Search Results (17)

Rice blast disease, caused by Magnaporthe oryzae (M. oryzae), is a significant threat to global rice production. Conventional methods for disease management face limitations, emphasizing the importance of sustainable alternatives. In this study, two rice cultivars with different blast resistant abilities, the susceptible variety CO39 and the resistant variety Pi4b, were used as materials to study the effects of Piriformospora indica (Pi) on the resistance to M. oryzae infection and rice growth. The in vitro tests revealed no direct antagonistic interaction between Pi and M. oryzae. However, the in vivo experiments showed that Pi promoted plant growth by increasing root and shoot length, chlorophyll content, and nitrogen uptake, particularly in CO39 during pathogen infection. Pi inoculation also significantly reduced disease severity, which was indicated by smaller lesion areas and shorter lesion lengths in both cultivars but a more pronounced effect in CO39. This occurred due to the decreasing levels of MDA and the modulating activity of antioxidant enzymes in Pi-inoculated rice plants. At the early stage of M. oryzae infection, the expression of the ethylene signaling gene OsEIN2 and the gibberellin biosynthesis gene OsGA20ox1 in Pi-inoculated CO39 decreased but significantly increased in both rice cultivars at the later stage. The reverse was found for the pathogenesis-related (PR) genes OsPR10 and OsPBZ1 and the blast-resistant genes OsBRG1, OsBRG2, and OsBRW1, suggesting early growth suppression for rice resilience to blast followed by a later shift back to growth. Meanwhile, Pi inoculation increased OsCesA9 expression in rice to strengthen cell walls and establish the primary defense barrier against M. oryzae and upregulated the expression of OsNPR1 without a significant difference in CO39 but downregulated it in Pi4b to activate PR genes to enhance plant resistance. In summary, these results underscore the potential of Pi as a sustainable biological control agent for rice blast disease, which is particularly beneficial for blast-susceptible rice cultivars. Full article

Low temperatures cause serious threat to rice seed emergence, which has become one of the main limiting factors in the production of direct seeding rice. It is of great importance to study the genes controlling low-temperature tolerance during seed germination and to mine the possible regulatory mechanism for developing new rice varieties with immense low-temperature germination ability. In the current research study, two types of mutants of nal11 and gasr9, derived from the WT (wild type) ZH11, were used for the analysis of low-temperature germinability. The results showed that the nal11 and gasr9 mutants displayed no significant difference in germination rate with ZH11 at room temperature, but the mutants showed significantly lower germination rates, germination potential and germination index, and slowed seedling growth in the simulated direct seeding experiments at low temperatures compared to ZH11. Additionally, the activity of POD, SOD, CAT, and anti-superoxide anion radial activity were significantly reduced, but the levels of MDA and H₂O₂ were significantly higher in the nal11 and gasr9 mutant seeds that were germinated at low temperatures compared to ZH11. Further analysis revealed that the levels of total active GA, especially GA4 and GA7, were significantly lower in the nal11 and gasr9 mutants than that in ZH11 during low-temperature germination. Based on qRT-PCR analysis, the expression levels of some GA synthesis-related genes were higher, whereas some were lower in the nal11 and gasr9 mutants than those in ZH11, however, the GA metabolism-related genes OsGA2ox8 and OsGA2ox10 and the GA signaling negative regulator gene SLR1 were significantly up-regulated in both nal11 and gasr9 mutants at several time points during low-temperature germination. This may explain the lower GA levels in the nal11 and gasr9 mutants. Furthermore, the interaction between the OsNAL11 and OsGASR9 proteins was confirmed by Y2H, LUC, and Co-IP assays. This study provides preliminary insights into the regulatory mechanism of the OsNAL11 and OsGASR9 genes, which control the low-temperature germination of rice seeds by affecting the GA pathway. Our study will provide the basis for further mining the molecular mechanisms of low-temperature germination in rice and valuable theoretical reference for breeding varieties with strong low-temperature germinability. Full article

Rice (Oryza sativa L.), a fundamental global staple, nourishes over half of the world’s population. The identification of the ddt1 mutant in rice through EMS mutagenesis of the indica cultivar Shuhui527 revealed a dwarf phenotype, characterized by reduced plant height, smaller grain size, and decreased grain weight. Detailed phenotypic analysis and map-based cloning pinpointed the mutation to a single-base transversion in the LOC_Os03g04680 gene, encoding a cytochrome P450 enzyme, which results in a premature termination of the protein. Functional complementation tests confirmed LOC_Os03g04680 as the DDT1 gene responsible for the observed phenotype. We further demonstrated that the ddt1 mutation leads to significant alterations in gibberellic acid (GA) metabolism and signal transduction, evidenced by the differential expression of key GA-related genes such as OsGA20OX2, OsGA20OX3, and SLR1. The mutant also displayed enhanced drought tolerance, as indicated by higher survival rates, reduced water loss, and rapid stomatal closure under drought conditions. This increased drought resistance was linked to the mutant’s improved antioxidant capacity, with elevated activities of antioxidant enzymes and higher expression levels of related genes. Our findings suggest that DDT1 plays a crucial role in regulating both plant height and drought stress responses. The potential for using gene editing of DDT1 to mitigate the dwarf phenotype while retaining improved drought resistance offers promising avenues for rice improvement. Full article

