaf-to-fruit ratios, without affecting the amino acids/sugar/organic acids ratios. Transcript abundance analysis revealed that several anthocyanin biosynthetic genes (CHS2, CHS3, CHI, F3H, DFR, LODX, UFGT, MybA1 and MybA2) were decreased under low leaf-to-fruit ratio, whereas some genes (CHI, F3H, F3’5’H, LODX, UFGT, MybA1 and MybA2) were up-regulated after exogenous ABA treatment. Exogenous ABA had little effect on the transcript abundance of sugar accumulation related genes, and leaf-to-fruit ratio also had little effect on ABA biosynthetic genes. Berry in vitro culture experiments showed that ABA and elevated concentrations of glucose synergistically induced anthocyanins biosynthesis during ripening. Both ABA and glucose decreased Phenylalanine concentration in ripening berries, most probably due to their role in promoting anthocyanins synthesis, and two-way crossed ANOVA analysis indicate a significant interaction between sugar and ABA levels and anthocyanin accumulation in berries.In conclusion, our results showed that ABA and sugar signaling synergistically interact to regulate the expression of anthocyanin biosynthetic genes and increase anthocyanins accumulation during berry ripening. Thus, exogenous ABA application was able to increase the ratio of anthocyanins to sugar under low leaf-to-fruit ratio at harvest, and combining leaf-to-fruit ratio manipulation with exogenous ABA applications may offer a fine-tuned way to reduce sugar concentration, while maintaining anthocyanin concentrations in grape berry, potentially offering a way to partially alleviate climate change related high temperature effects.
... GABA is a non-protein amino acid widely distributed in nature that is a neurotransmitter in the brain and spinal cord of mammals (Bown et al., 1999). GABA health benefits include: diuretic effects, tranquillising effects (Jakobs et al., 1993), Alzheimer's disease prevention (Ito & Ishikawa, 2004), regulation of blood pressure and heart rate, pain and anxiety relief (Kono & Himeno, 2000), improved insulin secretion to prevent diabetes (Huang et al., 2007), or inhibition of cancer cell proliferation (Oh & Oh, 2004). ...
... GABA health benefits include: diuretic effects, tranquillising effects (Jakobs et al., 1993), Alzheimer's disease prevention (Ito & Ishikawa, 2004), regulation of blood pressure and heart rate, pain and anxiety relief (Kono & Himeno, 2000), improved insulin secretion to prevent diabetes (Huang et al., 2007), or inhibition of cancer cell proliferation (Oh & Oh, 2004). GABA is produced primarily by the decarboxylation of L-glutamic acid, catalysed by the glutamate decarboxylase enzyme (GAD) during the germination process of brown rice (Bown et al., 1999). The amount of GABA in germinated brown rice in Japan was noticed to be ten times more as compared to milled white rice and two times more than that of brown rice (Kayahara & Tsukahara, 2000). ...
Introduction: Many people are currently interested in improving and maintaining their health status by changing their dietary habits, like eating more natural foods; thus sprout products are becoming increasingly popular. In this context, sprouted brown rice grains are an excellent example of functional food, because besides their nutritive value, they also lower the risk of various diseases and/or exert healthpromoting effects. In this paper, we focused on the bioactive compound γ-aminobutyric acid (GABA) in germinated brown rice. GABA is known as an important amino acid that can help reduce hypertension and inhibit cancer cells development. Methods: We investigated the hydration characteristics of brown rice by drying them in a moisture analyser at 130°C until reaching a constant weight. The effects of soaking (duration and pH of soaking solution), as well as incubation conditions (temperature and time) on GABA biosynthesis in MangBuk brown rice of Vietnam were measured. Quantification of GABA was measured using a spectrophotometer. Results: GABA content in MangBuk type 1 brown rice was higher than in type 2. GABA content reached its highest value at 691.88 µg/g for type 1 rice and 596.48 µg/g for type 2 rice when MangBuk brown rice was soaked in a pH 7 water at 30°C for 12 hours, and then incubated at 35°C for 30 hours in aerobic condition. Conclusion: Germination conditions modified the content of biologically active compounds in MangBuk soft and hard rice varieties. GABA was synthesised during germination based on three factors, namely time of incubation, temperature of incubation, and pH of solution.
... Increased concentrations of GABA under drought stress has shown to induce stomatal closure via activation of anion channel, aluminium-activated malate transporters (ALMTs) [33]. Accumulation of GABA under stress conditions also helps activating plant's innate defence potential and pre-conditioning of the plant to next drought event through synthesising osmolytes [34], enhancing photochemical efficiency and WUE and ROS detoxification [35][36][37]. ...
... Before imposition of the drought stress, similar GABA levels were detected in all three clones. In line with previous studies [28,29,34,35], 2-fold and 12fold increase in GABA concentration was observed in DS and DT clones respectively at D1. However, in commercial clones, leaf GABA levels did not change in response to soil moisture content. ...
The Australian wine industry is currently under pressure to sustain its profitability due to climate change. Therefore, there is a pressing need to explore grapevine genetic diversity and identify superior clones with improved drought resistance. We previously characterised more than 15,000 dry-farmed (for over 65 years) Cabernet Sauvignon clones in a vineyard and identified three drought-tolerant (DT) clones, which can maintain significantly higher intrinsic water use efficiency (WUEi) under limited soil moisture than drought-sensitive (DS) clones. To understand whether DT clones grown under multi-decadal cyclical drought can prime their vegetatively-propagated clonal progenies for future drought events, in this study, all DT and DS vegetative progenies were propagated with commercial clones in the glasshouse. Their physiological and molecular responses were investigated under well-watered and two recurrent drought (D1 and D2) conditions. We observed that concentration of a natural priming agent, γ-amino butyric acid (GABA), were significantly higher in all DT progenies relative to other progenies under drought. Both commercial and DT progenies exhibited improved gas exchange, photosynthetic performance and WUEi under recurrent drought events relative to DS progenies. Our results suggest that DT progenies have adapted to be in a primed state to withstand future drought events.
... In plants, GABA is synthesized from glutamate and Δ 1 -pyrroline. The synthesis of GABA from glutamate via Glutamate Decarboxylase (GAD) is considered the pathway contributing the most to GABA production in plants (Shelp et al. 1999). This was demonstrated to be the case at least in roots, where the mutation of the rootspecific GAD1 resulted in a dramatical decrease of GABA levels (Bouché et al. 2004). ...
... Both GABA and proline have been reported to accumulate in many plant species under diverse abiotic stress conditions (Table 12.1). GABA was initially considered to accumulate in response to diverse stresses such as hypoxia, wounding, drought, heat or cold stress (Shelp et al. 1999), but in many other cases reductions in GABA have been reported as well (Table 12.1), which usually is interpreted as a mechanism to provide the necessary energy to cope with the stress condition through the GABA shunt. Other authors have reported that the regulation of the GABA shunt is the key response, independently of the GABA levels. ...
Environmental and biotic stresses induce metabolic changes in plants which led to the accumulation of specific metabolites. For instance, the accumulation of gamma-aminobutyric acid (GABA) and proline is a conserved response of plants to a wide range of stresses. In particular, these amino acids accumulate the most under abiotic stress conditions and, to a lesser extent, under biotic stress conditions. Multiple shared roles have been assigned to these molecules, including osmoprotection, osmotic adjustment, carbon source, redox balance and antioxidant functions. For some of these roles, however, the physiological function of these molecules has been a matter of debate. Recent studies point to unexplored functions of these molecules. For instance, proline accumulation was suggested to contribute to sustaining photosynthesis under stress conditions and proline catabolism was suggested to be relevant to induce plant autophagy. Moreover, many enzymes of proline metabolism were suggested to be important for optimal response to biotic stress. This is even clearer for GABA, for which several mechanisms of action were already described to explain its capacity to protect the plant against pathogens. However, the role of GABA under biotic stress is highly dependent on the biotic stressor involved. Furthermore, GABA was recently demonstrated to enhance biotic stress tolerance when applied as a priming agent and its effect seems to be dependent on ethylene. Here we will summarize and integrate the current knowledge of GABA and proline accumulation in response to stress and discuss future perspectives.
... Gamma-amino butyric acid (GABA), also known as 4-aminobutyric acid, is a natural, non-protein functional amino acid with substantial medical value widely present in animals, plants, bacteria, fungi, and algae (Shelp et al. 1999). In mammals and humans, GABA is mainly found in the central nervous system and is an important inhibitory neurotransmitter involved in several metabolic and physiological processes. ...
Gamma-amino butyric acid (GABA) is a natural non-protein amino acid involved in stress, signal transmission, carbon and nitrogen balance, and other physiological processes in plants. In the human body, GABA has the effects of lowering blood pressure, anti-aging, and activating the liver and kidneys. However, there are few studies on the molecular regulation mechanism of genes in the metabolic pathways of GABA during grain development of giant embryo rice with high GABA content. In this study, three glant embryo (ge) mutants of different embryo sizes were obtained by CRISPR/Cas9 knockout, and it was found that GABA, protein, crude fat, and various mineral contents of the ge mutants were significantly increased. RNA-seq and qRT-PCR analysis showed that in the GABA shunt and polyamine degradation pathways, the expression levels of most of the genes encoding enzymes promoting GABA accumulation were significantly upregulated in the ge-1 mutant, whereas, the expression levels of most of the genes encoding enzymes involved GABA degradation were significantly downregulated in the ge-1 mutant. This is most likely responsible for the significant increase in GABA content of the ge mutant. These results help reveal the molecular regulatory network of GABA metabolism in giant embryo rice and provide a theoretical basis for the study of its development mechanisms, which is conducive to the rapid cultivation of GABA-rich rice varieties, promoting human nutrition, and ensuring health.
... GABA in higher plants is mainly produced from the glutamate, catalyzed by glutamate decarboxylase (GAD) (Chi et al., 2021;Shelp et al., 1999), some of which also comes from polyamines degradation (Kim et al., 2013;Liao et al., 2017). The pathway of putrescine-derived GABA formation involves a two-step reaction, including diamine oxidase (DAO), or amine oxidase (CuAO), or polyamine oxidase (PAO) catalytic process and followed by an aminoaldehyde dehydrogenase (AMADH) degradation (Moschou et al., 2012;Zhang et al., 2022). ...
Our previous study has shown that copper-containing amine oxidase ( CuAO ) and aminoaldehyde dehydrogenase ( AMADH ) could regulate the accumulation of γ-aminobutyric acid (GABA) in tea plants by participating in polyamine degradation pathway. However, the involvement of these genes in drought tolerance has not been underlined. In this study, CsCuAO1 associated with CsAMADH1 confers drought tolerance, which modulates GABA levels in tea plants, was conducted. The results showed that exogenous GABA spraying effectively alleviated the drought induced physical damage. Overexpression Arabidopsis lines of CsCuAO1 and CsAMADH1 exhibited enhanced resistance to drought, which promoted the synthesis of GABA and putrescine, by alerting reactive oxygen species scavenging capacity and stomatal movement. However, suppression of CsCuAO1 or CsAMADH1 in tea plants exhibited increased sensitivity during drought treatment. Moreover, the co-overexpressed plants increased the accumulation of GABA both in the Agrobacterium -mediated Nicotiana benthamiana transient assay and transgenic Arabidopsis plants. In addition, a GABA transporter, CsGAT1 , was identified, whose expression is strongly correlated with GABA accumulation levels in different tissues under drought stress. Taken together, CsCuAO1 and CsAMADH1 were involved in response to drought stress through a dynamic balance between GABA and putrescine. Our data will greatly contribute to the characterization of GABA biological functions in response to environmental stresses in plants.
... Although the precise pathway(s) causing hypoxia succinate accumulation is yet unknown, it has been proposed that the increased activity of fumarate reductase and GABA shunt under hypoxia may be responsible [24,75]. The GABA shunt starts with the synthesis of GABA from glutamate, followed by the synthesis of succinic semialdehyde, and eventually, succinate [76]. Therefore, hypoxic succinate accumulation may be caused by increased activity of the GABA shunt. ...
