SummaryD-1-pyrroline-5-carboxylate synthetase enzymes, which catalyse the rate-limiting step of proline biosynthesis, are encoded by two closely related P5CS genes in Arabidopsis. Transcription of the P5CS genes is differentially regulated by drought, salinity and abscisic acid, suggesting that these genes play specific roles in the control of proline biosynthesis. Here we describe the genetic characterization of p5cs insertion mutants, which indicates that P5CS1 is required for proline accumulation under osmotic stress. Knockout mutations of P5CS1 result in the reduction of stress-induced proline synthesis, hypersensitivity to salt stress, and accumulation of reactive oxygen species. By contrast, p5cs2 mutations cause embryo abortion during late stages of seed development. The desiccation sensitivity of p5cs2 embryos does not reflect differential control of transcription, as both P5CS mRNAs are detectable throughout embryonic development. Cellular localization studies with P5CS-GFP gene fusions indicate that P5CS1 is sequestered into subcellular bodies in embryonic cells, where P5CS2 is dominantly cytoplasmic. Although proline feeding rescues the viability of mutant embryos, p5cs2 seedlings undergo aberrant development and fail to produce fertile plants even when grown on proline. In seedlings, specific expression of P5CS2-GFP is seen in leaf primordia where P5CS1-GFP levels are very low, and P5CS2-GFP also shows a distinct cell-type-specific and subcellular localization pattern compared to P5CS1-GFP in root tips, leaves and flower organs. These data demonstrate that the Arabidopsis P5CS enzymes perform non-redundant functions, and that P5CS1 is insufficient for compensation of developmental defects caused by inactivation of P5CS2.
A family tree of the multifunctional proteins, glutathione transferases (GSTs, EC 2.5.1.18) was created in Solanum lycopersicum based on homology to known Arabidopsis GSTs. The involvement of selected SlGSTs was studied in salt stress response of tomato primed with salicylic acid (SA) or in un-primed plants by real-time qPCR. Selected tau GSTs (SlGSTU23, SlGSTU26) were up-regulated in the leaves, while GSTs from lambda, theta, dehydroascorbate reductase and zeta classes (SlGSTL3, SlGSTT2, SlDHAR5, SlGSTZ2) in the root tissues under salt stress. Priming with SA exhibited a concentration dependency; SA mitigated the salt stress injury and caused characteristic changes in the expression pattern of SlGSTs only at 10(-4) M concentration. SlGSTF4 displayed a significant up-regulation in the leaves, while the abundance of SlGSTL3, SlGSTT2 and SlGSTZ2 transcripts were enhanced in the roots of plants primed with high SA concentration. Unexpectedly, under high salinity the SlDHAR2 expression decreased in primed roots as compared to the salt-stressed plants, however, the up-regulation of SlDHAR5 isoenzyme contributed to the maintenance of DHAR activity in roots primed with high SA. The members of lambda, theta and zeta class GSTs have a specific role in salt stress acclimation of tomato, while SlGSTU26 and SlGSTF4, the enzymes with high glutathione conjugating activity, characterize a successful priming in both roots and leaves. In contrast to low concentration, high SA concentration induced those GSTs in primed roots, which were up-regulated under salt stress. Our data indicate that induction of GSTs provide a flexible tool in maintaining redox homeostasis during unfavourable conditions.
SummaryTotal GST (glutathione S-transferase, EC 2.5.1.18) and glutathione peroxidase activity were measured spectrophotometrically in Triticum aestivum cv. MV Emese and cv. Plainsman (drought tolerant) and cv. GK Élet and Cappelle Desprez (drought sensitive) flag leaves under control and drought stress conditions during the grain filling period, in order to reveal possible roles of different GST classes in the senescence of flag leaves. Six wheat GSTs, members of 3 GST classes, were selected and their regulation by drought and senescence was investigated. High glutathione peroxidase activity (EC 1.11.1.9) was measured in wellwatered controls of the drought tolerant Plainsman cultivar. At the same time TaGSTU1B and TaGSTF6 sequences, investigated by Real-Time PCR showed high expression levels that increased with time indicating that the gene products of these genes may play important roles in monocarpic senescence of wheat. The expression levels of these genes were also induced by drought stress in all of the four investigated cultivars, but extremely high transcript amounts were detected in cv. Plainsman. Our data indicate genotypic variations of wheat GSTs; high expression levels and early induction of two senescenceassociated GSTs under drought during grain filling in flag leaves correlated with high yield stability.
The activity and expression of glutathione transferases (GSTs) depend on several less-known endogenous and well-described exogenous factors, such as the developmental stage, presence, and intensity of different stressors, as well as on the absence or presence and quality of light, which to date have received less attention. In this review, we focus on discussing the role of circadian rhythm, light quality, and intensity in the regulation of plant GSTs. Recent studies demonstrate that diurnal regulation can be recognized in GST activity and gene expression in several plant species. In addition, the content of one of their co-substrates, reduced glutathione (GSH), also shows diurnal changes. Darkness, low light or shade mostly reduces GST activity, while high or excess light significantly elevates both the activity and expression of GSTs and GSH levels. Besides the light-regulated induction and dark inactivation of GSTs, these enzymes can also participate in the signal transduction of visible and UV light. For example, red light may alleviate the harmful effects of pathogens and abiotic stressors by increasing GST activity and expression, as well as GSH content in leaves of different plant species. Based on this knowledge, further research on plants (crops and weeds) or organs and temporal regulation of GST activity and gene expression is necessary for understanding the complex regulation of plant GSTs under various light conditions in order to increase the yield and stress tolerance of plants in the changing environment.
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