Regulates Salt Stress Tolerance Research Articles

A maize calcineurin B-like interacting protein kinase ZmCIPK42 confers salt stress tolerance.

Calcineurin B-like (CBL) and CBL-interacting protein kinase (CIPK) play a crucial role in biotic and abiotic stress responses. However, the roles of different CIPKs in biotic and abiotic stress responses are less well characterized. In this study, we identified a mutation leading to an early protein termination of the maize CIPK gene ZmCIPK42 that undergoes a G to A mutation at the coding region via searching for genes involved in salt stress tolerance and ion homeostasis from maize with querying the EMS mutant library of maize B73. The mutant zmcipk42 plants have less branched tassel and impaired salt stress tolerance at the seedling stage. Quantitative real-time PCR analysis revealed that ZmCIPK42was expressed in diverse tissues and was induced by NaCl stress. A yeast two-hybrid screen identified a proteinase inhibitor (ZmMPI) as well as calcineurin B-like protein 1 and protein 4 (ZmCBL1, ZmCBL4) as interaction partners of ZmCIPK42. These interactions were further confirmed by bimolecular fluorescence complementation in plant cells. Moreover, over-expressing ZmCIPK42 resulted in enhanced tolerance to high salinity in both maize and Arabidopsis. These findings suggest that ZmCIPK42 is a positive regulator of salt stress tolerance and is a promising candidate gene to improve salt stress tolerance in maize through genetic manipulation.

TGase regulates salt stress tolerance through enhancing bound polyamines-mediated antioxidant enzymes activity in tomato

The accumulation of free polyamines (PAs) plays critical role in plants under abiotic stress conditions, while protective traits of bound PAs receiving much less attention, our understanding of the regulation of free PAs converted to bound PAs is limited. In this study, we demonstrated that the PA mediator transglutaminase (TGase) play an important role in improving tomato tolerance to salinity stress, in part because its critical role in inducing accumulation of bound PAs under salt stress. TGase expression was induced upon salt stress. Exogenous phenanthroline (O-phen, a TGase and bound PAs synthesis inhibitor) abolished the TGase-mediated salt tolerance with terminating bound PAs accumulation in TGaseOE plants. O-phen also had a negatively altered on Na+/K+ uptake or homeostasis under salt stress in leaves of TGaseOE plants. Instead, the overexpression of TGase effectively alleviated salt-induced oxidative damage and reduced reactive oxygen species (ROS) over-production, but it was significantly aggravated by O-phen in the TGaseOE plants. Although the ROS generating enzymes (PA oxidase, diamine oxidase and NADPH oxidase) was not affected by application of O-phen, the antioxidant enzyme (superoxide dismutase, catalase, ascorbate peroxidase and monodehydroascorbate reductase) activity was significantly decreased by exogenous O-phen in plants. Furthermore, further analysis showed that the antioxidant enzyme activity positively and closely correlated with the level of bound PAs in TGaseOE plant. Together, these results uncovered a probably function mechanism of TGase-mediated antioxidant enzyme activity to reduced salt-induced oxidative damage, which is partially depended on bound PAs accumulation.

The effect of ABRE BINDING FACTOR 4-mediated FYVE1 on salt stress tolerance in Arabidopsis

FYVE1 encodes a protein that is localized to the peripheral membrane of late endosomal compartments, and is involved in the regulation of mulitivesicular/prevacuolar compartment protein sorting. It was found that FYVE1 attenuates ABA signaling through degrading ABA receptors PYR1 and PYL4 by ESCRT pathway, and also interacts with transcription factors ABF4 and ABI5 to transcriptionally inhibit ABA signaling pathway by reducing their binding to the cis-regulatory sequences of their downstream genes. However, the mechanisms underlying the transcriptional regulation of FYVE1 and its biological function in salt stress are largely unknown. Here, we show that fyve1 knockdown-mutants show enhanced tolerance to salt stress, while overexpression of FYVE1 results in increased sensitivity to salt stress. Further analysis shows that FYVE1 negatively regulates salt stress tolerance, which is associated with ABA signaling pathway. ABRE BINDING FACTOR 4 (ABF4) directly binds to promoter of FYVE1 to activate its transcription. Moreover, FYVE1 interacts with and promotes degradation of all ABA PYR/PYL receptors. Thus, our results suggest that FYVE1 negatively modulates salt stress tolerance in Arabidopsis via a negative feedback loop.