Hypoxic stress, caused by the low cellular oxygen in the events of flooding or waterlogging, limits crop productivity in many regions of the world. Hypoxic stress in plants is often dynamic and followed by a reoxygenation process that returns the oxygen level to normal. Although metabolic responses to hypoxia have been studied in many plants, less is known about the recovery processes following stress removal. To better understand the dynamic metabolic shift from a low-oxygen environment to a reoxygenated environment, we performed time-course measurements of metabolites in Arabidopsis seedlings at 0, 6, 12, and 24 h of reoxygenation recovery after 24 h of hypoxia stress (100% N2 environment). Among the 80 metabolic features characterized using GC-MS, 60% of them were significantly changed under hypoxia. The reoxygenation phase was accompanied by progressively fewer metabolic changes. Only 26% significantly changed metabolic features by the 24 h reoxygenation. Hypoxia-induced metabolic changes returned to normal levels at different speeds. For example, hypoxia-induced accumulation of lactate decreased to a basal level after 6 h of reoxygenation, whereas hypoxia-induced accumulation of alanine and GABA showed partial recovery after 24 h of reoxygenation. Some metabolites, such as gluconate, xylose, guanine, and adenosine, constantly increased during hypoxia reoxygenation. These dynamic metabolic changes demonstrate the flexibility and complexity of plant metabolism during hypoxia stress and subsequent reoxygenation recovery.
... Glutamic acid supports plant growth (Al-Juthery et al.,2020 andEl-Metwally et al., 2022). Glutamic acid consider the precursor of γ-aminobutyric acid (GABA) and proline under stress conditions (Shelp et al., 1999 andShang et al., 2011) The purpose of this research was to investigate the possibility of glutamic acid could alleviate the effect of water stress on growth, chemical composition and volatile oil of sweet basil plant cultivated in new reclaimed soil. ...
The present study was conducted during 2019 and 2020 seasons in the Experimental Farm of EL-Quassassin, Horticulture
Research Station, Ismailia Governorate, Egypt to investigate the effect of foliar application of glutamic acid on sweet basil
(Ocimum basilicum, L. var. Grand Vert) plants grown under different irrigation levels. A split-plot design was used, the
main factor was irrigation levels (50%, 75% and 100% of crop evapotranspiration (ETc)), while the glutamic acid at (0, 5
and 10 mM) were assigned in sub-factor. The results showed that irrigation rates had a significant effect on most aspects for
the two cuts in both seasons. Where the irrigation level at 100% ETc was the most effective treatment. The foliar spraying
with glutamic acid on sweet basil plants had a positive effect on plants, it significantly increased growth, yield parameters
and chemical composition in treated plants as compared to un treated ones. Regarding the interaction treatments, it was
worth to be mentioned that, there was insignificant difference, when spraying with glutamic acid under irrigation level at
100 or 75% ETc in most studied traits. Also, the results of Gc- Ms analysis of sweet basil volatile oil indicated that, the
highest linalool content which consider the main component was produced with irrigation level at 75% ETc +spraying with
10mM glutamic acid. Furthermore, glutamic acid foliar application improve irrigation water utilization efficiency
(IW.Ut.E). The highest IW.Ut.E was recorded at the interaction of irrigation level at 50% ETc with foliar application
by10mM glutamic acid.
... Above and beyond this plants have developed powerful tactics to counter biotic stress through hormonal defense mechanisms, especially jasmonic acid and ethylene (ET), which specifically act against necrotrophic pathogens and herbivores, and salicylic acid (SA) and cytokinin (CK) acting against biotrophic and hemibiotrophic pathogens (Gao et al., 2015;Conrath et al., 2015). In these defense responses, several molecules are produced by the plants such as reactive oxygen species (ROS) (Inze and Van Montagu, 2002) and GABA (Shelp et al., 1999;Kinnersley and Turano, 2000), and some defenses are also induced to neighboring plants, which are known as systemic acquired resistance (SAR) and induced systemic resistance (ISR) (Pieterse et al., 2014). ...
... Succinic semialdehyde dehydrogenase (SSADH), GABA transaminase (GABA-T), and glutamine decarboxylase are the three enzymes found to be involved in the TCA cycle (Bouche & Fromm, 2004;Bown & Shelp, 1997). GABA plays multiple roles in plant cells, which include redox regulation, biotic and abiotic defense response, pH regulation, signaling, energy balance, and C-N metabolism (Deewatthanawong et al., 2010). ...
Derived from tryptophan, melatonin (MT; N-acetyl-5-methoxytryptamine) is a ubiquitous indoleamine that is widely distributed in species ranging from microorganisms to mammals. It has been associated to play vital roles in the human body like cardiac rhythms, immunological enhancement, and antioxidant activity. In plants, it is a pleiotropic molecule with multiple roles, as it not only performs extensive functions like delaying senescence, growth and development regulation, exerting antioxidant effects, and facilitating adaption of plants to certain biotic and abiotic stress responses but also provides resistance to chilling injury and disease development. This paper reviews the latest progress in the multiple roles of MT in fruits, summarizes the pathways for its biosynthesis, and presents the relation of MT with plant hormones, like auxin, cytokinin, ethylene, gibberellins, abscisic acid, jasmonic, and salicylic acid, and highlights the effect of postharvest application of MT on physiology and quality of fruits, action mechanisms, and safety regulations of MT. Recent trends focus on using alternatives that are safe for postharvest produce and do not have major side effects. MT is a better alternative to hazardous chemicals being commercially used in the postharvest management of fruits and providing future directions for its utilization.
... In our study, a possible explanation as to why cysteine stimulates long-chain fatty acid synthesis is that it presumably improves membrane integrity for acid resistance to improve cell performance for GABA production. GABA production occurred in acidic milieu since the glutamic acid decarboxylation required H + consumption (Bown and Shelp 1997), which meant cells should regulate intracellular pathways including cell membrane integrity to alleviate acid stress. Some studies clarified fatty acid synthesis regulated membrane integrity and stability. ...
Levilactobacillus brevis NPS-QW-145 isolated from kimchi is deficient in glutamate dehydrogenase-encoding gene (gdhA) to form glutamate, hence it required exogenous supplementation of glutamate/monosodium glutamate (MSG) for decarboxylation reaction to produce γ-aminobutyric acid (GABA). However, GABA conversion rate from MSG was relatively low. The individual effect of 20 amino acids on regulating GABA biosynthesis was investigated. Cysteine was selected to significantly improve GABA production from MSG. It was found that Lb. brevis was capable of producing H2O2, cysteine protected Lb. brevis against H2O2-induced oxidative damage to increase cell viability for the enhancement of GABA production. Moreover, cysteine promoted glucose consumption to produce acetyl-CoA for synthesizing long-chain fatty acids to significantly up-regulate GABA biosynthesis. These findings deciphered antioxidative capability of cysteine in Lb. brevis 145 and provided a theoretical basis for fatty acids synthesis-mediated GABA synthesis in Lb. brevis 145, and possibly in other lactic acid bacteria.
... Considering the positive role of GABA in cold tolerance (Zhu et al., 2019), higher GABA content conferred greater cold tolerance in LJ43. GABA is mainly synthesized from L-glutamate by glutamate decarboxylase, whose activity is Ca 2+ /calmodulin-dependent (Shelp et al., 1999). The calcium signaling pathway is different between cold-resistant and cold-susceptible tea plant cultivars , and there may be a connection between gene expression and the content of related metabolites. ...
Cold stress is a major environmental factor that affects tea production and quality. In this study, the global profiles of metabolites in response to cold acclimation (CA) of three tea plant cultivars with contrasting cold tolerances were investigated using UPLC/MS and GC/MS analyses. A total of 167 and 68 metabolites, which were CA responsive and showed differential accumulation in the susceptible and resistant cultivars under CA, were found from UPLC/MS and GC/MS analyses, respectively. The analysis revealed that higher levels of primary metabolites (amino acids, ascorbic acid, intermediates of the tricarboxylic acid cycle, and carbohydrates) were present in the resistant cultivar. Flavonoids (kaempferol, quercetin, myricetin, and catechin), which are secondary metabolites, also showed high accumulation in the cold-resistant cultivar. Accordingly, under CA, catechin component content in the four cold-resistant accessions was higher than that in the three cold-susceptible accessions, indicating that catechins play important roles in the cold response. Moreover, exogenous epigallocatechin gallate (EGCG) application conferred tolerance to freezing stress in tea plants. Tea leaves treated with EGCG exhibited decreased levels of relative electrolyte leakage and malondialdehyde content and increased reactive oxygen species scavenging activity and Fv/Fm under freezing conditions. Expression analysis of cold-regulated genes indicated that EGCG facilitated the transcriptional activation of CsICE1-CsCBF-CsCOR pathway to improve the freezing tolerance of tea plants. Taken together, the induction of stress tolerance-related metabolites was greater in the cold-resistant cultivar than in susceptible cultivars. This study highlights the important roles of catechins, especially EGCG, in cold tolerance.
... The roles of GABA in plants remain incompletely defined, but it likely participates in intercellular signalling (Kinnersley and Turano 2000), as a reserve of C and N during flooding (together with alanine), in osmoregulation and in the metabolism of reactive oxygen species (Bouché and Fromm 2004, Fait et al. 2008, Mustroph et al. 2014. Production of GABA via glutamate decarboxylation also consumes protons, and may therefore help to alleviate acidification resulting from lactate synthesis (Shelp et al. 1999). Indeed, accumulation of lactate in G. ulmifolia FloC plants at 24 h coincided with GABA accumulation suggesting a link between these metabolites (Figure 4). ...
Flood tolerance is crucial to the survival of tree species subject to long periods of flooding such as those present in the Amazonian Varzea. Tolerance can be mediated by adjustments of metabolism, physiology and morphology, reinforcing the need for investigation of the physiological and biochemical mechanisms used by tropical tree species to survive this stress. Moreover, such mechanisms may vary between populations that are subjected to differences in the frequency of flooding events. Here, we therefore aimed to identify the mechanisms used by two populations of the tropical tree Guazuma ulmifolia (Lam.) to tolerate flooding; an Amazonian population frequently exposed to flooding and a Cerrado population, adapted to a dry environment. Young plants were subjected to flooding of the roots and lower stem for 32 days, followed by 17 days of recovery. Amazonian plants exhibited greater increases in shoot length and higher maximum photosynthetic rate (Amax) compared to non-flooded plants from 7 days of flooding onwards, whilst increased Amax occurred later in flooded Cerrado plants and was not accompanied by increased shoot length. Lactate accumulated in roots of Cerrado plants after 24 h flooding, together with transcripts coding for lactate dehydrogenase in roots of both Cerrado and Amazonian plants. After 7 days of flooding lactate decreased and alcohol dehydrogenase activity increased transiently, together with concentrations of alanine, GABA and succinate, indicating activation of metabolic processes associated with low oxygen availability. Other amino acids also increased in flooded Cerrado plants, revealing more extensive metabolic changes than in Amazonian plants. Wetland and dryland populations of Guazuma ulmifolia revealed great capacity to tolerate flooding stress through a suite of alterations in photosynthetic gas exchange and metabolism. However, the integrated physiological, biochemical and molecular analyses realized here indicated that wetland plants acclimatized more efficiently with increased shoot elongation and more rapid restoration of normal metabolism.
... The reaction mixture (1 mL) containing 50 mM acetate buffer (pH 5.0) and 10 mM 2,2′-azino-di-3-ethylbenzathiazoline-6-sulfo nate (ABTS) was mixed with 200 μL culture filtrate and incubated at 25°C for 5 min with 3 mM of hydrogen peroxide. Absorbance was evaluated at 420 nm for the measurement of peroxidase activity (Shelp et al., 1999). ...
Aims:
To study the altered metabolic pathways and metabolites produced in overexpression and knockdown mutants of a global regulator named MoLAEA, which was recently found to regulate the expression of the genes involved in secondary metabolism in one of the most destructive plant pathogens, Magnaporthe oryzae.
Methods and results:
Mass spectrometry-based global untargeted metabolomic profiling was used to identify altered metabolites. Metabolites were extracted from the mutant strains of MoLAEA using two extraction methods viz., aqueous and organic extraction and data acquired using liquid chromatography-tandem mass spectrometry (LC-MS/MS) in positive and negative polarities. Levels of metabolites involved in various biological pathways such as amino acid as well as polyamine biosynthesis, fatty acid and pyrimidine metabolism showed remarkable change in the mutant strains. Interestingly, metabolites involved in stress responses were produced in higher quantities in the overexpression strain whereas, certain overproduced metabolites were associated with distinctive phenotypic changes in the overexpression strain compared to the wild-type. Further, the expression of several genes involved in the stress responses was found to have higher expression in the overexpression strain.
