Over-expression of a specific soybean GmGSTU4 isoenzyme improves chloroacetanilide herbicide tolerance of transgenic tobacco plants

Plant glutathione transferases (GSTs) have a major role in herbicide detoxification. Soybean (Glycine max L.) GmGSTs have been well studied for their correlation in herbicides selectivity towards diphenyl ether, chloroacetanilide and sulfonylurea herbicides. Chloroacetanilide herbicide tolerance was assayed in vitro by measuring the growth inhibition of wild type (wt) and transgenic tobacco seedlings from cultivars (Basmas, Virginia, Burley) in the presence of 7.5 and 15 mg/L of alachlor and metolachlor. Alachlor caused strong inhibition of shoot and root growth of wt tobacco plants. All the transgenic Basmas lines showed significantly higher shoot and root elongation at 7.5 mg/L alachlor, with line BAGST-3 exhibiting the greatest tolerance. However, at 15 mg/L alachlor, growth was highly reduced in transgenic and wt plants. In Burley, only line BUGST-2 has statistically significant greater mean of root and shoot length compared to wt under the two doses. On the contrary, Virginia has reduced growth which was similar to the wt. Metolachlor toxicity was less severe compared to alachlor. Growth of the transgenic lines of the three cultivars was not significantly greater in either metolachlor concentrations tested, compared to wt plants, except line BAGST-3 which exhibited significantly greater mean of shoot and root elongation at 7.5 mg/L. Transgene expression was determined quantitatively using Real Time qPCR, lines BAGST-3 and BUGST-2 showed greater expression of Gmgstu4 in shoot compared to root. These results confirm that overexpression of GmGSTU4 in tobacco provides higher catalytic activities towards xenobiotics, resulting for future use in environmental cleanup of alachlor

Maintenance of metabolic homeostasis and induction of cytoprotectants and secondary metabolites in alachlor-treated GmGSTU4-overexpressing tobacco plants, as resolved by metabolomics

Herbicides are an invaluable tool for agricultural production scaling up. However, their continuous and intensive use has led to an increased incidence of herbicide resistant weeds and environmental pollution. Plant glutathione transferases (GSTs) are tightly connected with crop and weed herbicide tolerance capacitating their efficient metabolic detoxification, thus GSTs can be biotechnologically exploited towards addressing those issues. However, information on their effects at a “systems” level in response to herbicides is lacking. Here, we aimed to study the effects of the chloroacetanilide herbicide alachlor on the metabolome of wild-type and tobacco plants overexpressing the soybean tau class glutathione transferase GmGSTU4. Alachlor-treated wild-type plants This system, naturally serving the detoxification of endogenous exhibited an abiotic stress-like response with increased abundance of compatible solutes, decrease in TCA cycle intermediates and decreased sugar and amino acid content. Transgenic plants responded distinctly, exhibiting an increased induction of abiotic stress responsive metabolites, accumulation of secondary metabolites and its precursors, and metabolic detoxification by-products compared to wild-type plants. These results suggest that the increased metabolic capacity of GmGSTU4 overexpressing plants is accompanied by pleiotropic metabolic alterations, which could be the target for further manipulation in order to develop herbicide resistant crops, plants with increased phytoremediation potential, as well as efficient management of non-target site, GST induced, herbicide resistance in weeds

Morphological, Physiological and Metabolomic Response of transgenic tobacco plants (N. tabacum L.) overexpressing GmGSTU4 under Drought Stress

GSTs appear to have a significant role in plants’ adaptation under abiotic stress as many isoenzymes are found to be differentially expressed under these conditions yet, little is known about the regulatory functions of GSTs. Wild type and transgenic tobacco plants over-expressing the soybean GmGSTU4 of cultivars Basmas, Burley and Virginia were grown in vitro under 100 and 200mM mannitol or in soil (plant pots) by withholding watering for 15 days. However, GmGSTU4 plants did not exhibit significant differences in drought tolerance compared to wild-type plants. Morphological (shoot length, total and root fresh weight) and physiological (chlorophyll content, relative water content and photosynthetic capacity) parameters of transgenic plants did not differ from the wild-type in the presence of 100 or 200mM mannitol or in the soil when watering was halted. Metabolite profiling was used to understand the dynamics between the wild-type and transgenic tobacco response to drought stress. Different metabolic pathways are involved in production of osmoprotectants. These molecules accumulate in plants under stress conditions as adaptive mechanism, which can provide stress tolerance. GmGSTU4 plants did not exhibit difference in drought tolerance compared to wild-type plants, however metabolomics analysis indicated alterations in metabolite profile and increased concentration of sorbitol, glycerol and pyruvic acid. In conclusion, overexpression of GmGSTU4 in transgenic plants did not affect their drought stress tolerance although it has altered their metabolite profile possible because of diverse effects on plant stress tolerance mechanism

Plant behaviour under combined stress : tomato responses to combined salinity and pathogen stress

Crop plants are subjected to a variety of stresses during their lifecycle, including abiotic stress factors such as salinity and biotic stress factors such as pathogens. Plants have developed a multitude of defense and adaptation responses to these stress factors. In the field, different stress factors mostly occur concurrently resulting in a new state of stress, the combined stress. There is evidence that plant resistance to pathogens can be attenuated or enhanced by abiotic stress factors. With stress tolerance research being mostly focused on plant responses to individual stresses, the understanding of a plant's ability to adapt to combined stresses is limited. In the last few years, we studied powdery mildew resistance under salt stress conditions in the model crop plant tomato with the aim to understand the requirements to achieve plant resilience to a wider array of combined abiotic and biotic stress combinations. We uncovered specific responses of tomato plants to combined salinity-pathogen stress, which varied with salinity intensity and plant resistance genes. Moreover, hormones, with their complex regulation and cross-talk, were shown to play a key role in the adaptation of tomato plants to the combined stress. In this review, we attempt to understand the complexity of plant responses to abiotic and biotic stress combinations, with a focus on tomato responses (genetic control and cross-talk of signaling pathways) to combined salinity and pathogen stresses. Further, we provide recommendations on how to design novel strategies for breeding crops with a sustained performance under diverse environmental conditions

