Analysis of Transgenic Cotton Plants Containing Universal Stress Protein (GaUSP-1, GaUSP-2) and Zinc Finger Transcriptional Factor (GaZnF) Genes under Drought Stress.

Water is the most limiting factor for plant growth and crop productivity. Drought stress adversely affects crop yield throughout the world. Up to 50% of crop yield in Pakistan is severely affected by the shortage of water. Cotton is an important cash crop for Pakistan known as "white gold." It accounts for 8.2% of the value added in agriculture and about 3.2% of GDP. Besides, being the world's fourth-largest cotton producer, our yield per acre ranks 13th in the world. If we look at the Pakistan scenario, water deficiency is one of the major yield-limiting factors. Limitations related to conventional breeding and the advancements in plant genomics and biotechnology applications have opened new horizons to plant improvements. Therefore, in the current study, we carry out a comparative analysis to evaluate the morphological, physiological biochemical and molecular parameters in transgenic plants containing GaUSP-1, GaUSP-2 and GaZinc Finger genes under different drought stress conditions. Data showed that transgenic plants showed more tolerance as compared to non-transgenic plants. Transgenic and non-transgenic assist us in our better understanding of the drought-responsive mechanism and its effect on different plant growth traits, so, in this way, we would be able to explore drought tolerance mechanism and this will open the doors for the identification of drought-related genes.

A member of NF-Y family, OsNF-YC5 negatively regulates salt tolerance in rice.

NF-Y, a critical transcription factor, binds to the CCAAT-box in target gene promoters, playing a pivotal role in plant development and abiotic stress response. OsNF-YC5, encodes a putative subunit of the NF-Y transcription factor in rice, had an undetermined function. Our research revealed that OsNF-YC5 is induced by high salinity and exogenous abscisic acid (ABA). Subcellular localization studies showed that OsNF-YC5 is nuclear- and cytoplasm-localized. Using CRISPR-Cas9 to disrupt OsNF-YC5, we observed significantly enhanced rice salinity tolerance and ABA-hypersensitivity. Compared to the wild-type, osnf-yc5 mutants exhibited reduced H2O2 and malondialdehyde (MDA) levels, increased catalase (CAT) activity, and elevated OsCATA transcripts under salt stress. Moreover, ABA-dependent (OsABI2 and OsLEA3) and ABA-independent (OsDREB1A, OsDREB1B, and OsDREB2A) marker genes were upregulated in mutant lines in response to salinity. These results indicate that disrupting OsNF-YC5 enhances rice salinity tolerance, potentially by boosting CAT enzyme activity and modulating gene expression in both ABA-dependent and ABA-independent pathways. Therefore, this study provides a valuable theoretical foundation and genetic resources for developing novel salt-tolerant rice varieties.

The apple lipoxygenase MdLOX3 positively regulates zinc tolerance.

Various abiotic stresses, especially heavy metals near factories around the world, limit plant growth and productivity worldwide. Zinc is a light gray transition metal, and excessive zinc will inactivate enzymes in the soil, weaken the biological function of microorganisms, and enter the food chain through enrichment, thus affecting human health. Lipoxygenase (LOX) can catalyze the production of fatty acid derivatives from phenolic triglycerides in plants and is an important pathway of fatty acid oxidation in plants, which usually begins under unfavorable conditions, especially under biotic and abiotic stresses. Lipoxygenase can be divided into 9-LOX and 13-LOX. MdLOX3 is a homolog of AtLOX3 and has been identified in apples (housefly apples). MdLOX3 has a typical conserved lipoxygenase domain, and promoter analysis shows that it contains multiple stress response elements. In addition, different abiotic stresses and hormonal treatments induced the MdLOX3 response. In order to explore the inherent anti-heavy metal mechanism of MdLOX3, this study verified the properties of MdLOX3 based on genetic analysis and overexpression experiments, including plant taproots length, plant fresh weight, chlorophyll, anthocyanins, MDA, relative electrical conductivity, hydrogen peroxide and superoxide anion, NBT\DAB staining, etc. In the experiment, overexpression of MdLOX3 in apple callus and Arabidopsis effectively enhanced the tolerance to zinc stress by improving the ability to clear ROS. Meanwhile, tomato materials with overexpression of ectopia grew better under excessive zinc ion stress. These results indicated that MdLOX3 had a good tolerance to heavy metal zinc. Homologous mutants are more sensitive to zinc, which proves that MdLOX3 plays an important positive role in zinc stressed apples, which broadens the range of action of LOX3 in different plants.