Conclusions:
The global regulator MoLAEA is involved in secondary metabolism in the plant pathogen M. oryzae such that the mutant strains showed altered level of several metabolites involved in the biosynthesis pathways compared to the wild-type. Also, metabolites involved in stress responses were overproduced in the overexpression strain and this can be seen in the higher growth in media amended with stress-inducing agents or higher expression of genes involved in stress response in the overexpression strain compared to the wild-type.
Significance and impact:
This is the first report of metabolite profiling relative to the global regulation of secondary metabolism in M. oryzae, where secondary metabolism is poorly understood. It opens up avenues for more relevant investigations on the genetic regulation of several of the metabolites found in the analysis, which have not been previously characterized in M. oryzae.
... In MTU 1010 and KNM 118, carbohydrate content of raw and GBR flours at different germination hours (12, (Shelp et al., 1999). Aurisano et al. (1995) found that germinating seed rice under anaerobic conditions for 24 h induced GABA accumulation in the shoot and root. ...
Germinated brown rice (GBR) is an emerging health food that has received attention due to its nutritional composition, especially Gamma Amino Butyric Acid (GABA). The objective of this research was to germinate two brown rice varieties MTU 1010 and KNM 118 at various germination hours (0, 12, 24, 36, 48, 72 hours) to determine the best yield and correlate with the nutritional composition and GABA content. Brown rice soaked in water for 12 hours at 28 ± 2ºC followed by 24 to 36 hours germination was found to have optimum yield (85%) with good nutritional and GABA content. Protein, fat, crude fiber, carbohydrate content and energy content among all samples were highest at 24 hours and 36 hours of germination. Pearson correlation coefficient performed between yield, germination hours and nutritional parameters of both varieties, indicated a positive correlation between germination hours and GABA content in both varieties.
... GABA can accumulate rapidly under biotic and abiotic stress conditions and take part in the defense system against those conditions (Shelp et al., 1999). Accumulation of GABA has been reported in response to low O2 storage in tomatoes (Deewatthanawong et al., 2010). ...
... It consists of three major reactions that are catalyzed by the cytosolic enzyme glutamate decarboxylase (GAD, E.C. 4 In a nonreversible reaction, which limits the GABA synthesis rate, glutamic acid decarboxylase (GAD) mainly catalyzes the a-decarboxylation of glutamate to GABA [16]. GABA is transformed to succinic semialdehyde (SSA) by GABA-T, which is then transformed to succinate by SSADH [17], as described in Figure 1. [18]. ...
aminobutyric acid (GABA) is a non-protein amino acid involved in various physiological processes; it aids in the protection of plants against abiotic stresses, such as drought, heavy metals, and salinity. GABA tends to have a protective effect against drought stress in plants by increasing osmolytes and leaf turgor and reducing oxidative damage via antioxidant regulation. Guard cell GABA production is essential, as it may provide the benefits of reducing stomatal opening and transpiration and controlling the release of tonoplast-localized anion transporter, thus resulting in increased water-use efficiency and drought tolerance. We summarized a number of scientific reports on the role and mechanism of GABA-induced drought tolerance in plants. We also discussed existing insights regarding GABA's metabolic and signaling functions used to increase plant tolerance to drought stress.
... To detect the activities of extracellular peroxidases and laccases in Momyo1 knockdown mutants, mycelial plugs (5 × 5 mm) were firstly cultured on CM amended with 2,2′-azino-di-3-ethylbenzthiazoline-6-sulphonate (ABTS) and/or Congo Red Song et al. 2010). The laccase and peroxidase activities of the Guy1l and Momyo1 knockdown mutants were determined after 48 and 120 h, respectively Shelp et al. 1999). In liquid CM, the extracellular enzymes activity was determined according to the previously established procedures ). ...
In eukaryotes, myosin provides the necessary impetus for a series of physiological processes, including organelle movement, cytoplasmic flow, cell division, and mitosis. Previously, three members of myosin were identified in Magnaporthe oryzae, with class II and class V myosins playing important roles in intracellular transport, fungal growth, and pathogenicity. However, limited is known about the biological function of the class I myosin protein in the rice blast fungus. Here, we found that Momyo1 is highly expressed during conidiation and infection. Functional characterization of this gene via RNA interference (RNAi) revealed that Momyo1 is required for vegetative growth, conidiation, melanin pigmentation, and pathogenicity of M. oryzae. The Momyo1 knockdown mutant is defective in formation of appressorium-like structures (ALS) at the hyphal tips. In addition, Momyo1 also displays defects on cell wall integrity, hyphal hydrophobicity, extracellular enzyme activities, endocytosis, and formation of the Spitzenkörper. Furthermore, Momyo1 was identified to physically interact with the MoShe4, a She4p/Dim1p orthologue potentially involved in endocytosis, polarization of the actin cytoskeleton. Overall, our findings provide a novel insight into the regulatory mechanism of Momyo1 that is involved in fungal growth, cell wall integrity, endocytosis, and virulence of M. oryzae.
Key points
• Momyo1 is required for vegetative growth and pigmentation of M. oryzae.
• Momyo1 is essential for cell wall integrity and endocytosis of M. oryzae.
• Momyo1 is involved in hyphal surface hydrophobicity of M. oryzae.
... Polyaminederived GABA synthesis can also have a significant impact on plant function under certain scenarios (Zarei et al., 2016). GABA synthesis in plants is stimulated by stress, and its known or proposed roles-as a metabolite in plants-were traditionally thought to be confined to processes such as pH regulation, redox status, and carbon-nitrogen balance (Shelp et al., 1999;Batushansky et al., 2014;Bor and Turkan, 2019). ...
While the proposal that γ-aminobutyric acid (GABA) acts a signal in plants is decades old, a signaling mode of action for plant GABA has been unveiled only relatively recently. Here, we review the recent research that demonstrates how GABA regulates anion transport through aluminum-activated malate transporters (ALMTs) and speculation that GABA also targets other proteins. The ALMT family of anion channels modulates multiple physiological processes in plants, with many members still to be characterized, opening up the possibility that GABA has broad regulatory roles in plants. We focus on the role of GABA in regulating pollen tube growth and stomatal pore aperture, and we speculate on its role in long-distance signaling and how it might be involved in cross talk with hormonal signals. We show that in barley (Hordeum vulgare), guard cell opening is regulated by GABA, as it is in Arabidopsis (Arabidopsis thaliana), to regulate water use efficiency, which impacts drought tolerance. We also discuss the links between glutamate and GABA in generating signals in plants, particularly related to pollen tube growth, wounding, and long-distance electrical signaling, and explore potential interactions of GABA signals with hormones, such as abscisic acid, jasmonic acid, and ethylene. We conclude by postulating that GABA encodes a signal that links plant primary metabolism to physiological status to fine tune plant responses to the environment.
... Gamma aminobutyric acid (GABA) is a four-carbon non-proteinogenic amino acid existing in both animals and plants as a considerable component in free amino acid pool (Bown and Shelp 1997). It is synthesized by decarboxylation of glutamic acid and can also be obtained by the degradation of polyamines (Shelp et al. 1999). As an endogenous signaling molecule, GABA is essential for growth regulation and plant development. ...
Heat stress has detrimental impacts on wheat growth and yield formation. Conferring heat tolerance through applying plant growth regulators is a feasible strategy to reduce loss. Gamma aminobutyric acid (GABA) is a four-carbon non-proteinogenic amino acid existing in organisms and accumulates in response to stress. In this study, spring wheat Liaochun17 and winter wheat Jingdu 40 were used to investigate the function of exogenous GABA on the heat tolerance of wheat seedlings. Data displayed that GABA treatment not only reduced the production of reactive active oxygen (ROS), but also improved the scavenging capacity of diphenyl picryl phenyl hydrazine active oxygen under heat stress, thus alleviating the accumulation of malondialdehyde and the damage of cell membrane. In addition, analysis of protein and amino acids revealed that GABA effectively promoted the accumulation of soluble protein and coordinated amino acid homeostasis. Summarily, our current findings revealed that GABA strengthened the resistance of wheat seedling to heat stress by maintaining the metabolism balance of ROS and amino acids.
... GABA is a non-proteinogenic natural amino acid commonly found in animals, plants and microorganisms. It has been demonstrated that GABA possesses many physiological functions such as regulation of blood pressure, heart rate and hormone levels, reduction of blood lipid, as well as improvement of liver and kidney function and so on (Bown and Shelp 1997;Hayakawa et al. 2004;Ma et al. 2020). Body builders also use it to increase muscle growth. ...
γ-aminobutyric acid (GABA) is an important inhibitory neurotransmitter in the human body, but its content decreases with age. So it is suitable to supplement the body's GABA from diet. Hongqu wine is popular because of the addition of Monascus strains in the saccharification process, which makes the wine rich in functional ingredients such as GABA, and monacolin K. In this study, the fermentation parameters of Hongqu wine were optimised to maximise the GABA content through response surface methodology (RSM). The optimal conditions were as follows: 500 g of steamed rice was mixed with 115.4% of boiled water containing 10 g of sodium glutamate and adjusted to pH 3.8 with lactic acid, and then 32% of Hongqu seed inoculum was added. After 5 days of fermentation at 28 °C, 1.5 g of activated yeast was inoculated for ethanol fermentation at 30 °C for 5 days. Finally, the average content of GABA in Hongqu wine amounted to 710.24 mg L<sup>–1</sup>, which is close to the value predicted by RSM model (692.44 mg L<sup>–1</sup>), indicating the statistical fit is good. This provided technical support and theoretical guidance for the production of Hongqu wine rich in GABA by two-stage fermentation.
... The compound γ-aminobutyric acid (GABA) is a four-carbon non-protein amino acid and a natural active ingredient, which has the biological activities of improving brain blood circulation, lowering blood pressure, treating epilepsy and enhancing liver and kidney function Luo et al. 2021;Shelp et al. 1999). Nowadays, GABA is used considerably in pharmaceutical, medical, cosmetics and feed industries (Barrett et al. 2012;Luo et al. 2021;Park et al. 2005). ...
The compound γ-aminobutyric acid (GABA) was widely used in various fields. To enhance the production of GABA in Escherichia coli BL21(DE3), the enzymes of the regeneration pathway of the coenzyme factor pyridoxal 5’-phosphate (PLP) were engineered. The recombinant E. coli strain was screened and identified. The initial concentrations of L-monosodium glutamate (L-MSG) had an obvious influence on the production of GABA. The highest concentration of GABA in recombinant E. coli BL21/pET28a-gadA was 5.54 g/L when the initial L-MSG concentration was 10 g/L, whereas it was 8.45 g/L in recombinant E. coli BL21/pET28a-gadA-SNO1-SNZ1 at an initial L-MSG concentration of 15 g/L. The corresponding conversion yields of GABA in these two strains were 91.0% and 92.7%, respectively. When the initial concentrations of L-MSG were more than 15 g/L, the concentrations of GABA in E. coli BL21/pET28a-gadA-SNO1-SNZ1 were significantly higher as compared to those in recombinant E. coli BL21/pET28a-gadA, and it reached a maximum of 13.20 g/L at an initial L-MSG concentration of 25 g/L, demonstrating that the introduction of the enzymes of the regeneration pathway of PLP favored to enhance the production of GABA. This study provides new insight into producing GABA effectively in E. coli BL21(DE3).
... increase in the levels of alanine and glutamate after 7-and 14-day treatments under all water saturation levels. The GABA shunt has been associated with physiological responses, such as cytosolic pH regulation [86], nitrogen metabolism, and carbon fluxes into the Krebs cycle [87], protection against ROS production [88], and osmoregulation and signaling [89]. The GABA shunt is also involved in carbon and nitrogen metabolism, maintenance of cell membrane integrity [90], and minimizing the negative effects of abiotic stresses on plant metabolism [91]. ...