In model plants, the BRI1-EMS suppressor 1 (BES1)/brassinazole-resistant 1 (BZR1) transcription factors play vital roles in regulating growth, development, and stimuli response. However, the roles of maize ZmBES1/BZR1 members are largely unknown. In this research, the ZmBES1/BZR1-9 gene was ectopically expressed in Arabidopsis and rice for the phenotyping of flowering. We found that the complementation and overexpression of ZmBES1/BZR1-9 in bes1-D mutant and wild type Arabidopsis both resulted in early flowering that was about 10 days shorter than in the untransformed control under long-day conditions. In addition, there was no difference in the rosette leaf number between all transgenic lines and the control. Subsequently, the ZmBES1/BZR1-9 gene was overexpressed in rice. It was found that overexpression lines of rice exhibited early flowering with heading dates that were 8 days shorter compared with untransformed plants. Moreover, the results of RNA-seq and qRT-PCR showed that five flowering-regulated genes, namely At2-MMP, AtPCC1, AtMYB56, AtPELPK1, and AtPRP10, were significantly up-regulated in all complementary and overexpressing lines of Arabidopsis. Meanwhile, the results of RNA-seq showed that 69 and 33 differentially expressed genes (DEGs) were up- and down-regulated in transgenic rice, respectively. Four flowering-related genes, namely OsGA20OX1, OsCCR19, OsBTBN19, and OsRNS4 were significantly up-regulated in transgenic lines. To sum up, our findings demonstrate that ZmBES1/BZR1-9 is involved in controlling flowering and provide insights into further underlying roles of BES1/BZR1s in regulating growth and development in crops. Full article

Arsenic (As) is one of the toxic heavy metal pollutants found in the environment. An excess of As poses serious threats to plants and diminishes their growth and productivity. NAC transcription factors revealed a pivotal role in enhancing crops tolerance to different environmental stresses. The present study investigated, for the first time, the functional role of SNAC3 in boosting As stress tolerance and grain productivity in rice (Oryza sativa L.). Two SNAC3-overexpressing (SNAC3-OX) and two SNAC3-RNAi transgenic lines were created and validated. The wild-type and transgenic rice plants were exposed to different As stress levels (0, 25, and 50 µM). The results revealed that SNAC3 overexpression significantly improved rice tolerance to As stress and boosted grain yield traits. Under both levels of As stress (25 and 50 µM), SNAC3-OX rice lines exhibited significantly lower levels of oxidative stress biomarkers and OsCRY1b (cryptochrome 1b) expression, but they revealed increased levels of gas exchange characters, chlorophyll, osmolytes (soluble sugars, proteins, proline, phenols, and flavonoids), antioxidant enzymes (SOD, CAT, APX, and POD), and stress-tolerant genes expression (OsSOD-Cu/Zn, OsCATA, OsCATB, OsAPX2, OsLEA3, OsDREB2B, OsDREB2A, OsSNAC2, and OsSNAC1) in comparison to wild-type plants. By contrast, SNAC3 suppression (RNAi) reduced grain yield components and reversed the aforementioned measured physio-biochemical and molecular traits. Taken together, this study is the first to demonstrate that SNAC3 plays a vital role in boosting As stress resistance and grain productivity in rice through modulating antioxidants, photosynthesis, osmolyte accumulation, and stress-related genes expression, and may be a useful candidate for further genetic enhancement of stress resistance in many crops. Full article

Clade A protein phosphatase 2Cs (PP2CAs) negatively regulate abscisic acid (ABA) signaling. Here, we investigated the functions of OsPP2CAs and their crosstalk with ABA and gibberellic acid (GA) signaling pathways in rice (Oryza sativa). Among the nine OsPP2CAs, OsPP2C08 had the highest amino acid sequence similarity with OsPP2C51, which positively regulates GA signaling in rice seed germination. However, OsPP2C08 was expressed in different tissues (internodes, sheaths, and flowers) compared to OsPP2C51, which was specifically expressed in seeds, and showed much stronger induction under abiotic stress than OsPP2C51. Transgenic rice lines overexpressing OsPP2C08 (OsPP2C08-OX) had a typical ABA-insensitive phenotype in a post-germination assay, indicating that OsPP2C08, as with other OsPP2CAs, negatively regulates ABA signaling. Furthermore, OsPP2C08-OX lines had longer stems than wild-type (WT) plants due to longer internodes, especially between the second and third nodes. Internode cells were also longer in OsPP2C08-OX lines than in the WT. As GA positively regulates plant growth, these results suggest that OsPP2C08 might positively regulate GA biosynthesis. Indeed, the expression levels of GA biosynthetic genes including gibberellin 20-oxidase (OsGA20ox4) and Ent-kaurenoic acid oxidase (OsKAO) were increased in OsPP2C08-OX lines, and we observed that GIBBERELLIN 2-OXIDASE 4 (OsGA2ox4), encoding an oxidase that catalyzes the 2-beta-hydroxylation of several biologically active GAs, was repressed in the OsPP2C08-OX lines based on a transcriptome deep sequencing and RT-qPCR analysis. Furthermore, we compared the accumulation of SLENDER RICE 1 (SLR1), a DELLA protein involved in GA signaling, in OsPP2C08-OX and WT plants, and observed lower levels of SLR1 in the OsPP2C08-OX lines than in the WT. Taken together, our results reveal that OsPP2C08 negatively regulates ABA signaling and positively regulates GA signaling in rice. Our study provides valuable insight into the molecular mechanisms underlying the crosstalk between GA and ABA signaling in rice. Full article