MdNup62 involved in salt and osmotic stress tolerance in apple.

Abiotic stress of plants has serious consequences on the development of the apple industry. Nuclear pore complexes (NPCs) control nucleoplasmic transport and play an important role in the regulation of plant abiotic stress response. However, the effects of NPCs on apple salt and osmotic stress responses have not been reported yet. In this study, we analyzed the expression and function of NUCLEOPORIN 62 (MdNup62), a component of apple NPC. MdNup62 expression was significantly increased by salt and mannitol (simulated osmotic stress) treatment. The MdNup62-overexpressing (OE) Arabidopsis and tomato lines exhibited significantly reduced salt stress tolerance, and MdNup62-OE Arabidopsis lines exhibited reduced osmotic stress tolerance. We further studied the function of HEAT SHOCK FACTOR A1d (MdHSFA1d), the interacting protein of MdNup62, in salt and osmotic stress tolerance. In contrast to MdNup62, MdHSFA1d-OE Arabidopsis lines showed significantly enhanced tolerance to salt and osmotic stress. Our findings suggest a possible interaction of MdNup62 with MdHSFA1d in the mediation of nuclear and cytoplasmic transport and the regulation of apple salt and osmotic stress tolerance. These results contribute to the understanding of the salt and osmotic stress response mechanism in apple.

Involvement of the Calmodulin-like Protein Gene VaCML92 in Grapevine Abiotic Stress Response and Stilbene Production.

Calmodulin-like proteins (CMLs) are an important family of plant calcium sensor proteins that sense and decode changes in the intracellular calcium concentration in response to environmental and developmental stimuli. Nonetheless, the specific functions of individual CML family members remain largely unknown. This study aims to explore the role of the Vitis amurensis VaCML92 gene in the development of its high stress resistance and the production of stilbenes. The expression of VaCML92 was sharply induced in V. amurensis cuttings after cold stress. The VaCML92 gene was cloned and its role in the abiotic stress responses and stilbene production in grapevine was further investigated. The VaCML92-overexpressing callus cell cultures of V. amurensis and soil-grown plants of Arabidopsis thaliana exhibited enhanced tolerance to cold stress and, to a lesser extent, to the drought, while their tolerance to heat stress and high salinity was not affected. In addition, the overexpression of VaCML92 increased stilbene production in the V. amurensis cell cultures by 7.8-8.7-fold. Taken together, the data indicate that the VaCML92 gene is involved as a strong positive regulator in the rapid response to cold stress, the induction of cold stress resistance and in stilbene production in wild grapevine.

Simultaneous Promotion of Salt Tolerance and Phenolic Acid Biosynthesis in Salvia miltiorrhiza via Overexpression of Arabidopsis MYB12.

Transcription factors play crucial roles in regulating plant abiotic stress responses and physiological metabolic processes, which can be used for plant molecular breeding. In this study, an R2R3-MYB transcription factor gene, AtMYB12, was isolated from Arabidopsis thaliana and introduced into Salvia miltiorrhiza under the regulation of the CaMV35S promoter. The ectopic expression of AtMYB12 resulted in improved salt tolerance in S. miltiorrhiza; transgenic plants showed a more resistant phenotype under high-salinity conditions. Physiological experiments showed that transgenic plants exhibited higher chlorophyll contents, and decreased electrolyte leakage and O2- and H2O2 accumulation when subjected to salt stress. Moreover, the activity of reactive oxygen species (ROS)-scavenging enzymes was enhanced in S. miltiorrhiza via the overexpression of AtMYB12, and transgenic plants showed higher superoxide dismutase (SOD), catalase (CAT), and peroxidase (POD) activities compared with those of the wild type (WT) under salt stress, coupled with lower malondialdehyde (MDA) levels. In addition, the amount of salvianolic acid B was significantly elevated in all AtMYB12 transgenic hair roots and transgenic plants, and qRT-PCR analysis revealed that most genes in the phenolic acid biosynthetic pathway were up-regulated. In conclusion, these results demonstrated that AtMYB12 can significantly improve the resistance of plants to salt stress and promote the biosynthesis of phenolic acids by regulating genes involved in the biosynthetic pathway.