The physiological and biochemical role of the γ-aminobutyric acid (GABA) shunt pathway in green pea seedlings (Pisum sativum L.) was studied in response to soil water holding capacity levels: 80%, 60%, 40%, 20%, and 10% grown under continuous light at 25 °C for 7 days and 14 days, separately. Characterization of seeds germination pattern, seedlings growth (plant height, fresh and dry weight, and chlorophyll contents), GABA shunt metabolite (GABA, glutamate, and alanine) levels, total protein and carbohydrate levels, and oxidative damage (MDA level) were examined. Data showed a significant effect of drought stress on seed germination, plant growth, GABA shunt metabolites level, total protein and carbohydrate contents, and MDA level. A significant decline in seed germination percentage was recorded at a 20% drought level, which indicated that 20% of soil water holding capacity is the threshold value of water availability for normal germination after 14 days. Seedling fresh weight, dry weight, and plant height were significantly reduced with a positive correlation as water availability was decreased. There was a significant decrease with a positive correlation in Chl a and Chl b contents in response to 7 days and 14 days of drought. GABA shunt metabolites were significantly increased with a negative correlation as water availability decreased. Pea seedlings showed a significant increase in protein content as drought stress was increased. Total carbohydrate levels increased significantly when the amount of water availability decreased. MDA content increased slightly but significantly after 7 days and sharply after 14 days under all water stress levels. The maximum increase in MDA content was observed at 20% and 10% water levels. Overall, the significant increases in GABA, protein and carbohydrate contents were to cope with the physiological impact of drought stress on Pisum sativum L. seedlings by maintaining cellular osmotic adjustment, protecting plants from oxidative stress, balancing carbon and nitrogen (C:N) metabolism, and maintaining cell metabolic homeostasis and cell turgor. The results presented in this study indicated that severe (less than 40% water content of the holding capacity) and long-term drought stress should be avoided during the germination stage to ensure proper seedling growth and metabolism in Pisum sativum L.
... Among the dysregulated metabolites found in the metabolomic result, some were neurotransmitters or had neuroactive functions, such as gamma-aminobutyric acid (GABA), serotonin, and kynurenic acid. GABA is a chief inhibitory neurotransmitter found in the nervous systems and acts by binding to specific transmembrane receptors in the plasma membrane of both pre-and postsynaptic neurons [28]. It has been reported that changes in GABAergic inputs in the paraventricular nucleus maintain sympathetic vasomotor tone in chronic heart failure [29]. ...
Heart failure is a syndrome with symptoms or signs caused by cardiac dysfunction. In clinic, four stages (A, B, C, and D) were used to describe heart failure progression. This study was aimed to explore plasma metabolomic and lipidomic profiles in different HF stages to identify potential biomarkers. Metabolomics and lipidomics were performed using plasma of heart failure patients at stages A (n = 49), B (n = 61), and C+D (n = 26). Analysis of Variance (ANOVA) was used for screening dysregulated molecules. Bioinformatics was used to retrieve perturbed metabolic pathways. Univariate and multivariate receiver operating characteristic curve (ROC) analyses were used for potential biomarker screening. Stage A showed significant difference to other stages, and 142 dysregulated lipids and 134 dysregulated metabolites were found belonging to several metabolic pathways. Several marker panels were proposed for the diagnosis of heart failure stage A versus stage B-D. Several molecules, including lysophosphatidylcholine 18:2, cholesteryl ester 18:1, alanine, choline, and Fructose, were found correlated with B-type natriuretic peptide or left ventricular ejection fractions. In summary, using untargeted metabolomic and lipidomic profiling, several dysregulated small molecules were successfully identified between HF stages A and B-D. These molecules would provide valuable information for further pathological researches and biomarker development.
... The shunt pathway regulates GABA metabolism under stress conditions, strikes a balance of carbon/nitrogen metabolism and influences other physiological processes including plant growth and development (Bouché et al. 2003;Fait et al. 2008;Ansari et al. 2014;Al-Quraan and Al-Share 2016;Jalil et al. 2017Jalil et al. , 2019a. The production of GABA in plants is significantly increased in response to hostile environmental conditions such as hypoxia, low temperature, mechanical stimulation, γ radiation, and low pH (Lane and Stiller 1970;Wallace et al. 1984;Bown and Shelp 1997;Kenersley and Turano 2000). These stresses trigger signal transduction pathway in which enhanced cytosolic Ca 2+ stimulates Ca 2+ /CaM activity of GAD. ...
Unfavorable environmental conditions such as heat, cold, drought, metal/metalloid toxicity, and pathogens enhance production of intra-and inter-cellular levels of reactive oxygen species (ROS) in plants. ROS, acting as signaling molecules, activate signal transduction pathways in response to various stresses. Alternatively, ROS cause irreversible cellular damage due to lipid peroxidation, oxidation of protein, inactivation of enzymes, DNA damage, and interact with other vital constituents of plant cells through their strong oxidative properties, which drastically alter plant morphological structures, becoming disadvantageous for survival and productivity. Higher plants have complex defense systems to scavenge ROS. Being a central molecule of the defense system, gamma-aminobutyric acid (GABA) is ubiquitous from prokaryotes to eukaryotes cells. GABA helps mitigate ROS in plants and GABA shunt pathway plays a key role either as metabolites or endogenous signaling molecules in several regulatory mechanisms under stress conditions. The GABA transporters (GATs) being activated with the attachment of GABA under environmental stress stimuli facilitate high content of Ca2+ into the cytosol. Ca2+ combines with calmodulin (CaM) -binding domain that activates the glutamate decarboxylase (GAD) enzyme for the conversion of glutamate into GABA. This synchronized process regulates GABA shunt gene expressions under stress conditions and improves defense mechanisms in plants. This review highlights the regulatory aspects of GABA shunt pathway for ROS production as well as in the defense mechanism of plants.
... Such substrates that may play a role in cold adaptation include D-mannitol, g-aminobutyric acid, Tween 40, and Tween 80. D-mannitol is a well-known compatible solute, capable of providing cryopreservation, and osmoprotection to psychrophiles in addition to carbon, nitrogen, and energy (Weinstein et al., 1997;Tribelli and López 2018). g-Aminobutyric acid is a compatible solute produced by plants to withstand various stresses such as cold temperatures (Shelp et al., 1999;Mazzucotelli et al., 2006), and given our results, may do the same in bacteria. ...
Martian lava tube caves resulting from a time when the planet was still volcanically active are proposed to contain deposits of water ice, a feature that may increase microbial habitability. In this study, we taxonomically characterized and directly measured metabolic activity of the microbial communities that inhabit lava tube ice from Lava Beds National Monument, an analogue environment to martian lava tubes. We investigated whether this environment was habitable to microorganisms by determining their taxonomic diversity, metabolic activity, and viability using both culture-dependent and culture-independent techniques. With 16S rRNA gene sequencing, we recovered 27 distinct phyla from both ice and ice-rock interface samples, primarily consisting of Actinobacteria, Proteobacteria, Bacteroidetes, Firmicutes, and Chloroflexi. Radiorespiration and Biolog EcoPlate assays found these microbial communities to be metabolically active at both 5°C and -5°C and able to metabolize diverse sets of heterotrophic carbon substrates at each temperature. Viable cells were predominantly cold adapted and capable of growth at 5°C (1.3 × 104 to 2.9 × 107 cells/mL), and 24 of 38 cultured isolates were capable of growth at -5°C. Furthermore, 14 of these cultured isolates, and 16 of the 20 most numerous amplicon sequences we recovered were most closely related to isolates and sequences obtained from other cryophilic environments. Given these results, lava tube ice appears to be a habitable environment, and considering the protections martian lava tubes offer to microbial communities from harsh surface conditions, similar martian caves containing ice may be capable of supporting extant, active microbial communities.
... Drosophila lymph gland blood progenitor cells internalize GABA via GABA transporter (Gat) and catabolize it into succinate (Fig. 1A, Madhwal et al., 2020). Succinic-semialdehyde dehydrogenase (Ssadh), the last and the rate-limiting step of GABA catabolic pathway (Shelp et al., 1999) is responsible for generation of succinate in progenitor cells. Succinate via inhibition of hydroxy prolyl hydroxylase (Hph) activity, stabilizes Sima protein in progenitor cells. ...
The importance of reactive oxygen species (ROS) in myeloid cell development and function is well-established. However, a comprehensive understanding of metabolic states controlling ROS levels during hematopoiesis remains elusive. Myeloid-like blood progenitor cells of the Drosophila larvae reside in a specialized hematopoietic organ called the lymph gland. We find that these progenitors in homeostasis, utilize TCA to generate ROS. Excessive activation of TCA however raises ROS levels causing them to precociously differentiate and leads to retardation of lymph gland size. Thus, to maintain ROS homeostasis, progenitor cells utilize systemically derived GABA. GABA internalization and catabolism via inhibiting hydroxy prolyl hydroxylase (Hph) activity, promotes pyruvate dehydrogenase kinase enzyme activity (PDK). PDK controls inhibitory phosphorylation of pyruvate dehydrogenase (PDH), the rate-limiting enzyme, connecting pyruvate to TCA cycle and OXPHOS. Thus, by regulating PDK, GABA regulates progenitor TCA activity and ROS levels. In addition to this, GABA-catabolism/Hph axis via Hif/Sima drives a glycolytic state in progenitor cells. The dual control established by GABA on PDK and Sima maintains progenitor cell metabolism and sustains ROS homeostasis necessary for their development. Taken together, our study demonstrates the metabolic underpinnings of GABA in myeloid ROS regulation and their development, the relevance of which may be broadly conserved.
... The GABA shunt is a closed system that circumvents multiple steps in the tricarboxylic acid (TCA) cycle to regulate the balance of nitrogen to carbon and involves glutamic acid and proline metabolism (Bouché and Fromm 2004;Signorelli et al. 2015). In this pathway, α-ketoglutarate reacts with glutamate dehydrogenase to yield glutamic acid, and GABA is formed by decarboxylation of glutamic acid (Shelp et al. 1999). Glutamic acid is known to serve as a reservoir of carbon for δ-aminolevulinate during chlorophyll synthesis (Beale et al. 1975), and δ-aminolevulinate is formed when glutamic acid reacts with tRNA, indicating the importance of glutamic acid in chlorophyll biosynthesis (Kannangara et al. 1988). ...
Plant defense against heat stress involves adjustments in amino acid metabolism. The objective of this study was to identify major amino acids and associated metabolic pathways differentially regulated by γ-aminobutyric acid (GABA) and proline that may contribute to augmentation of heat tolerance in cool-season grass species. Creeping bentgrass (Agrostis stolonifera L. cv. ‘Penncross’) was exposed to non-stress (22/18 °C, day/night) or heat stress (35/30 °C, day/night) conditions for 35 d in controlled-environment growth chambers. Non-stressed and heat-stressed plants were foliar-sprayed with water (untreated control), GABA, proline, or ammonium nitrate (N) as a nitrogen source control. Under heat stress, foliar application of GABA, proline, or N significantly increased turf quality and leaf chlorophyll content compared to untreated control plants. N application had nutritional effects, resulting in increases in the content of all amino acids under heat stress. Application of GABA under heat stress significantly increased endogenous content of glutamic acid, GABA, and threonine. Plants treated with proline under heat stress had significantly higher endogenous levels of proline, GABA, glutamic acid, aspartic acid, lysine, isoleucine, leucine, valine, serine, alanine, threonine, and tryptophan compared to untreated controls. The improved heat tolerance in creeping bentgrass pathway by GABA was mainly associated with regulation of amino acid metabolism in the GABA shunt and oxaloacetate pathways. Proline-enhanced heat tolerance involved the regulation of five metabolic pathways (GABA shunt, oxaloacetate, 3-phosphoglycerate, secondary metabolism, and pyruvate). GABA and proline, as well as their responsive amino acids could be used as biomarkers to improve heat tolerance in cool-season grass species.
... Intracellularly, GABA can be 269 catabolized via the GABA-shunt pathway to generate succinate in two steps (Shelp et 270 al., 1999). The final step catalyzed by succinic-semialdehyde dehydrogenase (Ssadh, 271 Figure 2A) is the rate-limiting and critical step of the GABA-catabolic pathway 272 (Shelp et al., 1999). Hence, we manipulated this step by expressing Ssadh RNAi in blood 273 progenitor cells. ...
We show that neuronally released GABA derived upon olfactory stimulation, is utilized by blood-progenitor cells as a metabolite and through its catabolism, these cells stabilize Sima/HIFα protein. Sima capacitates blood-progenitor cells with the ability to initiate lamellocyte differentiation. This systemic axis becomes relevant for larvae dwelling in wasp-infested environments where chances of infection are high. By co-opting the olfactory route, the pre-conditioned animals elevate their systemic GABA levels leading to the up-regulation of blood-progenitor cell Sima expression. This elevates their immune-potential and primes them to respond rapidly when infected with parasitic wasps.
