Manipulation of arginase expression modulates abiotic stress tolerance in Arabidopsis: effect on arginine metabolism and ROS accumulation.
ABSTRACT: Arginine is an important medium for the transport and storage of nitrogen, and arginase (also known as arginine amidohydrolase, ARGAH) is responsible for catalyse of arginine into ornithine and urea in plants. In this study, the impact of AtARGAHs on abiotic stress response was investigated by manipulating AtARGAHs expression. In the knockout mutants of AtARGAHs, enhanced tolerances were observed to multiple abiotic stresses including water deficit, salt, and freezing stresses, while AtARGAH1- and AtARGAH2-overexpressing lines exhibited reduced abiotic stress tolerances compared to the wild type. Consistently, the enhanced tolerances were confirmed by the changes of physiological parameters including electrolyte leakage, water loss rate, stomatal aperture, and survival rate. Interestingly, the direct downstream products of arginine catabolism including polyamines and nitric oxide (NO) concentrations significantly increased in the AtARGAHs-knockout lines, but decreased in overexpressing lines under control conditions. Additionally, the AtARGAHs-overexpressing and -knockout lines displayed significantly reduced relative arginine (% of total free amino acids) relative to the wild type. Similarly, reactive oxygen species accumulation was remarkably regulated by AtARGAHs under abiotic stress conditions, as shown from hydrogen peroxide (H2O2), superoxide radical ( ) concentrations, and antioxidant enzyme activities. Taken together, this is the first report, as far as is known, to provide evidence that AtARGAHs negatively regulate many abiotic stress tolerances, at least partially, attribute to their roles in modulating arginine metabolism and reactive oxygen species accumulation. Biotechnological strategy based on manipulation of AtARGAHs expression will be valuable for future crop breeding.
Project description:Environmental stresses such as drought, salinity, and cold are major factors that significantly limit agricultural productivity. NAC transcription factors play essential roles in response to various abiotic stresses. However, the paucity of wheat NAC members functionally characterized to date does not match the importance of this plant as a world staple crop. Here, the function of TaNAC2 was characterized in Arabidopsis thaliana. A fragment of TaNAC2 was obtained from suppression subtractive cDNA libraries of wheat treated with polyethylene glycol, and its full-length cDNA was obtained by searching a full-length wheat cDNA library. Gene expression profiles indicated that TaNAC2 was involved in response to drought, salt, cold, and abscisic acid treatment. To test its function, transgenic Arabidopsis lines overexpressing TaNAC2-GFP controlled by the cauliflower mosaic virus 35S promoter were generated. Overexpression of TaNAC2 resulted in enhanced tolerances to drought, salt, and freezing stresses in Arabidopsis, which were simultaneously demonstrated by enhanced expression of abiotic stress-response genes and several physiological indices. Therefore, TaNAC2 has potential for utilization in transgenic breeding to improve abiotic stress tolerances in crops.
Project description:Abiotic stresses are major environmental factors that affect agricultural productivity worldwide. NAC transcription factors play pivotal roles in abiotic stress signaling in plants. As a staple crop, wheat production is severely constrained by abiotic stresses whereas only a few NAC transcription factors have been characterized functionally. To promote the application of NAC genes in wheat improvement by biotechnology, a novel NAC gene designated TaNAC67 was characterized in common wheat. To determine its role, transgenic Arabidopsis overexpressing TaNAC67-GFP controlled by the CaMV-35S promoter was generated and subjected to various abiotic stresses for morphological and physiological assays. Gene expression showed that TaNAC67 was involved in response to drought, salt, cold and ABA treatments. Localization assays revealed that TaNAC67 localized in the nucleus. Morphological analysis indicated the transgenics had enhanced tolerances to drought, salt and freezing stresses, simultaneously supported by enhanced expression of multiple abiotic stress responsive genes and improved physiological traits, including strengthened cell membrane stability, retention of higher chlorophyll contents and Na(+) efflux rates, improved photosynthetic potential, and enhanced water retention capability. Overexpression of TaNAC67 resulted in pronounced enhanced tolerances to drought, salt and freezing stresses, therefore it has potential for utilization in transgenic breeding to improve abiotic stress tolerance in crops.
Project description:Ethylene regulates plant abiotic stress responses and tolerances, and ethylene-insensitive3 (EIN3)/EIN3-like (EIL) proteins are the key components of ethylene signal transduction. Although the functions of EIN3/EIL proteins in response to abiotic stresses have been investigated in model plants, little is known in non-model plants, including mulberry (Morus L.), which is an economically important perennial woody plant. We functionally characterized a gene encoding an EIN3-like protein from mulberry, designated as MnEIL3. A quantitative real-time PCR analysis demonstrated that the expression of MnEIL3 could be induced in roots and shoot by salt and drought stresses. Arabidopsis overexpressing MnEIL3 exhibited an enhanced tolerance to salt and drought stresses. MnEIL3 overexpression in Arabidopsis significantly upregulated the transcript abundances of ethylene biosynthetic genes. Furthermore, MnEIL3 enhanced the activities of the MnACO1 and MnACS1 promoters, which respond to salt and drought stresses. Thus, MnEIL3 may play important roles in tolerance to abiotic stresses and the expression of ethylene biosynthetic genes.
