Transgenic Rice Plants Research Articles

OsWRKY114 Negatively Regulates Drought Tolerance by Restricting Stomatal Closure in Rice.

The WRKY family of transcription factors plays a pivotal role in plant responses to biotic and abiotic stress. The WRKY Group III transcription factor OsWRKY114 is a positive regulator of innate immunity against Xanthomonas oryzae pv. oryzae; however, its role in abiotic stress responses is largely unknown. In this study, we showed that the abundant OsWRKY114 transcripts present in transgenic rice plants are reduced under drought conditions. The overexpression of OsWRKY114 significantly increased drought sensitivity in rice, which resulted in a lower survival rate after drought stress. Moreover, we showed that stomatal closure, which is a strategy to save water under drought, is restricted in OsWRKY114-overexpressing plants compared with wild-type plants. The expression levels of PYR/PYL/RCAR genes, such as OsPYL2 and OsPYL10 that confer drought tolerance through stomatal closure, were also markedly lower in the OsWRKY114-overexpressing plants. Taken together, these results suggest that OsWRKY114 negatively regulates plant tolerance of drought stress via inhibition of stomatal closure, which would otherwise prevent water loss in rice.

Isolation and Functional Characterization of a Green-Tissue Promoter in Japonica Rice (Oryza sativa subsp. Japonica).

Simple SummaryTransgenic applications have largely focused on constitutive promoters in plants. However, strong and continuous over-expression of certain genes may be redundant and even harmful to plant growth. Thus, tissue-specific promoters are the most suitable for regulating target gene expression. Although several tissue-specific promoters have been identified, the regulatory mechanism of tissue-specific gene expression remains unclear. By a series of GUS staining of 5′ and 3′ deletions, we uncover tissue-specific cis-acting elements in GSX7R, including ten light-responsive elements. The results reveal that GSX7R is a reverse green tissue-specific promoter, except in endosperm. In contrast, strong tissue-specific promoters that can be used for rice improvements are limited. In this study, we successfully showed that the GSX7R promoter can drive the Cry1Ab gene to resistant rice yellow stem borer. In addition, our study demonstrates an effective promoter to drive foreign genes for crop improvement.Plant promoters play a vital role in the initiation and regulation of gene transcription. In this study, a rice protein/gene of unknown expression, named Os8GSX7, was gained from a rice T-DNA capture line. The semi-quantitative RT-PCR analysis showed that the gene was only expressed in root, glume, and flower, but not in stem, leaf, embryo, and endosperm of japonica rice. The GUS activity analysis of the GSX7R promoter showed that it was a reverse green tissue expression promoter, except in endosperm. The forward promoter of GSX7 cannot normally drive the expression of the foreign GUS gene, while the reverse promoter of GSX7 is a green tissue-specific expression promoter, which can drive the expression of the foreign GUS gene. The region from −2097 to −1543 bp was the key region for controlling the green tissue-specific expression. The regulatory sequences with different lengths from the 2097 bp reverse sequence from the upstream region of the Os8GSX7 were fused with the GUS reporter gene and stably expressed in rice. Furthermore, transgenic rice plants carrying Cry1Ab encoding Bacillus thuringiensis endotoxin, regulated by GSX7R, were resistant to yellow stem borer. The analysis suggested that 10 light responsive elements of tissue-specific expression were found, including ACE, Box4, CAT-box, G-Box, G-box, GATA motif, GC motif, I-box, Sp1, and chs-unit1 M1. In addition, the results of 5′ and 3′ deletions further speculated that ACE and I-box may be the key elements for determining the green tissue-specific expression of GSX7R promoter.

Melatonin Mediates Axillary Bud Outgrowth by Improving Nitrogen Assimilation and Transport in Rice.