... This observation is in agreement with the previous reports in the literature. Maximum GABA production of L. brevis L-32 was observed between 36 to 72 h of cultivation and GABA was mainly produced during the stationary growth phase [45][46][47]. However, GABA production decreased when the cultivation time was prolonged further. ...
In the present study, we isolated and screened thirty strains of GABA-producing lactic acid bacteria (LAB) from Indonesian traditional fermented foods. Two strains were able to convert monosodium glutamate (MSG) to GABA after 24 h of cultivation at 37oC based on thin layer chromatography (TLC) screening. 16S rDNA sequencing and proteomic identification using MALDI-TOF MS identified these two strains as Lactobacillus plantarum designated as L. plantarum FNCC 260 and L. plantarum FNCC 343. The highest yield of GABA production obtained from the fermentation of L. plantarum FNCC 260 was 809.2 mg/l of culture medium after 60 h of cultivation. Supplementation of 0.6 mM pyridoxal 5’-phosphate (PLP) and 0.1 mM pyridoxine led to the increase in GABA production to 945.3 mg/l and 969.5 mg/l, respectively. The highest GABA production of 1226.5 mg/l of culture medium was obtained with 100 mM initial concentration of MSG added in the cultivation medium. The open reading frame (ORF) of 1410 bp of the gadB gene from L. plantarum FNCC 260 encodes 469 amino acids with a calculated molecular mass of 53.57 kDa. The production of GABA via enzymatic conversion of monosodium glutamate (MSG) using purified recombinant glutamate decarboxylase (GAD) from L. plantarum FNCC 260 expressed in Escherichia coli was found to be more efficient (5-fold higher within 6 h) than the production obtained from fermentation. L. plantarum FNCC 260 could be of interest for the synthesis of GABA.
... GABA is mostly created by glutamate decarboxylase in the cytosol, and transferred to the mitochondria. GABA succinic semi-aldehyde dehydrogenase transfer GABA into succinate in the TCA phase (Fait et al., 2008;Shelp et al., 1999). GABA metabolism was linked with carbon-nitrogen constancy (Bouche and Fromm, 2004;Song et al., 2010). ...
Heat stress has been defined as the rise of temperature for a period of time higher than a threshold level, thereby permanently affecting the plant growth and development. Day or night temperature is considered as the major limiting factor for plant growth. Earlier studies reported that night temperature is an important factor in the heat reaction of the plants. Tomato cultivars capable of setting viable fruits under night temperatures above 21°C are considered as heat-tolerant cultivars. The development of breeding objectives is generally summarized in four points: (a) cultivars with higher yield, (b) disease resistant varieties in the 1970s, (c) long shelf-life in 1980s, and (d) nutritive and taste quality during 1990s. Some unique varieties like the dwarf “Micro-Tom”, and the first transgenic tomato (FlavrSavr) were developed through breeding; they were distributed late in the 1980s.
High temperature significantly affects seed, pollen viability and root expansion. Researchers have employed different parameters to evaluate the tolerance to heat stress, including membrane thermo stability, floral characteristics (Stigma exertion and antheridia cone splitting), flower number, and fruit yield per plant. Reports on pollen viability and fruit set/plant under heat stress by comparing the pollen growth and tube development in heat-treated and non-heat-stressed conditions are available in literature. The electrical conductivity (EC) have been used to evaluate the tolerance of some tomato cultivars in vitro under heat stress conditions as an indication of cell damage due to electrolyte leakage; they classified the cultivars into three groups: (a) heat tolerant, (b) moderately heat tolerant, and (c) heat sensitive.
It is important to determine the range in genetic diversity for heat tolerance in tomatoes. Heat stress experiments under field conditions offer breeders information to identify the potentially heat tolerant germplasm.
The current worldwide context promoting agroecology and green agriculture require the discovery of new ecofriendly and sustainable plant protection tools. Plant resistance inducers, called also elicitors, are one of the most promising alternatives fitting with such requirements. We produced here a set of 30 molecules from pyroglutamic acid, bio-sourced from sugar beet byproducts, and examined for their biological activity on the major agro-economically pathosystem wheat-Zymoseptoria tritici. Foliar application of the molecules provided significant protection rates (up to 63% disease severity reduction) for 16 among them. Structure–activity relationship analysis highlighted the importance of all chemical groups of the pharmacophore in the bioactivity of the molecules. Further investigations using in vitro and in planta antifungal bioassays as well as plant molecular biomarkers revealed that the activity of the molecules did not rely on direct biocide activity towards the pathogen, but rather on the activation of plant defense mechanisms dependent on lipoxygenase, phenylalanine ammonia-lyase, peroxidase, and pathogenesis-related protein pathways. This study reports a new family of bio-sourced resistance inducers and provides new insights into the valorization of agro-resources to develop the sustainable agriculture of tomorrow.
γ-aminobutyric acid (GABA) is a non-protein amino acid which naturally and widely occurs in animals, plants, and microorganisms. As the chief inhibitory neurotransmitter in the central nervous system of mammals, it has become a popular dietary supplement and has promising application in food industry. The current article reviews the most recent literature regarding the physiological functions, preparation methods, enrichment methods, metabolic pathways, and applications of GABA. This review sheds light on developing GABA-enriched plant varieties and food products, and provides insights for efficient production of GABA through synthetic biology approaches.
Changing climatic conditions across Australia’s viticulture regions is placing increasing pressure on resources such as water and energy for irrigation. Therefore, there is a pressing need to identify superior drought tolerant grapevine clones by exploring the extensive genetic diversity of early European clones in old vineyards. Previously, in a field trial, we identified drought-tolerant (DT) dry-farmed Cabernet Sauvignon clones that had higher intrinsic water use efficiency (WUEi) under prolonged soil moisture deficiency compared to drought-sensitive (DS) clones. To investigate whether the field-grown clones have been primed and confer the drought-tolerant phenotypes to their subsequent vegetative progenies, we evaluated the drought responses of DT and DS progenies under two sequential drought events in a glasshouse alongside progenies of commercial clones. The DT clonal progenies exhibited improved gas exchange, photosynthetic performance and WUEi under recurrent drought events relative to DS clonal progenies. Concentration of a natural priming agent, γ-amino butyric acid (GABA), was significantly higher in DT progenies relative to other progenies under drought. Although DT and commercial clones displayed similar drought acclimation responses, their underlying hydraulic, stomatal and photosynthetic regulatory mechanisms were quite distinct. Our study provides fundamental insights into potential intergenerational priming mechanisms in grapevine.
Gamma-amino butyric acid (GABA) is a natural non-protein amino acid involved in plant stress, signal transmission, carbon and nitrogen balance, and other physiological effects in plants. In the human body, it has the effects of lowering blood pressure, anti-aging, and activating the liver and kidney. However, there are few studies on the molecular regulation mechanism of genes in the metabolic pathways of GABA during grain development of giant embryo rice with high GABA content. In this study, three glant embryo (ge) mutants of different embryo sizes were obtained by CRISPR/Cas9 knockout, and it was found that GABA, protein, crude fat, and various mineral contents of the ge mutants were significantly increased. RNA-seq and qRT-PCR analysis showed that in the GABA shunt and polyamine degradation pathway, the expression levels of most of genes encoding enzymes promoting GABA accumulation were significantly upregulated in the ge-1 mutant, while the expression levels of most of genes encoding enzymes conducive to GABA degradation were significantly downregulated in the ge-1 mutant. This is most likely responsible for the significant increase in GABA content of the ge mutant. These results are helpful in revealing the molecular regulatory network of GABA metabolism in giant embryo rice and provide a theoretical basis for the study of its development mechanisms, which is conducive to the rapid cultivation of GABA-rich rice varieties, promoting human nutrition, and ensuring health.
Effect of GABA and Spermine on Morphophysiological Traits of Rosa damascena Mill. in Zanjan Climate
The seasonal and daily dynamics of the content of nitrogen-containing compounds in the leaves of Hylotelephium triphyllum plants was studied. The maximum content of total nitrogen, soluble protein and free amino acids in the leaves was observed in the regrowth phase, followed by their reduction towards the end of the growing season. The daily dynamics was characterized by a high content of free amino acids and a low content of soluble protein in the morning hours. In the leaves of H. triphyllum , 23 free amino acids and 2 amides were found. The seasonal and daily dynamics of free amino acids was determined by the content of glutamine and glutamic acid. 4 non-proteinogenic amino acids were found: -aminobutyric, -alanine, ornithine, citrulline. The total proportion of non-theeinogenic amino acids increased with the aging of the leaves from 0,6% of the sum of amino acids during regrowth to 5,8%, in the fruiting phase, which corresponded to a decrease in the content of soluble protein in them. Seasonal and daily dynamics of non-theeinogenic amino acids was determined mainly by -aminobutyric acid. Alanine and -aminobutyric acid were predominated among the amino acids as an indicator of stress. It is assumed that -aminobutyric acid contributes to the regulation of acidity of cell sap in the leaves of H. triphyllum . At low air and soil humidity (flowering phase), the acidity of the cell sap significantly correlates with the content of free amino acids in the leaves.
Kombucha is a SCOBY-fermented tea beverage known for its taste, sensorial qualities, and high endogenous microbial load. In recent years, kombucha has become a popular functional food with a compound annual growth rate of 25% from 2015 to 2020 in American kombucha sales alone, and sales are predicted to keep increasing significantly over the next decade. However, kombucha is lacking in scientific research, and there is still much to be explored regarding its safety and native probiotic content. This research looked into the various routes of contamination of several kombucha systems as well as the feasibility of the producing a kombucha beverage with health-promoting characteristics derived from the inclusion of lactic probiotics. Although bacterial spore contamination and survival in the kombucha SCOBY have been documented, it is unknown whether spores can survive in the liquid, or whether they can be transmitted to daughter SCOBYs. The foodborne pathogen and spore-former Bacillus cereus was inoculated into the SCOBY, unfermented liquid, and fermented liquid of three different kombucha systems. Data suggest that neither the route of contamination nor the kombucha system influenced the transmission or survival of B. cereus spores. The spread of the spores between culture and liquid across generation was shown to be sporadic but possible, so hygienic handling of kombucha cultures and raw materials throughout the entire production process is crucial to prevent uptake of pathogenic organisms. There was no survival of B. cereus spores after short-term storage or secondary fermentation, indicating that implementation of a holding step may mitigate potential food safety threats. Kombucha is perceived to contain probiotics, but not all live cultures comprise probiotics. Some commercial kombucha products have validated probiotic strains added to them post-fermentation, but this can be costly. If probiotics, such as lactic acid bacteria, are inoculated into sweet tea prior to fermentation, they may be able to acidify the tea, replacing the need for utilizing previous kombucha or acetic acid, or survive and/or produce beneficial metabolites during fermentation in great enough amounts to convey a health benefit upon consumption. The survivability of six probiotic Lactobacillus sp. in acidified, sweetened tea at 25ºC during kombucha fermentation was established, and the medium (tea) and temperature (25ºC) were both revealed to affect the growth rates of the bacteria. Differences in pH indicated that the probiotics were unable to acidify the tea pre-fermentation. Although survival during fermentation was possible for four out of the six probiotics, it was concluded that probiotic Lactobacillus sp. are not well suited for a probiotic kombucha beverages, but out of the tested probiotics, Lactobacillus brevis and Lactobacillus fermentum were the most promising candidates.
Tepung beras pecah kulit berkecambah dapat dimanfaatkan sebagai bahan pangan fungsional karena mengandung senyawa γ-aminobutyric acid (GABA) dan komponen bioaktif lainnya. Penelitian ini bertujuan untuk menganalisis pengaruh waktu perendaman terhadap profil pasting dan komponen bioaktif tepung beras pecah kulit berkecambah. Perkecambahan dilakukan dengan merendam beras pecah kulit selama 120 jam dengan pengambilan sampel setiap 24 jam. Tepung beras pecah kulit berkecambah mengalami penurunan viskositas puncak, breakdown, dan setback seiring dengan lama waktu perendaman. Kandungan GABA mengalami peningkatan dan mencapai nilai tertinggi setelah perendaman 72 jam. Kandungan total fenol, kapasitas antioksidan, dan γ-orizanol mengalami penurunan seiring dengan lamanya waktu perendaman. Sementara itu, hasil analisis komposisi asam lemak tepung beras pecah kulit berkecambah pada perlakuan perendaman 120 jam menunjukkan adanya dominasi asam lemak tidak jenuh. Berdasarkan hasil penelitian ini, tepung beras kecambah dengan perendaman 72 jam dapat dipilih sebagai perlakuan terbaik karena memiliki akumulasi GABA paling tinggi.