A leucine-rich repeat receptor-like kinase, OsSTLK, modulates salt tolerance in rice

Leucine-rich repeat receptor-like kinases (LRR-RLKs) have been widely associated with plant abiotic stress responses. However, the functions of the majority of LRR-RLKs has not been well defined. Here, we identified a novel rice LRR-RLK member involved in salt tolerance and designated as OsSTLK (Oryza sativa L. Salt-Tolerance LRR-RLK). Transcript analysis showed that OsSTLK was significantly induced in response to salt stress in rice shoot and root in a time and dosage-dependent fashion. Phenotypic observations indicated that OsSTLK overexpression exhibited reduced salt sensitivity, and improved salt stress tolerance. Further physiological analysis showed that OsSTLK overexpression remarkably reduced electrolyte leakage, malondialdehyde (MDA) content, reactive oxygen species (ROS) accumulation under salt stress conditions by up-regulating ROS-scavenging activities and modifying stomatal patterning. Moreover, Na+/K+ ratio and MAPK phosphorylation level were also reduced in OsSTLK-overexpression transgenic rice plants compared with WT control. Taken together, our findings suggested that OsSTLK as an important positive regulator of salt stress tolerance perhaps through regulating ROS scavenging system, Na+/K+ ratio and MAPK signal pathway.

The wheat E3 ligase TaPUB26 is a negative regulator in response to salt stress in transgenic Brachypodium distachyon

Various abiotic stresses, including high salinity, affect the growth and yield of crop plants. We isolated a gene, TaPUB26, from wheat that encodes a protein containing a U-box domain and armadillo (ARM) repeats. The TaPUB26 transcript levels were upregulated by high salinity, temperature, drought and phytohormones, suggesting the involvement of TaPUB26 in abiotic stress responses. An in vitro ubiquitination assay revealed that TaPUB26 is an E3 ubiquitin ligase. We overexpressed TaPUB26 in Brachypodium distachyon to evaluate TaPUB26 regulation of salt stress tolerance. Compared with the wild type (WT) line, the overexpression lines showed higher salt stress sensitivity under salt stress conditions, but lower chlorophyll (Chl) content, lower photosynthetic levels and overall reduced salt stress tolerance. Additionally, the transgenic plants showed more severe membrane damage, lower antioxidant enzyme activity and more reactive oxygen species (ROS) accumulation than WT plants under salt stress, which might be related to the changes in the expression levels of some antioxidant genes. In addition, the transgenic plants also had higher Na+ and lower K+ contents, thus maintaining a higher cytosolic Na+/K+ ratio in leaves and roots than that in WT plants. Further analysis of the molecular mechanisms showed that TaPUB26 interacted with TaRPT2a, an ATPase subunit of the 26S proteasome complex in wheat. We speculated that TaPUB26 negatively regulates salt stress tolerance by interacting with other proteins, such as TaRPT2a, and that this mechanism involves altered antioxidant competition and cytosolic Na+/K+ equilibrium.

Transcription Factor CaSBP12 Negatively Regulates Salt Stress Tolerance in Pepper (Capsicum annuum L.).

SBP-box (Squamosa-promoter binding protein) genes are a type of plant-specific transcription factor and play important roles in plant growth, signal transduction, and stress response. However, little is known about the role of pepper SBP-box transcription factor genes in response to abiotic stress. Here, one of the pepper SBP-box gene, CaSBP12, was selected and isolated from pepper genome database in our previous study. The CaSBP12 gene was induced under salt stress. Silencing the CaSBP12 gene enhanced pepper plant tolerance to salt stress. The accumulation of reactive oxygen species (ROS) of the detached leaves of CaSBP12-silenced plants was significantly lower than that of control plants. Besides, the Na+, malondialdehyde content, and conductivity were significantly increased in control plants than that in the CaSBP12-silenced plants. In addition, the CaSBP12 over-expressed Nicotiana benthamiana plants were more susceptible to salt stress with higher damage severity index percentage and accumulation of ROS as compared to the wild-type. These results indicated that CaSBP12 negatively regulates salt stress tolerance in pepper may relate to ROS signaling cascades.