Melatonin plays an important role in plant resistance to biotic and abiotic stresses. However, whether melatonin is involved in the regulation of plant architecture, such as the formation of axillary bud outgrowth or tillering, in rice remains unknown. Here, we found that different concentrations of melatonin influenced axillary bud outgrowth in rice, and moderate melatonin concentrations also alleviated the inhibition of axillary bud outgrowth in the presence of high concentrations of basic amino acids lysine and arginine. Furthermore, transcriptome analysis demonstrated that genes involved in nitrogen metabolism and phytohormone signal transduction pathways may affect axillary bud outgrowth, which is regulated by melatonin. We determined that the differentially expressed genes glutamine synthetase OsGS2 and amino acid transporter OsAAP14, which are involved in nitrogen metabolism and are regulated by melatonin and basic amino acids, were the key regulators of axillary bud outgrowth in rice. In addition, we validated the functions of OsGS2 and OsAAP14 using rice transgenic plants with altered axillary bud outgrowth and tillers. Taken together, these results suggest that melatonin mediates axillary bud outgrowth by improving nitrogen assimilation and transport in rice.

Field-Evolved ΔG210-ppo2 from Palmer Amaranth Confers Pre-emergence Tolerance to PPO-Inhibitors in Rice and Arabidopsis.

Resistance to protoporphyrinogen IX oxidase (PPO)-inhibitors in Amaranthus palmeri and Amaranthus tuberculatus is mainly contributed by mutations in the PPO enzyme, which renders herbicide molecules ineffective. The deletion of glycine210 (ΔG210) is the most predominant PPO mutation. ΔG210-ppo2 is overexpressed in rice (Oryza sativa c. ‘Nipponbare’) and Arabidopsis thaliana (Col-0). A foliar assay was conducted on transgenic T1 rice plants with 2× dose of fomesafen (780 g ha−1), showing less injury than the non-transgenic (WT) plants. A soil-based assay conducted with T2 rice seeds confirmed tolerance to fomesafen applied pre-emergence. In agar medium, root growth of WT rice seedlings was inhibited >90% at 5 µM fomesafen, while root growth of T2 seedlings was inhibited by 50% at 45 µM fomesafen. The presence and expression of the transgene were confirmed in the T2 rice survivors of soil-applied fomesafen. A soil-based assay was also conducted with transgenic A. thaliana expressing ΔG210-ppo2 which confirmed tolerance to the pre-emergence application of fomesafen and saflufenacil. The expression of A. palmeri ΔG210-ppo2 successfully conferred tolerance to soil-applied fomesafen in rice and Arabidopsis. This mutant also confers cross-tolerance to saflufenacil in Arabidopsis. This trait could be introduced into high-value crops that lack chemical options for weed management.

Ectopic Overexpression of Pineapple Transcription Factor AcWRKY31 Reduces Drought and Salt Tolerance in Rice and Arabidopsis.

Pineapple (Ananas comosus (L.) Merr.) is an important tropical fruit with high economic value, and its growth and development are affected by the external environment. Drought and salt stresses are common adverse conditions that can affect crop quality and yield. WRKY transcription factors (TFs) have been demonstrated to play critical roles in plant stress response, but the function of pineapple WRKY TFs in drought and salt stress tolerance is largely unknown. In this study, a pineapple AcWRKY31 gene was cloned and characterized. AcWRKY31 is a nucleus-localized protein that has transcriptional activation activity. We observed that the panicle length and seed number of AcWRKY31 overexpression transgenic rice plants were significantly reduced compared with that in wild-type plant ZH11. RNA-seq technology was used to identify the differentially expressed genes (DEGs) between wild-type ZH11 and AcWRKY31 overexpression transgenic rice plants. In addition, ectopic overexpression of AcWRKY31 in rice and Arabidopsis resulted in plant oversensitivity to drought and salt stress. qRT-PCR analysis showed that the expression levels of abiotic stress-responsive genes were significantly decreased in the transgenic plants compared with those in the wild-type plants under drought and salt stress conditions. In summary, these results showed that ectopic overexpression of AcWRKY31 reduced drought and salt tolerance in rice and Arabidopsis and provided a candidate gene for crop variety improvement.