Project description:Although allelic diversity of genes has been shown to contribute to many phenotypic variations associated with different physiological processes in plants, information on allelic diversity of abiotic stress-responsive genes is limited. Here it is shown that the alleles OsWRKY45-1 and OsWRKY45-2 play different roles in abscisic acid (ABA) signalling and salt stress adaptation in rice. The two alleles had different transcriptional responses to ABA and salt stresses. OsWRKY45-1-overexpressing lines showed reduced ABA sensitivity, whereas OsWRKY45-1-knockout lines showed increased ABA sensitivity. OsWRKY45-1 transgenic plants showed no obvious difference from negative controls in response to salt stress. In contrast, OsWRKY45-2-overexpressing lines showed increased ABA sensitivity and reduced salt stress tolerance, and OsWRKY45-2-suppressing lines showed reduced ABA sensitivity and increased salt stress tolerance. OsWRKY45-1 and OsWRKY45-2 transgenic plants showed differential expression of a set of ABA- and abiotic stress-responsive genes, but they showed similar responses to cold and drought stresses. These results suggest that OsWRKY45-1 negatively and OsWRKY45-2 positively regulates ABA signalling and, in addition, OsWRKY45-2 but not OsWRKY45-1 negatively regulates rice response to salt stress. The different roles of the two alleles in ABA signalling and salt stress may be due to their transcriptional mediation of different signalling pathways.
Project description:Abiotic stresses negatively affect plants growth and development by inducing protein denaturation, and autophagy degrades the damaged proteins to alleviate their toxicity, however, little is known about the involvement of autophagy in pepper (Capsicum annuum L.) tolerances to abiotic stresses. In this study, we identified autophagy-related gene (ATG) members in the whole genome of pepper by HMM method and analyzed their expression profiles in response to heat and other abiotic stresses by quantitative real-time PCR. The results showed that the CaATG contained 15 core ATG members including 29 ATG proteins with their respective conserved functional domains, involving the whole process of autophagy. Under normal environmental condition, the expression of CaATG genes showed tissue- and developmental stage-specific patterns, while under abiotic stresses of salt, drought, heat, cold and carbohydrate starvation, the accumulation of autophagosome punctate increased and the expression level of CaATG genes changed with stress type-dependent pattern, which indicates the linkage of autophagy in pepper response to abiotic stresses. After treated with heat stress, both the number of up-regulated CaATG genes and the increment of autophagosome punctate were higher in pepper thermotolerant line R9 than those in thermosensitive line B6, implying an association of autophagy with heat tolerance. In addition, CaATG6 was predicted to interact with CaHSP90 family members. Our study suggests that autophagy is connected to pepper tolerances to heat and other abiotic stresses.
Project description:The growth, development, and production of banana plants are constrained by multiple abiotic stressors. However, it remains elusive for the tolerance mechanisms of banana responding to multiple abiotic stresses. In this study, we found that Fen Jiao (FJ) was more tolerant to osmotic, cold, and salt stresses than BaXi Jiao (BX) by phenotypic and physiological analyses. Comparative transcriptomic analyses highlighted stress tolerance genes that either specifically regulated in FJ or changed more than twofold in FJ relative to BX after treatments. In total, 933, 1644, and 133 stress tolerance genes were identified after osmotic, cold, and salt treatments, respectively. Further integrated analyses found that 30 tolerance genes, including transcription factor, heat shock protein, and E3 ubiquitin protein ligase, could be commonly regulated by osmotic, cold, and salt stresses. Finally, ABA and ROS signaling networks were found to be more active in FJ than in BX under osmotic, cold, and salt treatments, which may contribute to the strong stress tolerances of FJ. Together, this study provides new insights into the tolerance mechanism of banana responding to multiple stresses, thus leading to potential applications in the genetic improvement of multiple abiotic stress tolerances in banana.
Project description:MicroRNAs play an important role in abiotic stress responses in higher plants and animals, but their role in stress adaptation in algae remains unknown. In this study, the expression of identified and putative miRNAs in Chlamydomonas reinhardtii was assessed using quantitative polymerase chain reaction; some of the miRNAs (Cre-miR906-3p) were up-regulated, whereas others (Cre-miR910) were down-regulated when the species was subjected to multiple abiotic stresses. With degradome sequencing data, we also identified ATP4 (the d-subunit of ATP synthase) and NCR2 (NADPH: cytochrome P450 reductase) as one of the several targets of Cre-miR906-3p and Cre-miR910, respectively. Q-PCR data indicated that ATP4, which was expressed inversely in relation to Cre-miR906-3p under stress conditions. Overexpressing of Cre-miR906-3p enhanced resistance to multiple stresses; conversely, overexpressing of ATP4 produced the opposite effect. These data of Q-PCR, degradome sequencing and adaptation of overexpressing lines indicated that Cre-miR906-3p and its target ATP4 were a part of the same pathway for stress adaptation. We found that Cre-miR910 and its target NCR2 were also a part of this pathway. Overexpressing of Cre-miR910 decreased, whereas that of NCR2 increased the adaption to multiple stresses. Our findings suggest that the two classes of miRNAs synergistically mediate stress adaptation in algae.
