CML8 and GAD4 function in (Z)–3–hexenol–mediated defense by regulating GABA accumulation in Arabidopsis

(Z)–3–hexenol, a small gaseous molecule, is produced in plants under biotic stress and induces defense responses in neighboring plants. However, the research on little is known about how (Z)–3–hexenol induces plant defense–related signaling. In this study, we uncovered how (Z)–3–hexenol treatment enhances insect resistance by increasing γ–aminobutyric acid (GABA) contents in Arabidopsis thaliana leaves. First, (Z)–3–hexenol increases the intracellular content of the signaling molecule calcium in Arabidopsis leaf mesophyll cells. Both intracellular and extracellular calcium stores regulate these changes in calcium content. Then, CML8 and GAD4 are involved in calcium signaling. Yeast two–hybrid assays, firefly luciferase complementation imaging, and GST pull–down assays demonstrated that CML8 interacts with GAD4. Finally, (Z)–3–hexenol treatment increased the GABA contents in Arabidopsis leaves, thus increasing plant resistance to the insect Plutella xylostella. This study revealed the mechanism of activating plant insect defense induced by (Z)–3–hexenol, which is of great significance for the study of volatiles as biological control measures.

Optimization of γ-Aminobutyric Acid (GABA) Accumulation in Germinating Adzuki Beans (Vigna angularis) by Vacuum Treatment and Monosodium Glutamate, and the Molecular Mechanisms

This study aimed to investigate the optimal hypoxic and monosodium glutamate (MSG) stress conditions for the enrichment of γ-Aminobutyric acid (GABA) in germinating adzuki beans and to reveal the potential underlying molecular mechanisms of GABA accumulation. Using single-factor experiments and response surface model, we investigated the effects of germination time, germination temperature, vacuum time, and MSG concentration on GABA contents, and further explored the activity and gene expression of glutamate decarboxylase (GAD) and polyamine oxidase (PAO) critical rate restriction enzymes during GABA synthesis. The optimal soaking temperature, soaking time, and pH conditions were 35°C, 16 h, and 5, respectively. Furthermore, the optimal germination conditions for optimal GABA enrichment were 48 h, 1.99 mg/ml MSG concentration, germination temperature of 31.49°C, and vacuum time of 15.83 h. Under such conditions, the predicted GABA concentration was 443.57 ± 7.18 mg/100 g, with no significant difference between the predicted and experimental data. The vacuum + MSG (FZM) treatment has a maximum contribution rate of GABA to 38.29%, which significantly increase GABA content, and the increase was associated with increased GAD and PAO activity. In addition, MSG in combination with vacuum treatment could significantly induce VaGAD4 and VaGAD6 genes in 2 days germination of adzuki beans. According to the results of the present study, vacuum + MSG treatment is an effective approach to enhancing GABA accumulation in germinating adzuki beans, which could be employed in enhancing the functional quality of germinating adzuki beans.

Comparative study on the influence of silicon and selenium to mitigate arsenic induced stress by modulating TCA cycle, GABA and polyamine synthesis in rice seedlings

Arsenic contamination of groundwater is a major concern for its use as drinking water and crop irrigation in many regions of the world. Arsenic is absorbed by rice plants from arsenic contaminated water during irrigation, hampers growth and agricultural productivity. The aim of the study was to mitigate the activity of TCA cycle, synthesis of γ-aminobutyric acid (GABA) and polyamines (PAs) in rice (Oryza sativa L. cv. MTU-1010) seedlings under arsenate (As-V) stress [25 µM, 50 µM and 75 µM] by silicon (Si) [2 mM] and selenium (Se) [5 µM] amendments, and to investigate which chemical was more potential to combat this threat. As(V) application decreased the activities of tested respiratory enzymes while the levels of organic acids (OAs) were increased in the test seedlings. Co-application of Si and As(V) increased the activities of respiratory enzymes, consequently further increased accumulation of OAs that were more than Se with As(V) application in the test seedlings. GABA accumulation along with the activities of its regulatory enzymes were enhanced under As(V) stress. During joint application of Si and As(V) and Se and As(V) said parameters were decreased showing defensive role of these chemicals to resist As(V) toxicity in rice but amendment of Si was more potential than Se amendment resulted reduction of stress induced damage in the test seedlings. PAs trigger tolerance mechanism against stress in plants. PAs viz., Putrescine, spermidine and spermine were synthesized more during Si and Se amendments in As(V) contaminated rice seedlings to combat the effect of stress. Si amendment substantially modulated the toxic effects caused by As(V) over Se amendment in As(V) challenged test seedlings. Thus in future application Si enriched fertilizer will be beneficial than application of Se enriched fertilizer to grow rice plants with normal vigor in arsenic contaminated soil.

Characterization of Genes Involved in γ-Aminobutyric Acid Metabolic Pathways Response to Metabolites Accumulation in Embryos during Barley Germination

To reveal the key enzyme genes involved in γ-aminobutyric acid (GABA) metabolic pathways response to elevated metabolite storage in embryos during barley germination, this study investigated the GABA content, cloned GABA metabolic pathway genes and analyzed their expression levels, respectively. In barley embryos, GABA content continued to rise during the soaking process and then decreased after the germination. Three genes including glutamic acid decarboxylase (GAD), GABA transaminase (GABA-T) and succinic semialdehyde dehydrogenase (SSADH) involved in the GABA pathway were cloned and characterized from the barley embryos, respectively. Before the germination, the expression of GAD gene was up-regulated, while GABA-T gene expression was down-regulated. After the germination, GAD gene expression was lowered, but GABA-T gene expression was rapidly increased. The SSADH gene expression remained stable after soaking of 4 h, and then down-regulated. There is evidence that the high GABA content in germinating barley seeds is parallel with the upregulation of the GAD gene, and down-regulation of GABA-T gene. These results indicate that the expression level of the genes involved in GABA pathway is a crucial factor in GABA accumulation during soaking and germination. This study is beneficial for the development of GABA-rich barley products by germination. © 2021 Friends Science Publishers