Effects of suppression of chloroplast phosphoglycerate kinase on photosynthesis in rice.

As chloroplast phosphoglycerate kinase (cpPGK) is one of the enzymes which has the highest capacity among the Calvin-Benson cycle enzymes, it has not been regarded as a determinant for photosynthetic capacity. However, it was reported that the rate of CO2 assimilation decreased under high irradiance and normal [CO2] levels in the Arabidopsis cpPGK-knockdown mutant, implying that cpPGK has a control over photosynthetic capacity at a normal [CO2] level. In the present study, the contribution of cpPGK to photosynthetic capacity was evaluated in transgenic rice plants with decreased amounts of cpPGK protein under high irradiance and various [CO2] levels. The gene encoding cpPGK was suppressed using RNA interference techniques. Two lines of transgenic plants, Pi3 and Pi5, in which the amount of cpPGK protein decreased to 21% and 76% of that in wild-type plants, respectively, were obtained. However, there was no substantial difference in the rates of CO2 assimilation between wild-type and transgenic plants. The rates of CO2 assimilation decreased only slightly at elevated [CO2] levels in the transgenic line Pi3 and did not differ between wild-type plants and the transgenic line Pi5, irrespective of [CO2] level. These results clearly indicate that cpPGK does not have a strong control over photosynthetic capacity at various [CO2] levels in rice.

T-LOC: A comprehensive tool to localize and characterize T-DNA integration sites.

Scientists have developed many approaches based on PCR or next-generation sequencing to localize and characterize integrated T-DNAs in transgenic plants generated by Agrobacterium tumefaciens-mediated T-DNA transfer. However, none of these methods has the robust ability to handle all transgenic plants with diversified T-DNA patterns. Utilizing the valuable information in the whole-genome sequencing data of transgenic plants, we have developed a comprehensive approach (T-LOC) to localize and characterize T-DNA integration sites (TISs). We evaluated the performance of T-LOC on genome sequencing data from 48 transgenic rice (Oryza sativa) plants that provide real and unbiased resources of T-DNA integration patterns. T-LOC discovered 75 full TISs and reported a diversified pattern of T-DNA integration: the ideal single-copy T-DNA between two borders, multiple-copy of T-DNAs in tandem or inverted repeats, truncated partial T-DNAs with or without the selection hygromycin gene, the inclusion of T-DNA backbone, the integration at the genome repeat region, and the concatenation of multiple ideal or partial T-DNAs. In addition, we reported that DNA fragments from the two A. tumefaciens plasmids can be fused with T-DNA and integrated into the plant genome. Besides, T-LOC characterizes the genomic changes at TISs, including deletion, duplication, accurate repair, and chromosomal rearrangement. Moreover, we validated the robustness of T-LOC using PCR, Sanger sequencing, and Nanopore sequencing. In summary, T-LOC is a robust approach to studying the TISs independent of the integration pattern and can recover all types of TISs in transgenic plants.

Over-Expression of Dehydroascorbate Reductase Improves Salt Tolerance, Environmental Adaptability and Productivity in Oryza sativa.

Abiotic stress induces reactive oxygen species (ROS) generation in plants, and high ROS levels can cause partial or severe oxidative damage to cellular components that regulate the redox status. Here, we developed salt-tolerant transgenic rice plants that overexpressed the dehydroascorbate reductase gene (OsDHAR1) under the control of a stress-inducible sweet potato promoter (SWPA2). OsDHAR1-expressing transgenic plants exhibited improved environmental adaptability compared to wild-type plants, owing to enhanced ascorbate levels, redox homeostasis, photosynthetic ability, and membrane stability through cross-activation of ascorbate-glutathione cycle enzymes under paddy-field conditions, which enhanced various agronomic traits, including root development, panicle number, spikelet number per panicle, and total grain yield. dhar2-knockdown plants were susceptible to salt stress, and owing to poor seed maturation, exhibited reduced biomass (root growth) and grain yield under paddy field conditions. Microarray revealed that transgenic plants highly expressed genes associated with cell growth, plant growth, leaf senescence, root development, ROS and heavy metal detoxification systems, lipid metabolism, isoflavone and ascorbate recycling, and photosynthesis. We identified the genetic source of functional genomics-based molecular breeding in crop plants and provided new insights into the physiological processes underlying environmental adaptability, which will enable improvement of stress tolerance and crop species productivity in response to climate change.