The involvement of wheat U-box E3 ubiquitin ligase TaPUB1 in salt stress tolerance.

U-box E3 ubiquitin ligases play important roles in the ubiquitin/26S proteasome machinery and in abiotic stress responses. TaPUB1-overexpressing wheat (Triticum aestivum L.) were generated to evaluate its function in salt tolerance. These plants had more salt stress tolerance during seedling and flowering stages, whereas the TaPUB1-RNA interference (RNAi)-mediated knock-down transgenic wheat showed more salt stress sensitivity than the wild type (WT). TaPUB1 overexpression upregulated the expression of genes related to ion channels and increased the net root Na+ efflux, but decreased the net K+ efflux and H+ influx, thereby maintaining a low cytosolic Na+ /K+ ratio, compared with the WT. However, RNAi-mediated knock-down plants showed the opposite response to salt stress. TaPUB1 could induce the expression of some genes that improved the antioxidant capacity of plants under salt stress. TaPUB1 also interacted with TaMP (Triticum aestivum α-mannosidase protein), a regulator playing an important role in salt response in yeast and in plants. Thus, low cytosolic Na+ /K+ ratios and better antioxidant enzyme activities could be maintained in wheat with overexpression of TaPUB1 under salt stress. Therefore, we conclude that the U-box E3 ubiquitin ligase TaPUB1 positively regulates salt stress tolerance in wheat.

Histone deacetylase gene PtHDT902 modifies adventitious root formation and negatively regulates salt stress tolerance in poplar

Histone deacetylases (HDACs) regulate gene transcription, and play a critical role in plant growth, development and stress responses. HD2 proteins are plant specific histone deacetylases. In woody plants, functions of HD2s are not known. In this study, we cloned an HD2 gene PtHDT902 from Populus trichocarpa and investigated its sequence, expression, subcellular localization, and functions in root development and salt stress responses. Our findings indicated that PtHDT902 was a nuclear protein and its expression was regulated by abiotic stresses. The over-expression of PtHDT902 in both Arabidopsis and poplar increased the expression levels of gibberellin (GA) biosynthetic genes. The expression of PtHDT902 in Arabidopsis enhanced primary root growth, and its over-expression in poplar inhibited adventitious root formation. These phenotypes resulted from over-expression of PtHDT902 were consistent with the GA-overproduction phenotypes. In addition, the poplar plants over-expressing PtHDT902 exhibited lower tolerance to salt than non-transgenic plants. These findings indicated that PtHDT902 worked as an important regulator in adventitious root formation and salt stress tolerance in poplar.

Mulberry RGS negatively regulates salt stress response and tolerance

ABSTRACTRegulator of G-protein signaling (RGS) protein, the best-characterized accelerating GTPase protein in plants, regulates G-protein signaling and plays important role in abiotic stress tolerance. However, the detailed molecular mechanism of RGS involved in G-protein signaling mediated abiotic stress responses remains unclear. In this study, a mulberry (Morus alba L.) RGS gene (MaRGS) was transformed into tobacco, and the ectopic expression of MaRGS in tobacco decreased the tolerance to salt stress. The transgenic tobacco plants had lower proline content, higher malonaldehyde and H2O2 contents than wild type plants under salt stress condition. Meanwhile, MaRGS overexpression in mulberry seedlings enhances the sensitivity to salt stress and RNAi-silenced expression of MaRGS improves the salt stress response and tolerance. These results suggested that MaRGS negatively regulates salt stress tolerance. Further analysis suggested that D-glucose and autophagy may involve in the response of RGS to salt stress. This study revealed the role of MaRGS in salt stress tolerance and provides a proposed model for RGS regulates G-protein signaling in response to salt stress.

Retraction Note: Genetic regulation of salt stress tolerance revealed by RNA-Seq in cotton diploid wild species, Gossypium davidsonii

This article has been retracted.

CYSTM3 negatively regulates salt stress tolerance in Arabidopsis.