Background
As an important germplasm resource, wild soybean has good tolerance to complex stress environment stress. This study described the differences of physiological and metabolomic changes between common wild soybean (GS1) and the barren tolerance wild soybean (GS2) under low nitrogen (LN) stress.
Results
The result showed the barren tolerance wild soybean young leaves can maintain relatively stable chlorophyll content and increased the contents of Car;Photosynthetic rate and transpiration rate decreased significantly in in the barren tolerance wild soybean old leaves, but there was no significant change in young leaves; the barren tolerance wild soybean enhanced the enrichment of beneficial ion pairs such as zinc, calcium and phosphorus. The metabolism of amino acids and organic acids in the barren tolerance wild soybean old leaves was vigorous, a large number of beneficial amino acids such as GABA, asparagine and proline were enriched, and the metabolites related to TCA cycle were significantly increased.
Conclusion
the barren tolerance wild soybean can ensure the nitrogen supply of young leaves by inhibiting the photosynthetic response of old leaves; the relatively stable growth of young leaves also benefits from the effective transport and reuse of beneficial ions from old leaves; More importantly, the enhanced metabolism of specific amino acids and organic acids in GS2 old leaves seemed to play an important role in resisting LN stress. GABA and Asparagine played substantial roles in N storage, C/N balance, antioxidant defense and act as signaling molecule to help GS2 to resist LN stress. Difference organic acids in the old leaves of GS2 increased which could improve the utilization rate of N in the soil. In addition, the strength of fatty acids catabolism and TCA cycle in GS2 old leaves provided energy base for substance transport. The analysis of physiological and metabolite may provide a new perspective for revealing the importance of substance transport and reuse in different plant parts to resist abiotic stress.
Climate variability in arid and semi- arid regions that represents a large sector of Arab region and
East African. This change led to high decreasing in rain rates and consequently water shortage. In
addition to high full price and machine costs, all of that increase usage of hydroponics system. This
system constructed for producing pest organic animal forages. In this champers, we can control
internal condition by using some electronic devises that give the optimum spurting conditions
without adding any aid components. This spurting champers can produce a large amount of green
fodder from the smallest unit area and lese water and full usage. The hydroponic method need
optimum range of temperature, humidity, lighting and irrigation water, so this method is the
optimum for producing large amount of animal fodder with minimum amount of water (save about
90% irrigation water, fertilizers and mechanical works). It is save soil and under soil water from
chemical fertilizers pollination. The aim of study is to study the effect of some weather parameters as
temperature, humidity and lighting; some engineering parameters as selves height and its effect on
produced material from quantity and quality. The treatments include: four experiments of
temperature 20, 22, 24 and 26 ºC; four experiments of humidity 55, 60, 65 and 70, %; four duration
of lighting times 96, 120, 144 and 168, hours.; four experiments of shelves heights 30, 35, 40 and 45,
cm. The measurements include: Some chemical and physical analysis as: Protein percentage; Ash
content; N, %.; P, %.; K, %. and Length of plant. Temperature of 26º C and humidity of 70, % were
the best treatment for plant length, wet weight, N percentage and Protein percentage. It wasn't any
different between Temperature of 26º C and humidity of 70,%; and other experiments of
temperature and humidity in percentage of P,%; K,% and ash content, %, while it were lower in dry
weight. That was related to the effect of increasing temperature and humidity that were very
significant. The effect of increasing temperature and humidity were higher on physical and chemical
perpetrates of plant, that increasing economic benefits of barely spurting under Egyptian local
condition. This temperature and humidity didn’t need huge amount of energy for spurting rooms.
Lighting time of 168 hours was the best on effects of plant length, wet weight, dry weight, N, P, K
and protein percentage, while the best ash content was at 144 hour. There was any difference
between shelf height experiments, but the optimum shelf height was 35 cm. Shelf height of 35 cm
permit good lighting, ventilation, increasing number of used shelves and consequently increase unit
production. By observation, the best plant dry weight was at the second and the third experiment,
but didn't get the pest plant length. That is related to non-spurting barely seeds, little of roots side
extension and the amount of seeds were higher than roots. At lower lighting time, the yellow plant
color was related to lower lighting time. So, we recommended using: A temperature of 26 ± 1 ºC,
humidity of 70%, and lighting time of 168 hours and shelf height of 35 cm.
Background and Aims
Climate change is modifying grape berry composition and affecting wine quality and typicity. We evaluated the effects of leaf-to-fruit ratio and exogenous abscisic acid (ABA) application on the accumulation of primary and secondary metabolites in berries, in order to optimise climate change adaptation strategies.
Methods and Results
A range of leaf-to-fruit ratios (2, 4, 6, 8, 10 or 12 leaves per bunch) and exogenous ABA (400 mg/L) applied to Vitis vinifera L. cv. Cabernet Sauvignon fruiting-cuttings in a greenhouse prior to veraison were evaluated over six consecutive growing seasons (2013–2018). Reducing the leaf-to-fruit ratio decreased berry sugar and anthocyanin concentration, slightly increased total organic acids, and modified the composition of free amino acids. Exogenous ABA significantly enhanced sugar and anthocyanin concentration and partially restored the balance of sugar and anthocyanins under a low leaf-to-fruit ratio, without altering free amino acid concentration or sugar to acids ratios.
Conclusions
Combining manipulation of the leaf-to-fruit ratio with application of exogenous ABA offers a potential method to reduce berry sugar concentration, while maintaining anthocyanin concentration.
Significance of the Study
This study paves the way for possible adaptation strategies for viticulture to global climate change.
In this study, Bacillus strains were isolated from Korean traditional fermented foods, ‘Doenjang’ and ‘Kanjang’, and the fermentative properties of ‘Cheonggukjang’ prepared using these strains were analyzed. During the fermentation of ‘Chenoggukjang’, viable cell count, average β-glycosidase activity, and ammonia level were calculated as 9.49-13.13 log CFU/mL, 22.15 unit/g, and 77.52-273.63 mg/100 g, respectively. γ-aminobutyric acid (GABA) content was highest when using B. subtilis MGD07 (81.33 mg/100 g), whereas it was lowest when using B. cytotoxicus MGD01 (12.56 mg/100 g). Moreover, the presence of β-glycoside forms such as daidzin and genistin, decreased during fermentation, which corresponed to the increase in aglycone forms including daidzein and genistein, observed after fermentation. Additionally, the content of daidzein (249.78 μg/g), glycitein (39.23 μg/g), and genistein (23.61 μg/g) was highest in ‘Cheonggukjang’ prepared using B. licheniformis MGD05 isolated from ‘Doenjang’. Conversely, the total aglycone content was highest (305.55 μg/g) in ‘Cheonggukjang’ prepared using B. subtilis MGK04 isolated from ‘Kanjang’. In addition, the total polyphenol content and antioxidant activity, such as DPPH and ABTS radical scavenging, were higher in all ‘Cheonggukjang’ extracts than in steamed soybeans. In particular, B. amyloliquefaciens MGD02 and B. subtilis MGK02 produced low levels of ammonia and exhibited enhanced functional components (GABA and non-glycoside isoflavones).
Gamma-aminobutyric acid (GABA) is a ubiquitous four-carbon, non-protein amino acid. GABA has been widely studied in animal central nervous systems, where it acts as an inhibitory neurotransmitter. In plants, it is metabolized through the GABA shunt pathway, a bypass of the tricarboxylic acid (TCA) cycle. Additionally, it can be synthesized through the polyamine metabolic pathway. GABA acts as a signal in Agrobacterium tumefaciens-mediated plant gene transformation and in plant development, especially in pollen tube elongation (to enter the ovule), root growth, fruit ripening, and seed germination. It is accumulated during plant responses to environmental stresses and pathogen and insect attacks. A high concentration of GABA elevates plant stress tolerance by improving photosynthesis, inhibiting reactive oxygen species (ROS) generation, activating antioxidant enzymes, and regulating stomatal opening in drought stress. The transporters of GABA in plants are reviewed in this work. We summarize the recent research on GABA function and transporters with the goal of providing a review of GABA in plants.
The pretreatment of rice roots for 1 h in aerobic conditions with the Ca²⁺-channel blockers ruthenium red (RR) and verapamil and the calmodulin (CaM) antagonists N-(6-aminohexyl)-5-chloro-1-naphtylenesulfonamide (W-7) and trifluoperazine, induced during 3 h of anoxia: (i) inhibition of amino acid and γ-aminobutyric acid (Gaba) accumulation; (ii) a decline in the protein content; (iii) a release of amino acids and K⁺ into the growth media. The calcium ionophore A23187 reversed these effects in RR-treated roots. Moreover, the aerobic pretreatment of rice roots with A23187 alone or CaCl2 increased the accumulation of amino acids and Gaba. These data indicate that the Ca²⁺/CaM complex is involved in the transduction of an anaerobic signal by inhibiting proteolysis and solute release, and activating the Ca²⁺/CaM-dependent glutamate decarboxylase.
The pretreatment of rice roots for 1 h in aerobic conditions with the Ca2+-channel blockers ruthenium red (RR) and verapamil and the calmodulin (CaM) antagonists N -(6-aminohexyl)-5-chloro-1-naphtylenesulfonamide (W-7) and trifluoperazine, induced during 3 h of anoxia: (i) inhibition of amino acid and γ-aminobutyric acid (Gaba) accumulation; (ii) a decline in the protein content; (iii) a release of amino acids and K+ into the growth media. The calcium ionophore A23187 reversed these effects in RR-treated roots. Moreover, the aerobic pretreatment of rice roots with A23187 alone or CaCl 2 increased the accumulation of amino acids and Gaba. These data indicate that the Ca2+/CaM complex is involved in the transduction of an anaerobic signal by inhibiting proteolysis and solute release, and activating the Ca2+/CaM-dependent glutamate decarboxylase.
In excised rice roots, anaerobic degradation of proteins gave rise to an increase of free-amino acids. Anoxic accumulation of alanine, γ-aminobutyric acid and proline was the consequence of the interconversion of glutamate, aspartate and amides. The shift in the composition of the amino acid pool appears to be caused by changes in keto acid levels. The role of reactions involved in amino acid interconversion and the physiological significance of these interconversions are considered and discussed.
Molecular procedures have been applied to isolate plant calmodulin-binding proteins. A petunia cDNA expression library was screened with 35S-labeled recombinant calmodulin as a probe, and a cDNA coding for a Ca(2+)-dependent calmodulin-binding protein was isolated. The deduced amino acid sequence of the petunia protein (500 amino acid residues, 58 kDa) has 67% overall amino acid sequence similarity to glutamate decarboxylase (GAD) from Escherichia coli (466 amino acid residues, 53 kDa). The recombinant protein expressed in E. coli cells displays GAD activity, i.e. catalyzes the conversion of glutamic acid to gamma-aminobutyric acid and binds calmodulin, whereas E. coli GAD does not bind calmodulin. The calmodulin binding domain in the petunia GAD was mapped by binding truncated forms of GAD immobilized on nitrocellulose membranes to recombinant petunia 35S-calmodulin as well as to biotinylated bovine calmodulin and by binding truncated forms of GAD to calmodulin-Sepharose columns. The calmodulin binding domain in petunia GAD is part of a carboxyl end extension that is not present in E. coli GAD. Polyclonal antibodies raised against the recombinant petunia GAD detect a single protein band from plant extracts of gel mobility identical to that of the recombinant GAD. Moreover, the plant protein binds calmodulin in vitro. This is the first report of the isolation of a GAD gene from plants and of a calmodulin-binding GAD from any organism. Our results raise the possibility that intracellular Ca2+ signals via calmodulin are involved in the regulation of gamma-aminobutyric acid synthesis in plants.
Primary nitrogen metabolism in transformed root cultures of Datura stramonium was observed by in vivo 15N NMR. Treatment of the root cultures with the plant growth regulators α-naphthaleneacetic acid (NAA) and kinetin caused a
de-differentiation of the root tissue, together with perturbation of primary and secondary nitrogen metabolism. The levels
of newly-synthesized glutamine and glutamate during ammonium assimilation were depleted relative to control cultures, whereas
GABA biosynthesis was enhanced. Although GABA production could be stimulated by a decrease in cytoplasmic pH (whether imposed
artificially or induced by hypoxia), observation of the roots during phytohormone treatment by 31P NMR showed that the cytoplasmic pH remained stable, indicating that the perturbation of nitrogen metabolism in the de-differentiated
roots must be due to other causes.