A High-K+ Affinity Transporter (HKT) from Actinidia valvata Is Involved in Salt Tolerance in Kiwifruit.

Ion transport is crucial for salt tolerance in plants. Under salt stress, the high-affinity K+ transporter (HKT) family is mainly responsible for the long-distance transport of salt ions which help to reduce the deleterious effects of high concentrations of ions accumulated within plants. Kiwifruit is well known for its susceptibility to salt stress. Therefore, a current study was designed to decipher the molecular regulatory role of kiwifruit HKT members in the face of salt stress. The transcriptome data from Actinidia valvata revealed that salt stress significantly induced the expression of AvHKT1. A multiple sequence alignment analysis indicated that the AvHKT1 protein contains three conserved amino acid sites for the HKT family. According to subcellular localization analysis, the protein was primarily present in the cell membrane and nucleus. Additionally, we tested the AvHKT1 overexpression in 'Hongyang' kiwifruit, and the results showed that the transgenic lines exhibited less leaf damage and improved plant growth compared to the control plants. The transgenic lines displayed significantly higher SPAD and Fv/Fm values than the control plants. The MDA contents of transgenic lines were also lower than that of the control plants. Furthermore, the transgenic lines accumulated lower Na+ and K+ contents, proving this protein involvement in the transport of Na+ and K+ and classification as a type II HKT transporter. Further research showed that the peroxidase (POD) activity in the transgenic lines was significantly higher, indicating that the salt-induced overexpression of AvHKT1 also scavenged POD. The promoter of AvHKT1 contained phytohormone and abiotic stress-responsive cis-elements. In a nutshell, AvHKT1 improved kiwifruit tolerance to salinity by facilitating ion transport under salt stress conditions.

Chloroplast transformation for bioencapsulation and oral delivery using the immunoglobulin G fragment crystallizable (Fc) domain.

Proinsulin Like Growth Factor I (prolGF-I) and myostatin (Mstn) regulate muscle regeneration and mass when intravenously delivered. We tested if chloroplast bioencapsulated forms of these proteins may serve as a non-invasive means of drug delivery through the digestive system. We created tobacco (Nicotiana tabacum) plants carrying GFP-Fc1, proIGF-I-Fc1, and Mstn-Fc1 fusion genes, in which fusion with the immunoglobulin G Fc domain improved both protein stability and absorption in the small intestine. No transplastomic plants were obtained with the Mstn-Fc1 gene, suggesting that the protein is toxic to plant cells. proIGF-I-Fc1 protein levels were too low to enable in vivo testing. However, GFP-Fc1 accumulated at a high level, enabling evaluation of chloroplast-made Fc fusion proteins for oral delivery. Tobacco leaves were lyophilized for testing in a mouse system. We report that the orally administered GFP-Fc1 fusion protein (5.45 µg/g GFP-Fc1) has been taken up by the intestinal epithelium cells, evidenced by confocal microscopy. GFP-Fc1 subsequently entered the circulation where it was detected by ELISA. Data reported here confirm that chloroplast expression and oral administration of lyophilized leaves is a potential delivery system of therapeutic proteins fused with Fc1, with the advantage that the proteins may be stored at room temperature.

The Arabidopsis ubiquitin ligases ATL31 and ATL6 regulate plant response to salt stress in an ABA-independent manner.