CYSTM3, a small mitochondrial protein, acts as a negative regulator in salt stress response by preventing Na+ efflux and disturbing reactive oxygen species (ROS) homeostasis in Arabidopsis. Cysteine-rich transmembrane module (CYSTM) is a not well characterized small peptide family in plants. In this study, we identified a novel mitochondrion-localized CYSTM member CYSTM3 from Arabidopsis, which was ubiquitously expressed in different tissues and dramatically induced by salt stress. Transgenic plants overexpressing CYSTM3 (OE) displayed hypersensitivity to salt stress compared with wild type (WT) plants, whereas a knockout mutant cystm3 was more tolerant to high salinity than WT. Moreover, OE lines accumulated higher contents of Na+ and ROS than WT and cystm3 upon exposure to high salinity. Further analysis revealed that CYSTM3 could deter root Na+ efflux and inhibit the activities of a range of ROS scavenging enzymes in Arabidopsis. In addition, the transcripts of nuclear salt stress-responsive genes were over-activated in cystm3 than those in WT and OE lines. Taken together, Arabidopsis CYSTM3 acts as a negative regulator in salt stress tolerance.

Overexpression of the Wild Soybean R2R3-MYB Transcription Factor GsMYB15 Enhances Resistance to Salt Stress and Helicoverpa Armigera in Transgenic Arabidopsis.

Plant R2R3-MYB transcription factors (TFs) have been suggested to play crucial roles in the response to diverse abiotic and biotic stress factors but there is little molecular evidence of this role in soybean plants. In this work, we identified and functionally characterized an R2R3-MYB TF, namely, GsMYB15, from the wild soybean ED059. Protein and promoter sequence analysis indicated that GsMYB15 is a typical R2R3-MYB TF and contains multiple stress-related cis-elements in the promoter region. GsMYB15 is located in the nucleus and exhibits transcriptional activation activity. QPCR assays suggested that the expression of GsMYB15 could be induced by NaCl, insect attacks and defense-related hormones (MeJA and SA). Furthermore, GsMYB15 exhibited highest expression in pods compared to other tissues. Functional analysis of GsMYB15 demonstrated that overexpression of GsMYB15 could increase salt tolerance and enhance the resistance to H. armigera larvae in transgenic Arabidopsis plants. Moreover, overexpression of GsMYB15 also affected the expression levels of salt stress- and defense-related genes in the transgenic plants. Feeding with transgenic Arabidopsis plant leaves could significantly suppress the expression levels of immunity-related genes in H. armigera larvae. Overexpression of GsMYB15 also increased mesophyll cell levels in transgenic plants. Taken together, these results provide evidence that GsMYB15 is a positive regulator of salt stress tolerance and insect resistance in transformed Arabidopsis plants.

The Novel Rose MYB Transcription Factor RhMYB96 Enhances Salt Tolerance in Transgenic Arabidopsis

MYB transcription factors play diverse roles in plant development and responses to abiotic stresses. However, the biological roles of MYB family members in rose (Rosa hybrida) are still unknown. Here, we reported the isolation and functional characterization of a novel rose RhMYB96 gene. RhMYB96 have a close relationship to Arabidopsis AtMYB96, including the presence of typical R2- and R3-domains at the N-terminal. Salt, dehydration, or ethylene signaling induce the expression of RhMYB96. RhMYB96 localized in the nucleus when expressed in Arabidopsis thaliana. We generated three independent transgenic Arabidopsis overexpressing RhMYB96 to explore the function of RhMYB96 in salt tolerance. Overexpression of RhMYB96 in Arabidopsis resulted in NaCl insensitivity during germination. Furthermore, RhMYB96-overexpressing seedlings under salt stress had longer primary root length and increased biomass. Specifically, the RhMYB96-overexpressed Arabidopsis plants displayed higher capability of scavenging the reactive oxygen levels H2O2 and O2− under salt stress. In addition, overexpressing RhMYB96 improved abscisic acid (ABA) sensitivity during germination. Expression of abiotic stresses and ABA-related genes was stronger in RhMYB96-overexpressing plants. These findings suggest that the RhMYB96 plays and regulates salt stress tolerance modulating an ABA-mediated pathway. Main Conclusion The novel rose RhMYB96 regulates salt stress through the modulation of an ABA-mediated pathway in Arabidopsis.