Plant behaviour under combined stress : tomato responses to combined salinity and pathogen stress

Crop plants are subjected to a variety of stresses during their lifecycle, including abiotic stress factors such as salinity and biotic stress factors such as pathogens. Plants have developed a multitude of defense and adaptation responses to these stress factors. In the field, different stress factors mostly occur concurrently resulting in a new state of stress, the combined stress. There is evidence that plant resistance to pathogens can be attenuated or enhanced by abiotic stress factors. With stress tolerance research being mostly focused on plant responses to individual stresses, the understanding of a plant's ability to adapt to combined stresses is limited. In the last few years, we studied powdery mildew resistance under salt stress conditions in the model crop plant tomato with the aim to understand the requirements to achieve plant resilience to a wider array of combined abiotic and biotic stress combinations. We uncovered specific responses of tomato plants to combined salinity-pathogen stress, which varied with salinity intensity and plant resistance genes. Moreover, hormones, with their complex regulation and cross-talk, were shown to play a key role in the adaptation of tomato plants to the combined stress. In this review, we attempt to understand the complexity of plant responses to abiotic and biotic stress combinations, with a focus on tomato responses (genetic control and cross-talk of signaling pathways) to combined salinity and pathogen stresses. Further, we provide recommendations on how to design novel strategies for breeding crops with a sustained performance under diverse environmental conditions

The role of tomato WRKY genes in plant responses to combined abiotic and biotic stresses

In the field, plants constantly face a plethora of abiotic and biotic stresses that can impart detrimental effects on plants. In response to multiple stresses, plants can rapidly reprogram their transcriptome through a tightly regulated and highly dynamic regulatory network where WRKY transcription factors can act as activators or repressors. WRKY transcription factors have diverse biological functions in plants, but most notably are key players in plant responses to biotic and abiotic stresses. In tomato there are 83 WRKY genes identified. Here we review recent progress on functions of these tomato WRKY genes and their homologs in other plant species, such as Arabidopsis and rice, with a special focus on their involvement in responses to abiotic and biotic stresses. In particular, we highlight WRKY genes that play a role in plant responses to a combination of abiotic and biotic stresses.</p

Plant glutathione transferase-mediated stress tolerance : functions and biotechnological applications

Plant glutathione transferases (EC 2.5.1.18, GSTs) are an ancient, multimember and diverse enzyme class. Plant GSTs have diverse roles in plant development, endogenous metabolism, stress tolerance, and xenobiotic detoxification. Their study embodies both fundamental aspects and agricultural interest, because of their ability to confer tolerance against biotic and abiotic stresses and to detoxify herbicides. Here we review the biotechnological applications of GSTs towards developing plants that are resistant to biotic and abiotic stresses. We integrate recent discoveries, highlight, and critically discuss the underlying biochemical and molecular pathways involved. We elaborate that the functions of GSTs in abiotic and biotic stress adaptation are potentially a result of both catalytic and non-catalytic functions. These include conjugation of reactive electrophile species with glutathione and the modulation of cellular redox status, biosynthesis, binding, and transport of secondary metabolites and hormones. Their major universal functions under stress underline the potential in developing climate-resilient cultivars through a combination of molecular and conventional breeding programs. We propose that future GST engineering efforts through rational and combinatorial approaches, would lead to the design of improved isoenzymes with purpose-designed catalytic activities and novel functional properties. Concurrent GST–GSH metabolic engineering can incrementally increase the effectiveness of GST biotechnological deployment

Stress-inducible GmGSTU4 shapes transgenic tobacco plants metabolome towards increased salinity tolerance

The involvement of glutathione transferases (GSTs) in plant’s tolerance to abiotic stresses has been extensively studied; however, the metabolic changes occurring in the plants with altered GSTs expression have not been studied in detail. We have previously demonstrated that GmGSTU4 overexpression in tobacco plants conferred increased tolerance to herbicides, partly through its peroxidase activity. Here, we investigated GmGSTU4 transcriptional response to abiotic and chemical stimuli in soybean. Transgenic tobacco plants overexpressing GmGSTU4 were also evaluated regarding their phenotypic and metabolomics responses under salt stress. GmGSTU4 expression was highly induced after salt stress and atrazine treatment. Tobacco plants overexpressing GmGSTU4 were highly tolerant to 150 mM NaCl in vitro. Metabolomics comparison of plants growing under optimal conditions, indicating a shift of the transgenic plants metabolism towards the metabolic profiles observed under stress, increased concentration of precursors of glutathione biosynthesis and hexose concentration reduction. Under salt stress, transgenic plants maintained their cellular homeostasis in contrast to wild-type plants which exhibited deregulated energy metabolism. The metabolic response of the transgenic plants was characterized by higher concentration of protective metabolites such as proline and trehalose and greater induction of the oxidative pentose phosphate pathway. These results confirm GmGSTU4 contribution to salt stress tolerance, and outline a regulatory role that primes plants towards the up-regulation of protective and detoxification mechanisms under abiotic stress.</p