E3 ubiquitin ligases play critical roles in regulating plant response to salt stress. Arabidopsis Tóxicos En Levadura (ATL) is a subfamily of RING-type E3 ubiquitin ligases widely conserved in plant species. ATL genes have been shown to be involved in regulating plant response to biotic or abiotic stresses. We previously found that a pair of ATL genes, ATL31 and ATL6 positively regulated plant innate immunity. However, whether ATL31/6 are also involved in salt stress response remains to be investigated. Here, we demonstrate that ATL31/6 are induced by salt stress. The atl31 atl6 double mutant exhibits increased salt tolerance compared to the wild-type plants, while transgenic plants overexpressing ATL31 are more salt-sensitive. Notably, ATL31 and ATL6 do not participate in the abscisic acid (ABA) response. Furthermore, NaCl treatment induces the proteasomal degradation of ATL31 proteins. Together, we demonstrate that ATL31/6 positively regulate plant tolerance to salt stress, which is independent of ABA, and our work reveals that ATL31/6 are involved in regulating plant response to both biotic and abiotic stress.

Ectopic Expression of MADS-Box Transcription Factor VvAGL12 from Grape Promotes Early Flowering, Plant Growth, and Production by Regulating Cell-Wall Architecture in Arabidopsis.

The MADS-box family, a substantial group of plant transcription factors, crucially regulates plant growth and development. Although the functions of AGL12-like subgroups have been elucidated in Arabidopsis, rice, and walnut, their roles in grapes remain unexplored. In this study, we isolated VvAGL12, a member of the grape MADS-box group, and investigated its impact on plant growth and biomass production. VvAGL12 was found to localize in the nucleus and exhibit expression in both vegetative and reproductive organs. We introduced VvAGL12 into Arabidopsis thaliana ecotype Columbia-0 and an agl12 mutant. The resulting phenotypes in the agl12 mutant, complementary line, and overexpressed line underscored VvAGL12's ability to promote early flowering, augment plant growth, and enhance production. This was evident from the improved fresh weight, root length, plant height, and seed production, as well as the reduced flowering time. Subsequent transcriptome analysis revealed significant alterations in the expression of genes associated with cell-wall modification and flowering in the transgenic plants. In summary, the findings highlight VvAGL12's pivotal role in the regulation of flowering timing, overall plant growth, and development. This study offers valuable insights, serving as a reference for understanding the influence of the VvAGL12 gene in other plant species and addressing yield-related challenges.

Membrane-Bound Transcription Factor ZmNAC074 Positively Regulates Abiotic Stress Tolerance in Transgenic Arabidopsis.

Maize (Zea mays L.) is one of the most widely cultivated crops for humans, making a vital contribution to human nutrition and health. However, in recent years, due to the influence of external adverse environments, the yield and quality of maize have been seriously affected. NAC (NAM, ATAF1/2 and CUC2) transcription factors (TFs) are important plant-unique TFs, which are crucial for regulating the abiotic stress response of plants. Therefore, it is of great biological significance to explore the underlying regulatory function of plant NAC TFs under various abiotic stresses. In this study, wild-type and ZmNAC074-overexpressed transgenic Arabidopsis were used as experimental materials to dissect the stress-resistant function of ZmNAC074 in transgenic Arabidopsis at phenotypic, physiological and molecular levels. The analyses of seed germination rate, survival rate, phenotype, the content of chlorophyll, carotenoids, malondialdehyde (MDA), proline and other physiological indexes induced by distinct abiotic stress conditions showed that overexpression of ZmNAC074 could confer the enhanced resistance of salt, drought, and endoplasmic reticulum (ER) stress in transgenic Arabidopsis, indicating that ZmNAC074 plays an important regulatory role in plant response to abiotic stress, which provides an important theoretical foundation for further uncovering the molecular regulation mechanism of ZmNAC074 under abiotic stresses.

WRKY Transcriptional Factor IlWRKY70 from Iris laevigata Enhances Drought and Salinity Tolerances in Nicotiana tabacum.