Pectin methylesterase31 positively regulates salt stress tolerance in Arabidopsis

The alteration of cell wall component and structure is an important adaption to saline environment. Pectins, a major cell wall component, are often present in a highly methylesterified form. The level of methyl esterification determined by pectin methylesterases (PMEs) influences many important wall properties that are believed to relate to the adaption to saline stress. However, little is known about the function of PMEs in response to salt stress. Here, we established a link between pectin methylesterase31 (PME31) and salt stress tolerance. Salt stress significantly increases PME31 expression. PME31 is located in the plasma membrane and the expression level of PME31 was high in dry seeds. Knock-down mutants in PME31 conferred hypersensitive phenotypes to salt stress in seed germination and post-germination growth. Real-time PCR analysis revealed that the transcript levels of several stress genes (DREB2A, RD29A and RD29B) are lower in pme31-2 mutant than that in the wild type in response to salt stress. These results suggested that PME31 could positively modulate salt stress tolerance.

The 2'-O-methyladenosine nucleoside modification gene OsTRM13 positively regulates salt stress tolerance in rice.

Stress induces changes of modified nucleosides in tRNA, and these changes can influence codon-anticodon interaction and therefore the translation of target proteins. Certain nucleoside modification genes are associated with regulation of stress tolerance and immune response in plants. In this study, we found a dramatic increase of 2'-O-methyladenosine (Am) nucleoside in rice seedlings subjected to salt stress and abscisic acid (ABA) treatment. We identified LOC_Os03g61750 (OsTRM13) as a rice candidate methyltransferase for the Am modification. OsTRM13 transcript levels increased significantly upon salt stress and ABA treatment, and the OsTrm13 protein was found to be located primarily to the nucleus. More importantly, OsTRM13 overexpression plants displayed improved salt stress tolerance, and vice versa, OsTRM13 RNA interference (RNAi) plants showed reduced tolerance. Furthermore, OsTRM13 complemented a yeast trm13Δ mutant, deficient in Am synthesis, and the purified OsTrm13 protein catalysed Am nucleoside formation on tRNA-Gly-GCC in vitro. Our results show that OsTRM13, encoding a rice tRNA nucleoside methyltransferase, is an important regulator of salt stress tolerance in rice.

PacMYBA, a sweet cherry R2R3-MYB transcription factor, is a positive regulator of salt stress tolerance and pathogen resistance

Plant R2R3-MYB transcription factors play crucial roles in stress responses. We previously isolated a R2R3-MYB homolog from sweet cherry cv. Hong Deng, designated PacMYBA (GenBank accession No. KF974774). To explore the role of PacMYBA in the plant stress response, we heterologously expressed PacMYBA in transgenic Arabidopsis thaliana plants. In a previous study, we demonstrated that PacMYBA is mainly localized to the nucleus and could be induced by abscisic acid (ABA). Analysis of the promoter sequence of PacMYBA revealed that it contains several stress-related cis-elements. QPCR results showed that PacMYBA is induced by salt, salicylic (SA), and jasmonic acid (JA) in sweet cherry leaves. Transgenic Arabidopsis plants heterologously expressing PacMYBA exhibited enhanced salt-tolerance and increased resistance to Pseudomonas syringe pv. tomato (Pst) DC3000 infection. Overexpression of PacMYBA decreased the osmotic potential (OP), increased the free proline content, and increased the peroxidase content in transgenic Arabidopsis plants. Furthermore, overexpression of PacMYBA also affected the expression levels of salt stress- and pathogen defense-related genes in the transgenic plants. These results indicate that PacMYBA is a positive regulator of salt stress tolerance and pathogen resistance.