Molecular Regulation of Antioxidant Melatonin Biosynthesis by Brassinosteroid Acting as an Endogenous Elicitor of Melatonin Induction in Rice Seedlings.

Gibberellic acid (GA) was recently shown to induce melatonin synthesis in rice. Here, we examined whether brassinosteroids (BRs) also induce melatonin synthesis because BRs and GA show redundancy in many functions. Among several plant hormones, exogenous BR treatment induced melatonin synthesis by twofold compared to control treatment, whereas ethylene, 6-benzylaminopurine (BA), and indole-3-acetic acid (IAA) showed negligible effects on melatonin synthesis. Correspondingly, BR treatment also induced a number of melatonin biosynthetic genes in conjunction with the suppression of melatonin catabolic gene expression. Several transgenic rice plants with downregulated BR biosynthesis-related genes, such as DWARF4, DWARF11, and RAV-Like1 (RAVL1), were generated and exhibited decreased melatonin synthesis, indicating that BRs act as endogenous elicitors of melatonin synthesis. Notably, treatment with either GA or BR fully restored melatonin synthesis in the presence of paclobutrazol, a GA biosynthesis inhibitor. Moreover, exogenous BR treatment partially restored melatonin synthesis in both RAVL1 and Gα RNAi transgenic rice plants, whereas GA treatment fully restored melatonin synthesis comparable to wild type in RAVL1 RNAi plants. Taken together, our results highlight a role of BR as an endogenous elicitor of melatonin synthesis in a GA-independent manner in rice plants.

An F-box protein from wheat, TaFBA-2A, negatively regulates JA biosynthesis and confers improved salt tolerance and increased JA responsiveness to transgenic rice plants

Soil salinity is a serious problem encountered by agriculture worldwide, which will lead to many harmful effects on plant growth, development, and even crop yield. F-box protein is the core subunit of the Skp1-Cullin-F-box (SCF) complex E3 ligase and plays crucial roles in regulating the growth, development, biotic & abiotic stresses, as well as hormone signaling pathway in plants. In this study, an FBA type F-box gene TaFBA-2A was isolated from wheat (Triticum aestivum L.). This study showed that TaFBA-2A could interact with TaSKP1, and TaOPR2, the crucial enzyme involving in jasmonic acid (JA) biosynthesis. TaFBA-2A negatively regulates JA biosynthesis, probably by mediating the degradation of TaOPR2 via the ubiquitin-26S proteasome pathway. Ectopic expression of TaFBA-2A improved the salt tolerance and increased the JA responsiveness of the transgenic rice lines. In addition, some agronomic traits closely related to crop yield were significantly enhanced in the rice lines ectopic expressing TaFBA-2A. The data obtained in this study shed light on the function and mechanisms of TaFBA-2A in JA biosynthesis and the responses to salt stress and JA treatment; this study also suggested that TaFBA-2A has the potential in improving the salt tolerance and crop yield of transgenic rice plants.