Drought and high salinity greatly affect plant growth and development. WRKY transcription factors play a key role in plant tolerance to abiotic stress, but the functions of WRKYs in the ornamental monocotyledon Iris laevigata remain largely unexplored. In this study, we cloned IlWRKY70 and found that it is a Group III WRKY localized in the nucleus. The expression of IlWRKY70 was induced by NaCl and PEG-6000, which reached peaks (4.38 and 5.65 times) after 3 h and 1 h, respectively. The exogenous overexpression of IlWRKY70 in N. tabacum significantly improved the resistance under NaCl and drought treatments, as evidenced by higher germination rates, longer root lengths, and increased fresh weights compared to those of control plants. In addition, transgenic seedlings showed significantly reduced wilting, higher photosynthetic performance, higher Fv/Fm and chlorophyll content, and lower stomatal conductance. Moreover, transgenic lines showed higher antioxidant enzymatic activities, lower reactive oxygen species (ROS), and lower malondialdehyde contents. Accordingly, we also found higher expressions of antioxidant defense genes, including SOD, CAT, and POD, in transgenic lines compared to controls under salt and drought stresses. Thus, IlWRKY70 enhances the abilities of salt and drought tolerances in plants, at least partially, via ROS regulation and can be used for breeding I. laevigata possessing enhanced salt and drought resistances.

Overexpression of Grapevine VyTRXy Improves Drought Tolerance by Maintaining Photosynthesis and Enhancing the Antioxidant and Osmolyte Capacity of Plants.

Drought stress profoundly affects plant growth and development, posing a significant challenge that is extensively researched in the field. Thioredoxins (TRXs), small proteins central to redox processes, are crucial to managing both abiotic and biotic stresses. In this research, the VyTRXy gene, cloned from wild Yanshan grapes, was validated as a functional TRX through enzyme activity assays. VyTRXy was found to bolster photosynthesis, augment levels of osmotic regulators, stimulate antioxidant enzyme activities, and strengthen drought resilience in transgenic plants. These enhancements were evidenced by higher survival rates, optimized photosynthetic metrics, increased proline levels, augmented chlorophyll concentration, reduced electrolyte leakage, and decreased malondialdehyde and hydrogen peroxide (H2O2) levels. Furthermore, there was a surge in the activities of enzymes such as catalase, ascorbate peroxidase, glutathione peroxidase, dehydroascorbate reductase, and glutathione reductase, along with an increased expression of TRX peroxidase. Notably, under drought stress, there was a marked elevation in the expression of stress-responsive genes, including the adversity stress-inducible expression gene (NtRD29A) and DRE-binding protein (NtDREB), in transgenic tobacco. This investigation is pivotal in the quest for drought-resistant grapevine varieties and provides significant insights into the molecular functionality of VyTRXy in enhancing grapevine drought tolerance.

Wheat WRKY transcription factor TaWRKY24 confers drought and salt tolerance in transgenic plants.

Drought and salt stress are major environmental conditions that severely limit plant growth and productivity. WRKY transcription factors play a vital role in the responses against biotic or abiotic stress. In this study, TaWRKY24, a gene of the IIe WRKY family identified in wheat, was cloned and characterized. TaWRKY24 was mainly expressed in wheat leaf and stem and induced by treatment with PEG6000, salt, H2O2, ABA, MeJA, and ethrel. TaWRKY24 transient expression in onion epidermal cells suggested its nuclear localization and its transcriptional activation capability characteristics. Overexpression of TaWRKY24 in tobacco improved the seed germination rate and root growth of seedlings in transgenic lines when subjected to higher mannitol and NaCl concentrations. Further research showed that transgenic lines had higher proline and soluble sugars and lower levels of reactive oxygen species (ROS) and malondialdehyde (MDA). Moreover, compared to normal and negative control plants, TaWRKY24 silenced wheat seedlings had reduced growth under salt and drought stress. This study shows that wheat TaWRKY24 is crucial to plant stress, providing an excellent candidate gene for wheat resistance breeding.

CaRH57, a RNA helicase, contributes pepper tolerance to heat stress.