Methods for measuring plant cytoplasmic calcium using microelectrodes or microinjected fluorescent dyes are associated with extensive technical problems, so measurements have been limited to single or small groups of cells in tissue strips or protoplasts. Aequorin is a calcium-sensitive luminescent protein from the coelenterate Aequorea victoria (A. forskalea) which is formed from apoaequorin, a polypeptide of relative molecular mass approximately 22,000, and coelenterazine, a hydrophobic luminophore. Microinjected aequorin has been widely used for intracellular calcium measurement in animal cells, but its use in plants has been limited to exceptionally large cells. We show here that aequorin can be reconstituted in transformed plants and that it reports calcium changes induced by touch, cold-shock and fungal elicitors. Reconstituted aequorin is cytoplasmic and nonperturbing; measurements can be made on whole plants and a calcium indicator can be constituted in every viable cell. Now that apoaequorin can be targeted to specific organelles, cells and tissues, with the range of coelenterazines with differing calcium sensitivities and properties available, this new method could be valuable for determining the role of calcium in intracellular signalling processes in plants.
A tomato fruit cDNA library was differentially screened to identify mRNAs present at higher levels in fruit of the tomato ripening mutant rin (ripening inhibitor). Complete sequencing of a unique clone ERT D1 revealed an open reading frame with homology to several glutamate decarboxylases. The deduced polypeptide sequence has 80% overall amino acid sequence similarity to a Petunia hybrida glutamate decarboxylase (petGAD) which carries a calmodulin-binding site at its carboxyl terminus and ERT D1 appears to have a similar domain. ERT D1 mRNA levels peaked at the first visible sign of fruit colour change during normal tomato ripening and then declined, whereas in fruit of the ripening impaired mutant, rin, accumulation of this mRNA continued until at least 14 days after the onset of ripening. This mRNA was present at much lower levels in other tissues, such as leaves, roots and stem, and was not increased by wounding. Possible roles for GAD, and its product gamma-aminobutyric acid (GABA) in fruit, are discussed.
We point out an ability of certain amino acids to be recognized at a biological receptor site as though their amino group bore, instead of an alpha relationship to a carboxylate group, a beta, gamma or delta relationship to the same or a second carboxylate group. For aspartate, the unbalanced position of its amino group between a pair of carboxylates allows its occasional biorecognition as a beta-rather than as an alpha-amino acid, whereas for proline and its homologs, their cyclic arrangement may allow the imino group, without its being replicated, to be sensed analogously as falling at either of two distances from the single carboxylate group. The greater separation might allow proline to be seen as biologically analogous to gamma-aminobutyric acid. This more remote positioning of the imino group would allow the D-form of both amino acids to present its amino group in the orientation characteristic of the natural L-form. The dual modes of recognition should accordingly be signalled by what appears to be low stereospecificity, actually due to a distinction in the enantiorecognition of the two isomers. Competing recognition for transport between their respective D- and L-forms, although it does not prove that phenomenon, has been shown for proline and, significantly, even more strongly for its lower homolog, 2-azetidine carboxylate. Such indications have so far revealed themselves rather inconspicuously for the central nervous system binding of proline, reviewed here as a possible feature of a role suspected for proline in neurotransmission.
To date, only plants have been shown to possess a form of glutamate decarboxylase (GAD) that binds calmodulin. In the present study, a recombinant calmodulin-binding 58-kDa petunia GAD produced in Escherichia coli was purified to homogeneity using calmodulin-affinity chromatography, and its responsiveness to calcium and calmodulin was examined in vitro. At pH 7.0-7.5, the purified recombinant enzyme was essentially inactive in the absence of calcium and calmodulin, but it could be stimulated to high levels of activity (Vmax = 30 micromol of CO2 min-1 mg of protein-1) by the addition of exogenous calmodulin (K0.5 = 15 nM) in the presence of calcium (K0.5 = 0.8 microM). Neither calcium nor calmodulin alone had any effect on GAD activity. Recombinant GAD displayed hyperbolic kinetics at pH 7.3 (Km = 8.2 mM). A monoclonal antibody directed against the carboxyl-terminal region, which contains the calmodulin-binding domain of GAD, was able to fully activate GAD in a dose-dependent manner in the absence of calcium and calmodulin, whereas an antibody recognizing an epitope outside of this region was unable to activate GAD. This study provides the first evidence that the activity of the purified 58-kDa GAD polypeptide is essentially calcium/calmodulin-dependent at physiological pH. Furthermore, activation of GAD by two different proteins that interact with the calmodulin-binding domain, a monoclonal antibody or calcium/calmodulin, suggests that this domain plays a major role in the regulation of plant GAD activity.
Glutamate decarboxylase (GAD) catalyzes the decarboxylation of glutamate to CO2 and gamma-aminobutyrate (GABA). GAD is ubiquitous in prokaryotes and eukaryotes, but only plant GAD has been shown to bind calmodulin (CaM). Here, we assess the role of the GAD CaM-binding domain in vivo. Transgenic tobacco plants expressing a mutant petunia GAD lacking the CaM-binding domain (GADdeltaC plants) exhibit severe morphological abnormalities, such as short stems, in which cortex parenchyma cells fail to elongate, associated with extremely high GABA and low glutamate levels. The morphology of transgenic plants expressing the full-length GAD (GAD plants) is indistinguishable from that of wild-type (WT) plants. In WT and GAD plant extracts, GAD activity is inhibited by EGTA and by the CaM antagonist trifluoperazine, and is associated with a CaM-containing protein complex of approximately 500 kDa. In contrast, GADdeltaC plants lack normal GAD complexes, and GAD activity in their extracts is not affected by EGTA and trifluoperazine. We conclude that CaM binding to GAD is essential for the regulation of GABA and glutamate metabolism, and that regulation of GAD activity is necessary for normal plant development. This study is the first to demonstrate an in vivo function for CaM binding to a target protein in plants.
gamma-Aminobutyric acid (GABA), a nonprotein amino acid, is often accumulated in plants following environmental stimuli that can also cause ethylene production. We have investigated the relationship between GABA and ethylene production in excised sunflower (Helianthus annuus L.) tissues. Exogenous GABA causes up to a 14-fold increase in the ethylene production rate after about 12 h. Cotyledons fed with [14C]GABA did not release substantial amounts of radioactive ethylene despite its chemical similarity to 1-aminocyclopropane-1-carboxylic acid (ACC), indicating that GABA is not likely to be an alternative precursor for ethylene. GABA causes increases in ACC synthase mRNA accumulation, ACC levels, ACC oxidase mRNA levels, and in vitro ACC oxidase activity. In the presence of aminoethoxyvinylglycine or alpha-aminoisobutyric acid, GABA did not stimulate ethylene production. We therefore conclude that GABA stimulates ethylene biosynthesis mainly by promoting ACC synthase transcript abundance. Possible roles of GABA as a signal transducer are suggested.
Two distinct cDNA clones encoding for the glutamate decarboxylase (GAD) isoenzymes GAD1 and GAD2 from Arabidopsis (L.) Heynh. were characterized. The open reading frames for GAD1 and GAD2 were expressed in Escherichia coli and the recombinant proteins were purified by affinity chromatography. Analysis of the recombinant proteins by sodium dodecyl sulfate-polyacrylamide gel electrophoresis and immunoblot analysis suggest that GAD1 and GAD2 encode for 58- and 56-kD peptides, respectively. The enzymatic activities of the pure recombinant GAD1 and GAD2 proteins were stimulated 35- and 13-fold, respectively, by Ca2+/calmodulin but not by Ca2+ or calmodulin alone. Southern-blot analysis of genomic DNA suggests that there is only one copy of each gene in Arabidopsis. The GAD1 transcript and a corresponding 58-kD peptide were detected in roots only. Conversely, the GAD2 transcript and a corresponding 56-kD peptide were detected in all organs tested. The specific activity, GAD2 transcript, and 56-kD peptide increased in leaves of plants treated with 10 mM NH4Cl, 5 mM NH4NO3, 5 mM glutamic acid, or 5 mM glutamine as the sole nitrogen source compared with samples from plants treated with 10 mM KNO3. The results from these experiments suggest that in leaves GAD activity is partially controlled by gene expression or RNA stability. Results from preliminary analyses of different tissues imply that these tendencies were not the same in flower stalks and flowers, suggesting that other factors may control GAD activity in these organs. The results from this investigation demonstrate that GAD activity in leaves is altered by different nitrogen treatments, suggesting that GAD2 may play a unique role in nitrogen metabolism.
The acidic, bilobed protein calmodulin (CaM; molecular mass of 16.7 kDa) can activate some 40 distinct proteins in a calcium-dependent manner. The majority of the CaM-binding domain regions of the target proteins are basic and hydrophobic in nature, are devoid of multiple negatively charged residues, and have a propensity to form an alpha-helix. The CaM-binding domain in the C-terminal region of petunia glutamate decarboxylase (PGD) is atypical because it contains five negatively charged residues. Therefore, we chose to study the binding of calcium-CaM to a 26-residue synthetic peptide encompassing the C-terminal region of PGD. Gel band shift assays, fluorescence spectroscopy, and NMR titration studies showed that a single unique complex of calcium-CaM with two PGD peptides is formed. The formation of a 1:2 protein-peptide complex is unusual; normally, calcium-CaM forms 1:1 complexes with the majority of its target proteins. Circular dichroism spectroscopy showed that the bound PGD peptides have an alpha-helical structure. NMR studies of biosynthetically [methyl-13C]methionine-labeled CaM revealed that all the Met side chains in CaM are involved in the binding of the PGD peptides. Analysis of fluorescence spectra showed that the single Trp residue of the two peptides becomes bound to the N- and C-terminal lobes of CaM. These results predict that binding of calcium-CaM to PGD will give rise to dimerization of the protein, which may be necessary for activation. Possible models for the structure of the protein-peptide complex, such as a dimeric peptide structure, are discussed.
Tobacco (Nicotianum tabacum L.) plants engineered to express a sugar beet (Beta vulgaris L.) betaine aldehyde dehydrogenase (BADH) cDNA acquired not only BADH activity, but also three other aldehyde dehydrogenase activities (those measured with 3-dimethylsulfoniopropionaldehyde, 3-aminopropionaldehyde, and 4-aminobutyraldehyde, all of which are natural products). This shows that BADH is not, as believed up to now, a substrate-specific enzyme and that its role may not be limited to glycine betaine synthesis.
The hypothesis that rapid [gamma]-aminobutyrate (GABA) accumulation is a plant defense against phytophagous insects was investigated. Increasing GABA levels in a synthetic diet from 1.6 to 2.6 [mu]mol g-1 fresh weight reduced the growth rates, developmental rates, and survival rates of cultured Choristoneura rosaceana cv Harris larvae. Simulation of the mechanical damage resulting from phytophagous activity increased soybean (Glycine max L.) leaf GABA 10- to 25-fold within 1 to 4 min. Pulverizing leaf tissue resulted in a value of 2.15 ([plus or minus]0.11 SE) [mu]mol GABA g-1 fresh weight.
A cDNA coding for a 58-kD calcium-dependent calmodulin (CaM)-binding glutamate decarboxylase (GAD) previously isolated in our laboratory from petunia (Petunia hybrida) (G. Baum, Y. Chen, T. Arazi, H. Takatsuji, H. Fromm [1993] J Biol Chem 268: 19610-19617) was used to conduct molecular studies of GAD expression. GAD expression was studied during petunia organ development using the GAD cDNA as a probe to detect the GAD mRNA and by the anti-recombinant GAD serum to monitor the levels of GAD. GAD activity was studied in extracts of organs in the course of development. The 58-kD CaM-binding GAD is expressed in all petunia organs tested (flowers and all floral parts, leaves, stems, roots, and seeds). The highest expression levels were in petals of open flowers. Developmental changes in the abundance of GAD mRNA and the 58-kD GAD were observed in flowers and leaves and during germination. Moreover, developmental changes in GAD activity in plant extracts coincided in most cases with changes in the abundance of the 58-kD GAD. We conclude that the 58-kD CaM-binding GAD is a ubiquitous protein in petunia organs and that its expression is developmentally regulated by transcriptional and/or posttranscriptional processes. Thus, GAD gene expression is likely to play a role in controlling the rates of GABA synthesis during petunia seed germination and organ development.