A novel R3H protein, OsDIP1, confers ABA-mediated adaptation to drought and salinity stress in rice

AimsDehydration responsive element-binding factors (DBFs) have recently emerged as essential components of stress tolerance mechanisms in plants. In this work, we investigated the role of OsDIP1 that operate upstream of DBFs as a regulator of drought and salinity tolerance in rice.MethodsExperiments were conducted on rice plants (Oryza sativa) and included combination of physiological (plant phenotyping) and molecular methods. The latter involved real-time PCR (RT-qPCR); yeast two-hybrid, BiFC assay, GST pull-down, CoIP and firefly luciferase (LUC)/ renilla (REN) assays.ResultsOsDIP1 expression was induced by hydrogen peroxide (H2O2), ABA (abscisic acid), drought (polyethylene glycol treatment), and salt stress. Overexpression of OsDIP1 in rice enhanced drought and salinity tolerance while knocking out OsDIP1 by CRISPR-Cas9 editing resulted in drought and salt sensitive phenotype. The activity and gene expression of antioxidant defense enzymes, superoxide dismutase (SOD), catalase (CAT), was increased in OsDIP1-overexpressed transgenic rice plants, while the content of malondialdehyde (MDA) was decreased. In contrast, the content and gene expression of SODCc2 and CatB, decreased, and the content of MDA was increased in knockout of OsDIP1 rice plants, suggesting that overexpression of OsDIP1 enhances the antioxidant capacity of rice plants. The yeast two-hybrid screening test revealed that OsDIP1 interacted with ZFP36, a key zinc finger transcription factor involved in ABA-induced antioxidant defense. Moreover, OsDIP1 could modulate some key ABA-responsive genes via interacting with ZFP36.ConclusionsIn this work, we show that DIP1 plays a central role in modulating drought and salinity stress tolerance in rice.

Regulation of ppGpp Synthesis and Its Impact on Chloroplast Biogenesis during Early Leaf Development in Rice.

Guanosine tetraphosphate (ppGpp) is known as an alarmone that mediates bacterial stress responses. In plants, ppGpp is synthesized in chloroplasts from GTP and ATP and functions as a regulator of chloroplast gene expression to affect photosynthesis and plant growth. This observation indicates that ppGpp metabolism is closely related to chloroplast function, but the regulation of ppGpp and its role in chloroplast differentiation are not well understood. In rice, ppGpp directly inhibits plastidial guanylate kinase (GKpm), a key enzyme in GTP biosynthesis. GKpm is highly expressed during early leaf development in rice, and the GKpm-deficient mutant, virescent-2 (v2), develops chloroplast-deficient chlorotic leaves under low-temperature conditions. To examine the relationship between GTP synthesis and ppGpp homeostasis, we generated transgenic rice plants over-expressing RSH3, a protein known to act as a ppGpp synthase. When RSH3 was overexpressed in v2, the leaf chlorosis was more severe. Although the RSH3 overexpression in the wild type caused no visible effects, pulse amplitude modulation fluorometer measurements indicated that photosynthetic rates were reduced in this line. This finding implies that the regulation of ppGpp synthesis in rice is involved in the maintenance of the GTP pool required to regulate plastid gene expression during early chloroplast biogenesis. We further investigated changes in the expressions of RelA/SpoT Homolog (RSH) genes encoding ppGpp synthases and hydrolases during the same period. Comparing the expression of these genes with the cellular ppGpp content suggests that the basal ppGpp level is determined by the antagonistic action of multiple RSH enzymatic activities during early leaf development in rice.

Intronic long noncoding RNA, RICE FLOWERING ASSOCIATED (RIFLA), regulates OsMADS56-mediated flowering in rice

Long noncoding RNAs (lncRNAs) are known to play important roles in several plant processes such as flowering, organ development and stress response. However, studies exploring the diversity and complexity of lncRNAs and their mechanism of action in plants are far fewer that those in animals. Here, we show that an intronic lncRNA in rice (Oryza sativa L.), RICE FLOWERING ASSOCIATED (RIFLA), is required for the inhibition of OsMADS56 expression. RIFLA is produced from the first intron of the OsMADS56 gene. Overexpression of RIFLA in rice repressed OsMADS56 expression but activated the expression of flowering inducers Hd3a and RFT1. Additionally, RIFLA-overexpressing transgenic rice plants flowered earlier than the wild type. Under normal conditions, the transcript level of the rice enhancer of zeste gene OsiEZ1, a homolog of Arabidopsis histone H3K27-specific methyltransferase genes SWINGER (SWN) and CURLY LEAF (CLF), was as low as that of RIFLA, whereas the transcript level of OsMADS56 was relatively high. In the osiez1 mutant, OsMADS56 expression was upregulated, whereas RIFLA expression was downregulated. Additionally, RIFLA formed a complex with OsiEZ1. Together, these results suggest that the floral repressor activity of OsMADS56 is epigenetically regulated by RIFLA and OsiEZ1.