RNA helicases (RHs) are required for most aspects of RNA metabolism and play an important role in plant stress tolerance. Heat stress (HS) causes the deleterious effects on plant cells, such as membrane disruption and protein misfolding, which results in the inhibition of plant growth and development. In this study, CaRH57 was identified from pepper (Capsicum annuum) and encodes a DEAD-box RH. CaRH57 was induced by HS, and overexpression of CaRH57 in Atrh57-1 rescued the glucose-sensitive phenotype of Atrh57-1, suggesting the functional replacement of CaRH57 to AtRH57. The nucleolus-localized CaRH57 possessed a RH activity in vitro. CaRH57 knockdown impaired pepper heat tolerance, showing severe necrosis and enhanced ROS accumulation in the region of the shoot tip. Additionally, accumulation of aberrant-spliced CaHSFA1d and CaHSFA9d was enhanced, and the corresponding mature mRNA levels were reduced in the TRV2 (Tobacco rattle virus)-CaRH57-infected plants compared with the control plants under HS. Overall, these results suggested that CaRH57 acted as a RH to confer pepper heat tolerance and was required for the proper pre-mRNA splicing of some HS-related genes.

Iron and zinc biofortification of rice by synergistic expression of OsNAS2 gene with monocot (Pennisetum glaucum) and dicot (Phaseolus vulgaris) ferritins.

Iron and zinc deficiencies are the most prevalent cause of global hidden hunger. Rice, being one of the most consumed crops worldwide, is suitable to target for Fe and Zn biofortification. In present study, we generated rice transgenic lines to meet the recommended dietary requirement of iron and zinc through endosperm specific expression of dicot (kidney bean) and monocot (pearl millet) Ferritins along with constitutive expression of rice nicotianamine synthase 2 (OsNAS2) gene. Visualization through perls' prussian staining and quantification by ICP-MS showed significant improvement in grain iron content in all the transgenic lines. The transgenic lines expressing any of the three selected gene combinations (PvFerrtin-OsNAS2, feedPgFerrtin-OsNAS2 and foodPgFerritin-OsNAS2), showed the potential to surpass the 30% of the estimated average requirement (13 µg/g Fe and 28 µg/g Zn) proposed for rice in HarvestPlus breeding program. Though the expression of PvFerritin along with OsNAS2 gene in IET10364 (indica) variety showed the best result, providing up to 4.2- and 3.5-fold increase in iron (30.56 µg/g) and zinc (60.1 µg/g) content, respectively; in polished grains compared to non-transgenic control. Thus, the lines developed in our study can be used for further breeding purpose to enhance the iron and zinc content in commercial rice varieties.

The Transcription Factor ZmNAC89 Gene Is Involved in Salt Tolerance in Maize (Zea mays L.).

The NAC gene family has transcription factors specific to plants, which are involved in development and stress response and adaptation. In this study, ZmNAC89, an NAC gene in maize that plays a role in saline-alkaline tolerance, was isolated and characterized. ZmNAC89 was localized in the nucleus and had transcriptional activation activity during in vitro experiments. The expression of ZmNAC89 was strongly upregulated under saline-alkaline, drought and ABA treatments. Overexpression of the ZmNAC89 gene in transgenic Arabidopsis and maize enhanced salt tolerance at the seedling stage. Differentially expressed genes (DEGs) were then confirmed via RNA-sequencing analysis with the transgenic maize line. GO analyses showed that oxidation-reduction process-regulated genes were involved in ZmNAC89-mediated salt-alkaline stress. ZmNAC89 may regulate maize saline-alkali tolerance through the REDOX pathway and ABA signal transduction pathway. From 140 inbred maize lines, 20 haplotypes and 16 SNPs were found in the coding region of the ZmNAC89 gene, including the excellent haplotype HAP20. These results contribute to a better understanding of the response mechanism of maize to salt-alkali stress and marker-assisted selection during maize breeding.

Genome-Wide Analysis of Glycerol-3-Phosphate Acyltransferase (GPAT) Family in Perilla frutescens and Functional Characterization of PfGPAT9 Crucial for Biosynthesis of Storage Oils Rich in High-Value Lipids.