In vivo 15N NMR spectroscopy was used to monitor the assimilation of ammonium by cell-suspension cultures of carrot (Daucus carota L. cv Chantenay). The cell suspensions were supplied with oxygen in the form of either pure oxygen ("oxygenated cells") or air ("aerated cells"). In contrast to oxygenated cells, in which ammonium assimilation had no effect on cytoplasmic pH, ammonium assimilation by aerated cells caused a decrease in cytoplasmic pH of almost 0.2 pH unit. This led to a change in nitrogen metabolism resulting in the accumulation of [gamma]-aminobutyric acid. The metabolic effect of the reduced oxygen supply under aerated conditions could be mimicked by artificially decreasing the cytoplasmic pH of oxygenated cells and was abolished by increasing the cytoplasmic pH of aerated cells. The activity of glutamate decarboxylase increased as the cytoplasmic pH declined and decreased as the pH recovered. These findings are consistent with a role for the decarboxylation of glutamate, a proton-consuming reaction, in the short-term regulation of cytoplasmic pH, and they demonstrate that cytoplasmic pH influences the pathways of intermediary nitrogen metabolism.
[gamma]-Aminobutyric acid (GABA) synthesis (L-glutamic acid + H+ -> GABA + CO2) is rapidly stimulated by a variety of stress conditions including hypoxia. Recent literature suggests that GABA production and concomitant H+ consumption ameliorates the cytosolic acidification associated with hypoxia or other stresses. This proposal was investigated using isolated asparagus (Asparagus sprengeri Regel) mesophyll cells. Cell acidification was promoted using hypoxia, H+/L-glutamic acid symport, and addition of butyrate or other permeant weak acids. Sixty minutes of all three treatments stimulated the levels of both intracellular and extracellular GABA by values ranging from 100 to 1800%. At an external pH of 5.0, addition of 5 mM butyrate stimulated an increase in overall GABA level from 3.86 (0.56 [plus or minus] SE) to 20.4 (2.16 [plus or minus] SE) nmol of GABA/106 cell. Butyrate stimulated GABA levels by 200 to 300% within 15 s, and extracellular GABA was observed after 10 min. The acid load due to butyrate addition was assayed by measuring [14C]butyrate uptake. After 45 s of butyrate treatment, H+-consuming GABA production accounted for 45% of the imposed acid load. The cytosolic location of a fluorescent pH probe was confirmed using fluorescent microscopy. Spectrofluorimetry indicated that butyrate addition reduced cytosolic pH by 0.60 units with a half-time of approximately 2 s. The proposal that GABA synthesis ameliorates cytosolic acidification is supported by the data. The possible roles of H+ and Ca2+ in stimulating GABA synthesis are discussed.
Osmotic adjustment was studied in cultured cells of tomato (Lycopersicon esculentum Mill cv VFNT-Cherry) adapted to different levels of external water potential ranging from -4 bar to -28 bar. The intracellular concentrations of reducing sugars, total free amino acids, proline, malate, citrate, quaternary ammonium compounds, K(+), NO(3) (-), Na(+), and Cl(-) increased with decreasing external water potential. At any given level of adaptation, the maximum contribution to osmotic potential was from reducing sugars followed by potassium ions. The sucrose levels in the cells were 3- to 8-fold lower than reducing sugar levels and did not increase beyond those observed in cells adapted to -16 bar water potential. Concentrations of total free amino acids were 4- to 5-fold higher in adapted cells. Soluble protein levels declined in the adapted cell lines, but the total reduced nitrogen was not significantly different after adaptation. Uptake of nitrogen (as NH(4) (+) or NO(3) (-)) from the media was similar for adapted and unadapted cells. Although the level of quaternary ammonium compounds was higher in the nonadapted cells than that of free proline, free proline increased as much as 500-fold compared to only a 2- to 3-fold increase observed for quaternary ammonium compounds. Although osmotic adjustment after adaptation was substantial (up to -36 bar), fresh weight (volume increase) was restricted by as much as 50% in the adapted cells. Altered metabolite partitioning was evidenced by an increase in the soluble sugars and soluble nitrogen in adapted cells which occurred at the expense of incorporation of sugar into cell walls and nitrogen into protein. Data indicate that the relative importance of a given solute to osmotic adjustment may change depending on the level of adaptation.
In the present study we determined the effects of methionine, intermediates of polyamine catabolic pathways and inhibitors of either ethylene biosynthetic or polyamine catabolic pathways on polyamine accumulation in soybean leaves. Inhibitors to SAM decarboxylase and spermidine synthase, methylglyloxal-bis-(guanylhydrazone) and cyclohexylamine, respectively, suggest that methionine may provide aminopropyl groups for the synthesis of polyamine via S-adenosylmethionine (SAM). Results from experiments that utilized a combination of compounds which altered either ethylene or polyamine biosynthesis, namely, aminoethoxyvinyl glycine, CoSO4, 2,5-norbornadiene, and CuSO4, suggest the two pathways compete for a common precursor. However, exogenous addition of ethylene (via ethephon treatments) had little or no effect on polyamine biosynthesis. Likewise, polyamine treatments had little or no effect on ethylene biosynthesis. These data suggest that there are few or no inhibitory effects from the end products of one pathway on the synthesis of the other. Data from leaves treated with metabolic intermediates in the catabolic pathway of polyamines and inhibitors of enzymes in the catabolic pathway, i.e. aminoguanidine, hydroxyethyldrazine and gabaculine, suggest that the observed increases in polyamine titers were not due to decreased catabolism of the polyamines. One catabolic intermediate, gamma-aminobutyric acid (GABA), elevated putrescine, spermidine and spermine by 12-, 1.4-, and 2-fold, respectively. Ethylene levels decreased (25%) in GABA-treated leaves. This small decrease in ethylene could not account for such large increase in putrescine titers. Further analysis demonstrated that the GABA-mediated polyamine accumulation was inhibited by difluoromethylarginine, an inhibitor of arginine decarboxylase, but not by difluoromethylornithine, an inhibitor of ornithine decarboxylase. These data suggest that GABA directly or indirectly affects the biosynthesis of polyamines via arginine decarboxylase.
[gamma]-Aminobutyric acid (GABA), a nonprotein amino acid, is often accumulated in plants following environmental stimuli that can also cause ethylene production. We have investigated the relationship between GABA and ethylene production in excised sunflower (Helianthus annuus L.) tissues. Exogenous GABA causes up to a 14-fold increase in the ethylene production rate after about 12 h. Cotyledons fed with [14C]GABA did not release substantial amounts of radioactive ethylene despite its chemical similarity to 1-aminocyclopropane-1-carboxylic acid (ACC), indicating that GABA is not likely to be an alternative precursor for ethylene. GABA causes increases in ACC synthase mRNA accumulation, ACC levels, ACC oxidase mRNA levels, and in vitro ACC oxidase activity. In the presence of aminoethoxyvinylglycine or [alpha]-aminoisobutyric acid, GABA did not stimulate ethylene production. We therefore conclude that GABA stimulates ethylene biosynthesis mainly by promoting ACC synthase transcript abundance. Possible roles of GABA as a signal transducer are suggested.
During maturation, pollen undergoes a period of dehydration accompanied by the accumulation of compatible solutes.Solute import across the pollen plasma membrane, which occurs via proteinaceous transporters, is required to support pollen development and also for subsequent germination and pollen tube growth. Analysis of the free amino acid composition of various tissues in tomato revealed that the proline content in flowers was 60 times higher than in any other organ analyzed. Within the floral organs, proline was confined predominantly to pollen, where it represented >70% of total free amino acids. Uptake experiments demonstrated that mature as well as germinated pollen rapidly take up proline. To identify proline transporters in tomato pollen, we isolated genes homologous to Arabidopsis proline transporters. LeProT1 was specifically expressed both in mature and germinating pollen, as demonstrated by RNA in situ hybridization. Expression in a yeast mutant demonstrated that LeProT1 transports proline and γ-amino butyric acid with low affinity and glycine betaine with high affinity. Direct uptake and competition studies demonstrate that LeProT1 constitutes a general transporter for compatible solutes.
A comparative study between leaves of whole plants and calli from leaf was carried out in Lycopersicon exculentum (L.) Mill., L. pennellii (Correll) D'Arcy and their interspecific hybrid with the objective to obtain knowledge of the NaCl stress-induced changes in the total soluble sugars, organic acids, free amino acids and soluble proteins. Similar growth responses to salinity were found in leaf and calli of both parents and their hybrid, because leaf dry weight and the relative growth rate of calli increased in L. pennellii and decreased in L. esculentum and the hybrid at 70 mM NaCl with respect to controls. Total soluble sugar and organic acid contents were much higher in calli than in leaves. Sugars decreased with salinity in calli of L. pennellii and increased significantly in L. esculentum and the hybrid. However, these solutes are not related to salt tolerance in the leaves of whole plants. In general, total organic acids decreased with salinity in both parents and their hybrid at both organizational levels. An increase in total free amino acid contents with salinity was observed at both organizational levels in L. esculentum and the hybrid, while it was only found in leaves of L. pennellii, although to a lower degree. These increases were mainly due to proline and GABA in salt-sensitive L. esculentum and the hybrid, and Asn + Glu in L. pennellii. On the basis of these results, proline and GABA contents may be useful as possible salt-injury sensors, in both leaf and calli from leaf. Moreover, the relation between sugar content and salt-tolerance degree in calli indicates that leaf-cell culture is a good method to study the implication of sugar metabolism in salt-tolerance responses.
Glutamate metabolism was investigated in developing cotyledons of soybean (Glycine max (L.) Merrill). Intact excised cotyledons were injected with carrier-free 14C-glutamate and incubated, in the dark, in sealed vials containing a CO2 trap. Detection of 14C-radioactivity as released CO2, and ethanol-soluble and -insoluble fractions during 40-minute and 6-hour time courses, indicates that exogenously supplied glutamate was metabolized via CO2, 2-oxoglutarate, 4-aminobutyrate and Krebs-cycle intermediates. 14C-Labeled 4-aminobutyrate was recovered from the metabolism of [U-14C]glutamate, but not [1-14C]glutamate, demonstrating its production by decarboxylation of the carbon one of glutamate. The rapid metabolism of [14C]4-aminobutyrate and the recovery of 14C-radioactivity in protein-aspartate suggest that glutamate decarboxylation is not a response to stress, but could contribute Krebs-cycle-derived amino acids for protein synthesis in co-operation with glutamate deamination and transamination. Recovery of 14C as protein-arginine and -glutamate suggests that glutamate was also incorporated into arginine and protein, possibly without degradation. Multiple cellular amino acid pools were suggested from the rate of glutamate incorporation and the significant, but incomplete utilization of the soluble fraction.
Developing cotyledons of soybean. (Glycine max. (L.) Merrill) metabolize glutamate in situ via multiple routes including direct decarboxylation, deamination, and transmination reactions. The production of [14C]4-aminobutyrate . (GABA) from [U-14C]glutamate, but not from [1-14C]glutamate, confirmed that α-decarboxylation is responsible for the production of GABA. [U-14C]GABA was rapidly metabolized to [14C]succinate and [14C]malate, consistent with the entry of glutamate carbon into the Krebs cycle via GABA and succinic semialdehyde. Aminooxyacetate, at a concentration of 100 mM, reduced the in situ metabolism of [U-14C]glutamate and almost fully inhibited [U-14C]GABA synthesis. These data suggested that the GABA shunt . (glutamate → GABA → succinic semialdehyde →succinate) may be important in the glutamate metabolism of a developing soybean seed. Although the GA-BA shunt bypasses the 2-oxoglutarate dehydrogenase reaction of the Krebs cycle, metabolism of [1-14C]acetate or [U-14C]2-oxoglutarate indicated that this reaction is not restricted under our experimental conditions. The in situ glutamate flux through glutamate decarboxylase, as estimated from the changing specific activity of [14C]GABA during the early metabolism of [U-14C]glutamate, indicated that glutamate flux through the GABA shunt is comparable to direct incorporation of glutamate into protein.
Tracerkinetic experiments were performed using L-(guanidino-¹â´C)arginine, L-(U-¹â´C)arginine, L-(ureido-¹â´C)citrulline, and L-(1-¹â´C)ornithine to investigate arginine utilization in developing cotyledons of Gycine max (L.) Merrill. Excised co