Overexpression of OsHAK5 potassium transporter enhances virus resistance in rice (Oryza sativa).

Intracellular potassium (K+) transported by plants under the action of a number of transport proteins is crucial for plant survival under distinct abiotic and biotic stresses. A correlation between K+ status and disease incidence has been found in many studies, but the roles of K+ in regulating disease resistance to viral diseases remain elusive. Here, we report that HIGH‐AFFINITY K+ TRANSPORTER 5 (OsHAK5) regulates the infection of rice grassy stunt virus (RGSV), a negative‐sense single‐stranded bunyavirus, in rice (Oryza sativa). We found the K+ content in rice plants was significantly inhibited on RGSV infection. Meanwhile, a dramatic induction of OsHAK5 transcripts was observed in RGSV‐infected rice plants and in rice plants with K+ deficiency. Genetic analysis indicated that disruption of OsHAK5 facilitated viral pathogenicity. In contrast, overexpression of OsHAK5 enhanced resistance to RGSV infection. Our analysis of reactive oxygen species (ROS) including H2O2 and O2−, by DAB and NBT staining, respectively, indicated that RGSV infection as well as OsHAK5 overexpression increased ROS accumulation in rice leaves. The accumulation of ROS is perhaps involved in the induction of host resistance against RGSV infection in OsHAK5 transgenic overexpression rice plants. Furthermore, RGSV‐encoded P3 induced OsHAK5 promoter activity, suggesting that RGSV P3 is probably an elicitor for the induction of OsHAK5 transcripts during RGSV infection. These findings indicate the crucial role of OsHAK5 in host resistance to virus infection. Our results may be exploited in the future to increase crop yield as well as improve host resistance via genetic manipulations.

New recognition specificity in a plant immune receptor by molecular engineering of its integrated domain

Plant nucleotide-binding and leucine-rich repeat domain proteins (NLRs) are immune sensors that recognize pathogen effectors. Here, we show that molecular engineering of the integrated decoy domain (ID) of an NLR can extend its recognition spectrum to a new effector. We relied for this on detailed knowledge on the recognition of the Magnaporthe oryzae effectors AVR-PikD, AVR-Pia, and AVR1-CO39 by, respectively, the rice NLRs Pikp-1 and RGA5. Both receptors detect their effectors through physical binding to their HMA (Heavy Metal-Associated) IDs. By introducing into RGA5_HMA the AVR-PikD binding residues of Pikp-1_HMA, we create a high-affinity binding surface for this effector. RGA5 variants carrying this engineered binding surface perceive the new ligand, AVR-PikD, and still recognize AVR-Pia and AVR1-CO39 in the model plant N. benthamiana. However, they do not confer extended disease resistance specificity against M. oryzae in transgenic rice plants. Altogether, our study provides a proof of concept for the design of new effector recognition specificities in NLRs through molecular engineering of IDs.

Oryza-Specific Orphan Protein Triggers Enhanced Resistance to Xanthomonas oryzae pv. oryzae in Rice.