Glycerol-3-phosphate acyltransferase (GPAT) catalyzes the first step in triacylglycerol (TAG) biosynthesis. However, GPAT members and their functions remain poorly understood in Perilla frutescens, a special edible-medicinal plant with its seed oil rich in polyunsaturated fatty acids (mostly α-linolenic acid, ALA). Here, 14 PfGPATs were identified from the P. frutescens genome and classified into three distinct groups according to their phylogenetic relationships. These 14 PfGPAT genes were distributed unevenly across 11 chromosomes. PfGPAT members within the same subfamily had highly conserved gene structures and four signature functional domains, despite considerable variations detected in these conserved motifs between groups. RNA-seq and RT-qPCR combined with dynamic analysis of oil and FA profiles during seed development indicated that PfGPAT9 may play a crucial role in the biosynthesis and accumulation of seed oil and PUFAs. Ex vivo enzymatic assay using the yeast expression system evidenced that PfGPAT9 had a strong GPAT enzyme activity crucial for TAG assembly and also a high substrate preference for oleic acid (OA, C18:1) and ALA (C18:3). Heterogeneous expression of PfGPAT9 significantly increased total oil and UFA (mostly C18:1 and C18:3) levels in both the seeds and leaves of the transgenic tobacco plants. Moreover, these transgenic tobacco lines exhibited no significant negative effect on other agronomic traits, including plant growth and seed germination rate, as well as other morphological and developmental properties. Collectively, our findings provide important insights into understanding PfGPAT functions, demonstrating that PfGPAT9 is the desirable target in metabolic engineering for increasing storage oil enriched with valuable FA profiles in oilseed crops.

Overexpression of Lolium multiflorum LmMYB1 Enhances Drought Tolerance in Transgenic Arabidopsis.

Lolium multiflorum is one of the world-famous forage grasses with rich biomass, fast growth rate and good nutritional quality. However, its growth and forage yield are often affected by drought, which is a major natural disaster all over the world. MYB transcription factors have some specific roles in response to drought stress, such as regulation of stomatal development and density, control of cell wall and root development. However, the biological function of MYB in L. multiflorum remains unclear. Previously, we elucidated the role of LmMYB1 in enhancing osmotic stress resistance in Saccharomyces cerevisiae. Here, this study elucidates the biological function of LmMYB1 in enhancing plant drought tolerance through an ABA-dependent pathway involving the regulation of cell wall development and stomatal density. After drought stress and ABA stress, the expression of LmMYB1 in L. multiflorum was significantly increased. Overexpression of LmMYB1 increased the survival rate of Arabidopsis thaliana under drought stress. Under drought conditions, expression levels of drought-responsive genes such as AtRD22, AtRAB and AtAREB were up-regulated in OE compared with those in WT. Further observation showed that the stomatal density of OE was reduced, which was associated with the up-regulated expression of cell wall-related pathway genes in the RNA-Seq results. In conclusion, this study confirmed the biological function of LmMYB1 in improving drought tolerance by mediating cell wall development through the ABA-dependent pathway and thereby affecting stomatal density.

A Novel Isoprene Synthase from the Monocot Tree Copernicia prunifera (Arecaceae) Confers Enhanced Drought Tolerance in Transgenic Arabidopsis.

The capacity to emit isoprene, among other stresses, protects plants from drought, but the molecular mechanisms underlying this trait are only partly understood. The Arecaceae (palms) constitute a very interesting model system to test the involvement of isoprene in enhancing drought tolerance, as their high isoprene emissions may have contributed to make them hyperdominant in neotropical dry forests, characterized by recurrent and extended periods of drought stress. In this study we isolated and functionally characterized a novel isoprene synthase, the gene responsible for isoprene biosynthesis, from Copernicia prunifera, a palm from seasonally dry tropical forests. When overexpressed in the non-emitter Arabidopsis thaliana, CprISPS conferred significant levels of isoprene emission, together with enhanced tolerance to water limitation throughout plant growth and development, from germination to maturity. CprISPS overexpressors displayed higher germination, cotyledon/leaf greening, water usage efficiency, and survival than WT Arabidopsis under various types of water limitation. This increased drought tolerance was accompanied by a marked transcriptional up-regulation of both ABA-dependent and ABA-independent key drought response genes. Taken together, these results demonstrate the capacity of CprISPS to enhance drought tolerance in Arabidopsis and suggest that isoprene emission could have evolved in Arecaceae as an adaptive mechanism against drought.