The wheat MAP kinase phosphatase 1 alleviates salt stress and increases antioxidant activities in Arabidopsis

Mitogen-activated protein kinase phosphatases (MKPs) are important negative regulators in the MAPK signaling pathways, which play crucial roles in plant growth, development and stress responses. We have previously shown that the heterologous expression of a durum wheat MKP, TMKP1, results in increased tolerance to salt stress in yeast but its particular contribution in salt stress tolerance in plants was not investigated. Here, TMKP1 was overexpressed in Arabidopsis thaliana and physiological changes were assessed in transgenic plants exposed to stress conditions. Under salt stress and especially LiCl, the TMKP1 overexpressors displayed higher germination rates in comparison to wild type plants. The enhancement of salt stress tolerance was accompanied by increased antioxidant enzyme activities, namely superoxide dismutase, catalase and peroxydases. Such increases in antioxidant activities were concomitant with lower malondialdehyde, superoxide anion O2− and hydrogen peroxide levels in the TMKP1 transgenic seedlings. Moreover, we provide evidence that, in contrast to the Arabidopsis ortholog AtMKP1, TMKP1 acts as a positive regulator of salt stress tolerance via its ectopic expression in the Arabidopsis mkp1 mutant.

The nuclear protein GmbZIP110 has transcription activation activity and plays important roles in the response to salinity stress in soybean.

Plant basic-leucine zipper (bZIP) transcription factors play important roles in many biological processes and are involved in the regulation of salt stress tolerance. Previously, our lab generated digital gene expression profiling (DGEP) data to identify differentially expressed genes in a salt-tolerant genotype of Glycine soja (STGoGS) and a salt-sensitive genotype of Glycine max (SSGoGM). This DGEP data revealed that the expression (log2 ratio) of GmbZIP110 was up-regulated 2.76-fold and 3.38-fold in SSGoGM and STGoGS, respectively. In the present study, the salt inducible gene GmbZIP110 was cloned and characterized through phylogenetic analysis, subcellular localization and in silico transcript abundance analysis in different tissues. The functional role of this gene in salt tolerance was studied through transactivation analysis, DNA binding ability, expression in soybean composite seedlings and transgenic Arabidopsis, and the effect of GmbZIP110 on the expression of stress-related genes in transgenic Arabidopsis was investigated. We found that GmbZIP110 could bind to the ACGT motif, impact the expression of many stress-related genes and the accumulation of proline, Na+ and K+, and enhanced the salt tolerance of composite seedlings and transgenic Arabidopsis. Integrating all these results, we propose that GmbZIP110 plays a critical role in the response to salinity stress in soybean and has high potential usefulness in crop improvement.

Genetic regulation of salt stress tolerance revealed by RNA-Seq in cotton diploid wild species, Gossypium davidsonii

Cotton is an economically important crop throughout the world, and is a pioneer crop in salt stress tolerance research. Investigation of the genetic regulation of salinity tolerance will provide information for salt stress-resistant breeding. Here, we employed next-generation RNA-Seq technology to elucidate the salt-tolerant mechanisms in cotton using the diploid cotton species Gossypium davidsonii which has superior stress tolerance. A total of 4744 and 5337 differentially expressed genes (DEGs) were found to be involved in salt stress tolerance in roots and leaves, respectively. Gene function annotation elucidated salt overly sensitive (SOS) and reactive oxygen species (ROS) signaling pathways. Furthermore, we found that photosynthesis pathways and metabolism play important roles in ion homeostasis and oxidation balance. Moreover, our studies revealed that alternative splicing also contributes to salt-stress responses at the posttranscriptional level, implying its functional role in response to salinity stress. This study not only provides a valuable resource for understanding the genetic control of salt stress in cotton, but also lays a substantial foundation for the genetic improvement of crop resistance to salt stress.

An RNA chaperone, AtCSP2, negatively regulates salt stress tolerance

Cold shock domain (CSD) proteins are RNA chaperones that destabilize RNA secondary structures. Arabidopsis Cold Shock Domain Protein 2 (AtCSP2), one of the 4 CSD proteins (AtCSP1-AtCSP4) in Arabidopsis, is induced during cold acclimation but negatively regulates freezing tolerance. Here, we analyzed the function of AtCSP2 in salt stress tolerance. A double mutant, with reduced AtCSP2 and no AtCSP4 expression (atcsp2–3 atcsp4–1), displayed higher survival rates after salt stress. In addition, overexpression of AtCSP2 resulted in reduced salt stress tolerance. These data demonstrate that AtCSP2 acts as a negative regulator of salt stress tolerance in Arabidopsis.