All genomes carry lineage-specific orphan genes lacking homology in their closely related species. Identification and functional study of the orphan genes is fundamentally important for understanding lineage-specific adaptations including acquirement of resistance to pathogens. However, most orphan genes are of unknown function due to the difficulties in studying them using helpful comparative genomics. Here, we present a defense-related Oryza-specific orphan gene, Xio1, specifically induced by the bacterial pathogen Xanthomonas oryzae pv. oryzae (Xoo) in an immune receptor XA21-dependent manner. Salicylic acid (SA) and ethephon (ET) also induced its expression, but methyl jasmonic acid (MeJA) reduced its basal expression. C-terminal green fluorescent protein (GFP) tagged Xio1 (Xio1-GFP) was visualized in the nucleus and the cytosol after polyethylene glycol (PEG)-mediated transformation in rice protoplasts and Agrobacterium-mediated infiltration in tobacco leaves. Transgenic rice plants overexpressing Xio1-GFP showed significantly enhanced resistance to Xoo with reduced lesion lengths and bacterial growth, in company with constitutive expression of defense-related genes. However, all of the transgenic plants displayed severe growth retardation and premature death. Reactive oxygen species (ROS) was significantly produced in rice protoplasts constitutively expressing Xio1-GFP. Overexpression of Xio1-GFP in non-Oryza plant species, Arabidopsis thaliana, failed to induce growth retardation and enhanced resistance to Pseudomonas syringae pv. tomato (Pst) DC3000. Our results suggest that the defense-related orphan gene Xio1 plays an important role in distinctive mechanisms evolved within the Oryza and provides a new source of Oryza-specific genes for crop-breeding programs.

Impact of Drought Stress on Grains Filling in Rice and its Management: A Review

Rice (Oryza sativa L.) is the world’s single most important staple food source for over half of the world’s population, whose demand is expected to increase as human population increases almost exponentially every year. There are different kinds of abiotic and biotic stresses that a rice plant endures. Among the different forms of environmental stresses, drought hampers the production of rice grains notably. The reality of climate change is so stark that more and more droughts are expected to be evident in the distant future. Water, being a requisite for rice cultivation, is gradually becoming a highly limited resource across the earth. In comparison to other cereal crops, rice is more sensitive to the soil water content. Decrease of soil water content or the unavailability of water triggers drought stress in rice. Therefore, the management of drought stress for the growth of rice plants is essential as even moderate stress during reproductive growth may result in a drastic reduction of grain yield. The distinct changes brought about by drought stress not only range from morphological to physiological changes but also from biochemical to molecular changes as well. A number of starch synthesis related enzymes like granule bound starch synthase I (GBSSI), soluble starch synthase IIa (SSIIa) play important role in grain filling, which are also affected by limited water under drought environment. Due to water scarcity, the rate of photosynthesis decreases, as a result of which dry matter accumulation decreases, which ultimately affects grain filling in rice. Drought causes delayed flowering as well as spikelet sterility, followed by low grain yield during harvesting. The prevalence of drought over longer periods causes reactive oxygen species to form within the rice plants which subsequently denatures proteins and nucleic acids. In many places where drought continues for longer periods, rice plants die resulting in huge loss of grain yields. The effective strategies that have been adopted involves-the development of stable drought tolerant rice varieties through plant breeding programs, the development of short duration rice cultivars which have short growing cycle and harvesting periods that may help the plant to escape drought, the development of rice plants having deep root system such that it can penetrate the soil to find the available moisture and water required for growth, as well as the development of transgenic drought tolerant rice plants which over express specific genes to withstand drought, are few of the effective management strategies that need special attention. Thus, it can be said that drought management for rice production is a crucial step towards food security and food production in the world. The present discussions will emphasize on the negative impact of drought in rice plants and also discuss possible key management practices that can address the existing and imminent drought related problems on rice plants.

Nitrate-dependent regulation of miR444-OsMADS27 signalling cascade controls root development in rice.