Project description:Abiotic stresses are major limiting factors for growth, development, and productivity of crop plants. Here, we report on OsSKIPa, a rice homolog of human Ski-interacting protein (SKIP) that can complement the lethal defect of the knockout mutant of SKIP homolog in yeast and positively modulate cell viability and stress tolerance of rice. Suppression of OsSKIPa in rice resulted in growth arrest and reduced cell viability. The expression OsSKIPa is induced by various abiotic stresses and phytohormone treatments. Transgenic rice overexpressing OsSKIPa exhibited significantly improved growth performance in the medium containing stress agents (abscisic acid, salt, or mannitol) and drought resistance at both the seedling and reproductive stages. The OsSKIPa-overexpressing rice showed significantly increased reactive oxygen species-scavenging ability and transcript levels of many stress-related genes, including SNAC1 and rice homologs of CBF2, PP2C, and RD22, under drought stress conditions. More than 30 OsSKIPa-interacting proteins were identified, but most of these proteins have no matches with the reported SKIP-interacting proteins in animals and yeast. Together, these data suggest that OsSKIPa has evolved a specific function in positive modulation of stress resistance through transcriptional regulation of diverse stress-related genes in rice.
Project description:NACs are plant-specific transcription factors that have crucial roles in plant development and biotic and/or abiotic stress responses. This study characterized the functions of the soybean NAC gene GmNAC109 using an overexpression construct in Arabidopsis lines. Sequence analysis revealed that GmNAC109 is highly homologous to ATAF1 (Arabidopsis Transcription Activation Factor 1), which regulates biotic and abiotic stress responses. GmNAC109 protein localized to the nucleus and its C-terminal domain exhibited transcriptional activation activity. Salt, dehydration, and cold stresses significantly increased expression of GmNAC109 in soybean. Similarly, Arabidopsis plants overexpressing GmNAC109 were more tolerant to drought and salt stress than wild-type Col-0 plants. Stress response-related genes, such as DREB1A (drought-responsive element-binding 1A), DREB2A, AREB1 (ABSCISIC ACID-RESPONSIVE ELEMENT BINDING PROTEIN 1), AREB2, RD29A (RESPONSIVE TO Desiccation 29A), and COR15A (COLD REGULATED 15A) were upregulated in GmNAC109-overexpressing transgenic Arabidopsis lines. The transgenic lines showed upregulation of the ABA-responsive genes ABI1 (ABA INSENSITIVE 1) and ABI5 and hypersensitivity to ABA. However, GmNAC109 did not increase expression of the ABA-biosynthetic gene NCED3 (NINE-CIS-EPOXYCAROTENOID DIOXYGENASE 3) and endogenous ABA content in the transgenic lines. Overexpression of GmNAC109 significantly increased lateral root formation in transgenic Arabidopsis lines. Expression of AIR3 (AUXIN-INDUCED IN ROOT CULTURES 3) and ARF2 (AUXIN RESPONSE FACTOR 2) was increased and decreased in these transgenic lines, respectively, indicating that GmNAC109 is involved in the auxin signaling pathway and thereby helps to regulate hairy root formation. Our results provide a basis for development of soybean lines with improved tolerance to abiotic stresses via genetic manipulation.
Project description:RING finger proteins play an important role in plant adaptation to abiotic stresses. In the present study, a wild tomato (Solanum habrochaites) cold-induced RING-H2 finger gene, ShATL78L, was isolated, which has been identified as an abiotic stress responsive gene in tomato. The results showed that ShATL78L was constitutively expressed in various tissues such as root, leaf, petiole, stem, flower, and fruit. Cold stress up-regulated ShATL78L in the cold-tolerant S. habrochaites compared to the susceptible cultivated tomato (S. lycopersicum). Furthermore, ShATL78L expression was also regulated under different stresses such as drought, salt, heat, wound, osmotic stress, and exogenous hormones. Functional characterization showed that cultivated tomato overexpressing ShATL78L had improved tolerance to cold, drought and oxidative stresses compared to the wild-type and the knockdown lines. To understand the underlying molecular mechanism of ShATL78L regulating abiotic stress responses, we performed yeast one-hybrid and two-hybrid assays and found that RAV2 could bind to the promoter of ShATL78L and activates/alters its transcription, and CSN5B could interact with ShATL78L to regulate abiotic stress responses. Taken together, these results show that ShATL78L plays an important role in regulating plant adaptation to abiotic stresses through bound by RAV2 and interacting with CSN5B. Highlight: RAV2 binds to the promoter of ShATL78L to activates/alters its transcription to adapt the environmental conditions; furthermore, ShATL78L interacts with CSN5B to regulate the stress tolerance.