Astrocytic GABA Accumulation in Experimental Temporal Lobe Epilepsy

An imbalance of excitation and inhibition has been associated with the pathophysiology of epilepsy. Loss of GABAergic interneurons and/or synaptic inhibition has been shown in various epilepsy models and in human epilepsy. Despite this loss, several studies reported preserved or increased tonic GABAA receptor-mediated currents in epilepsy, raising the question of the source of the inhibitory transmitter. We used the unilateral intracortical kainate mouse model of temporal lobe epilepsy (TLE) with hippocampal sclerosis (HS) to answer this question. In our model we observed profound loss of interneurons in the sclerotic hippocampal CA1 region and dentate gyrus already 5 days after epilepsy induction. Consistent with the literature, the absence of interneurons caused no reduction of tonic inhibition of CA1 pyramidal neurons. In dentate granule cells the inhibitory currents were even increased in epileptic tissue. Intriguingly, immunostaining of brain sections from epileptic mice with antibodies against GABA revealed strong and progressive accumulation of the neurotransmitter in reactive astrocytes. Pharmacological inhibition of the astrocytic GABA transporter GAT3 did not affect tonic inhibition in the sclerotic hippocampus, suggesting that this transporter is not responsible for astrocytic GABA accumulation or release. Immunostaining further indicated that both decarboxylation of glutamate and putrescine degradation accounted for the increased GABA levels in reactive astrocytes. Together, our data provide evidence that the preserved tonic inhibitory currents in the epileptic brain are mediated by GABA overproduction and release from astrocytes. A deeper understanding of the underlying mechanisms may lead to new strategies for antiepileptic drug therapy.

Chitosan regulates metabolic balance, polyamine accumulation, and Na+ transport contributing to salt tolerance in creeping bentgrass

Background Chitosan (CTS), a natural polysaccharide, exhibits multiple functions of stress adaptation regulation in plants. However, effects and mechanism of CTS on alleviating salt stress damage are still not fully understood. Objectives of this study were to investigate the function of CTS on improving salt tolerance associated with metabolic balance, polyamine (PAs) accumulation, and Na+ transport in creeping bentgrass (Agrostis stolonifera). Results CTS pretreatment significantly alleviated declines in relative water content, photosynthesis, photochemical efficiency, and water use efficiency in leaves under salt stress. Exogenous CTS increased endogenous PAs accumulation, antioxidant enzyme (SOD, POD, and CAT) activities, and sucrose accumulation and metabolism through the activation of sucrose synthase and pyruvate kinase activities, and inhibition of invertase activity. The CTS also improved total amino acids, glutamic acid, and γ-aminobutyric acid (GABA) accumulation. In addition, CTS-pretreated plants exhibited significantly higher Na+ content in roots and lower Na+ accumulation in leaves then untreated plants in response to salt stress. However, CTS had no significant effects on K+/Na+ ratio. Importantly, CTS enhanced salt overly sensitive (SOS) pathways and also up-regulated the expression of AsHKT1 and genes (AsNHX4, AsNHX5, and AsNHX6) encoding Na+/H+ exchangers under salt stress. Conclusions The application of CTS increased antioxidant enzyme activities, thereby reducing oxidative damage to roots and leaves. CTS-induced increases in sucrose and GABA accumulation and metabolism played important roles in osmotic adjustment and energy metabolism during salt stress. The CTS also enhanced SOS pathway associated with Na+ excretion from cytosol into rhizosphere, increased AsHKT1 expression inhibiting Na+ transport to the photosynthetic tissues, and also up-regulated the expression of AsNHX4, AsNHX5, and AsNHX6 promoting the capacity of Na+ compartmentalization in roots and leaves under salt stress. In addition, CTS-induced PAs accumulation could be an important regulatory mechanism contributing to enhanced salt tolerance. These findings reveal new functions of CTS on regulating Na+ transport, enhancing sugars and amino acids metabolism for osmotic adjustment and energy supply, and increasing PAs accumulation when creeping bentgrass responds to salt stress.

GABA negatively regulates adventitious root development in poplar

γ-Aminobutyric acid (GABA) influences plant growth, but little is known about how this metabolite regulates adventitious root (AR) development. Here, we investigate the effects of GABA on ARs using poplar lines overexpressing glutamate decarboxilase 2 (GAD2) and by treating poplar stem cuttings with exogenous GABA or vigabatrin (VGB; a specific GABA transaminase inhibitor). Endogenous GABA accumulation not only inhibited AR growth, but it also suppressed or delayed AR formation. Anatomical observations revealed that the GABA and VGB treatments resulted in a 1 d delay in the formation of AR primordia and the appearance of ARs. This delay coincided with changes in primary metabolism, including transient increases in hexose and amino acid levels. GABA-dependent changes in the expression of genes related to hormone synthesis and signalling, as well as analysis of hormone levels revealed that ethylene-dependent pathways were decreased at the earliest stage of AR formation. In contrast, auxin and abscisic acid were increased at 1–5 d as well as GA4 over a 5 d period of AR formation. These results demonstrate that GABA plays a crucial role in AR development. Evidence is presented demonstrating that GABA can interact with hormone-related pathways as well as carbon/nitrogen metabolism. These findings also elucidate the functions of GABA in plant development.