Nitrate is an important nutrient and a key signalling molecule for plant development. A number of transcription factors involved in the response to nitrate and their regulatory mechanisms have been identified. However, little is known about the transcription factors involved in nitrate sensing and their regulatory mechanisms among crop plants. In this study, we identified functions of a nitrate-responsive miR444:MADS-box transcription factor OsMADS27 module and its downstream targets mediating rice root growth and stress responses. Transgenic rice plants expressing miR444 target mimic improved rice root growth. Although miR444 has the potential to target multiple genes, we identified OsMADS27 as the major miR444 target that regulates the expression of nitrate transporters, as well as several key genes including expansins, and those associated with auxin signalling, to promote root growth. In agreement with this, overexpression of miRNA-resistant OsMADS27 improved root development and tolerance to abiotic stresses, while its silencing suppressed root growth. OsMADS27 mediated robust stress tolerance in plants through its ability to bind to the promoters of specific stress regulators, as observed in ChIP-seq analysis. Our results provide evidence of a nitrate-dependent miR444-OsMADS27 signalling cascade involved in the regulation of rice root growth, as well as its surprising role in stress responses.

Meloidogyne-SP4 effector gene silencing reduces reproduction of root-knot nematodes in rice (Oryza sativa)

Summary The root-knot nematodes (RKN) Meloidogyne graminicola and M. incognita are responsible for rice yield losses worldwide, particularly in Asia and Africa. Previous studies demonstrated that nematode-secreted proteins are crucial for root invasion and establishment in the host. We present some characteristics of a pioneer effector, M. incognita-secreted protein 4 (Mi-SP4), which is conserved in RKN and required for infection in compatible rice-RKN interactions. In situ hybridisation assays revealed Mi-SP4 expression in the dorsal pharyngeal gland of M. incognita second-stage juveniles (J2). Meloidogyne-SP4 transcripts strongly accumulated in pre-parasitic J2 and decreased in later parasitic stages of M. incognita and M. graminicola. Transient expression of the nematode effector gene in Nicotiana benthamiana leaves and onion cells indicated that GFP-tagged Mi-SP4 was present in the cytoplasm and accumulated in the nucleus of the plant cells. In vitro RNA interference (RNAi) gene silencing, obtained by soaking J2 with small-interfering (si)RNA si4-1, decreased Mi -SP4 expression in J2 by 35% and significantly reduced M. incognita reproduction in rice by at least 30%. Similarly, host-mediated gene silencing of the nematode SP4 effector candidate gene in transgenic rice plants significantly reduced M. graminicola reproduction by 26% to 47%. The data obtained demonstrate that Mi -SP4 is a pioneer virulence effector, which plays an essential role in both M. incognita and M. graminicola pathogenicity on rice.

OsCyp2‐P, an auxin‐responsive cyclophilin, regulates Ca2+ calmodulin interaction for an ion‐mediated stress response in rice

OsCYP2-P is an active cyclophilin (having peptidyl-prolyl cis/trans-isomerase activity, PPIase) isolated from the wild rice Pokkali having a natural capacity to grow and yield seeds in coastal saline regions of India. Transcript abundance analysis in rice seedlings showed the gene is inducible by multiple stresses, including salinity, drought, high temperature, and heavy metals. To dissect the role of OsCYP2-P gene in stress response, we raised overexpression (OE) and knockdown (KD) transgenic rice plants with >2-3 folds higher and approximately 2-fold lower PPIase activity, respectively. Plants overexpressing this gene had more favorable physiological and biochemical parameters (K+ /Na+ ratio, electrolytic leakage, membrane damage, antioxidant enzymes) than wild type, and the reverse was observed in plants that were knocked down for this gene. We propose that OsCYP2-P contributes to stress tolerance via maintenance of ion homeostasis and thus prevents toxic cellular ion buildup and membrane damage. OE plants were found to have a higher harvest index and higher number of filled grains under salinity and drought stress than wild type. OsCYP2-P interacts with calmodulin, indicating it functions via the Ca-CaM pathway. Compared to the WT, the germinating OE seeds exhibited a substantially higher auxin level, and this hormone was below the detection limits in the WT and KD lines. These observations strongly indicate that OsCyp2-P affects the signaling and transport of auxin in rice.