Seed germination is a key contributing factor to the yield of direct seeding cultivation in rice. Unraveling the genetic architecture underlying rice seed germination is pivotal for breeding elite direct-seeded rice varieties. However, only a limited number of genes regulating seed germination have been characterized in rice. In this study, we implemented a genome-wide association study (GWAS) to dissect the genetic structure of seed germination by using 131 Japonica rice accessions. We identified six stable loci (qGR1.1, qGR2.1, qGR3.1, qGR7.1, qGR8.1 and qGR9) associated with seed germination in two consecutive years, all of which were co-localized with previously reported quantitative trait loci (QTLs). OsGA2ox5, encoding a gibberellin 2-oxidase, was identified as the most plausible candidate gene of the major locus qGR7.1. Knockout of OsGA2ox5 led to delayed seed germination and retarded seedling growth. A non-synonymous variant (Chr7-218,245) within the coding region of OsGA2ox5 might be closely associated with variation in seed germination among Japonica accessions. Low nucleotide diversity at the OsGA2ox5 locus in Japonica could be a result of selection during rice improvement. Taken together, our results provide an important foundation for elucidating the molecular mechanism underlying seed germination and genetic improvement of rice seed vigor in the future. Full article

Gibberellins (GAs), a pivotal plant hormone, play fundamental roles in plant development, growth, and stress response. In rice, gibberellin-dioxygenases (GAoxes) are involved in the biosynthesis and deactivation of gibberellins. However, a comprehensive genome-wide analysis of GA oxidases in rice was not uncovered. Here, a total of 80 candidate OsGAox genes were identified and 19 OsGAox genes were further analyzed. Studies on those 19 OsGAox genes, including phylogenetic tree construction, analysis of gene structure, exploration of conserved motifs and expression patterns, were conducted. Results showed that the GAox genes in Arabidopsis and rice were divided into four subgroups and shared some common features. Analysis of gene structure and conserved motifs revealed that splicing phase and motifs were well conserved during the evolution of GAox genes in Arabidopsis and rice, but some special conserved motifs possessed unknown functions need to be further studied. Exploration of expression profiles from RNA-seq data indicated that each GAox gene had tissue-specific expression patterns, although they varied greatly. The expression patterns of these genes under GA₃ treatment revealed that some genes, such as OsGA2ox1, OsGA2ox3, OsGA2ox4, OsGA2ox7, OsGA20ox1, and OsGA20ox4, may play a major role in regulating the level of bioactive GA. Taken together, our study provides a comprehensive analysis of the GAox gene family and will facilitate further studies on their roles in rice growth and development so that these genes can be better exploited. Full article

The plant hormones gibberellins (GAs) regulate plant growth and development and are closely related to the yield of cash crops. The GA oxidases (GAoxs), including the GA2ox, GA3ox, and GA20ox subfamilies, play pivotal roles in GAs’ biosynthesis and metabolism, but their classification and evolutionary pattern in Gramineae crops remain unclear. We thus conducted a comparative genomic study of GAox genes in six Gramineae representative crops, namely, Setaria italica (Si), Zea mays (Zm), Sorghum bicolor (Sb), Hordeum vulgare (Hv), Brachypodium distachyon (Bd), and Oryza sativa (Os). A total of 105 GAox genes were identified in these six crop genomes, belonging to the C₁₉-GA2ox, C₂₀-GA2ox, GA3ox, and GA20ox subfamilies. Based on orthogroup (OG) analysis, GAox genes were divided into nine OGs and the number of GAox genes in each of the OGs was similar among all tested crops, which indicated that GAox genes may have completed their family differentiations before the species differentiations of the tested species. The motif composition of GAox proteins showed that motifs 1, 2, 4, and 5, forming the 2OG-FeII_Oxy domain, were conserved in all identified GAox protein sequences, while motifs 11, 14, and 15 existed specifically in the GA20ox, C₁₉-GA2ox, and C₂₀-GA2ox protein sequences. Subsequently, the results of gene duplication events suggested that GAox genes mainly expanded in the form of WGD/SD and underwent purification selection and that maize had more GAox genes than other species due to its recent duplication events. The cis-acting elements analysis indicated that GAox genes may respond to growth and development, stress, hormones, and light signals. Moreover, the expression profiles of rice and maize showed that GAox genes were predominantly expressed in the panicles of the above two plants and the expression of several GAox genes was significantly induced by salt or cold stresses. In conclusion, our results provided further insight into GAox genes’ evolutionary differences among six representative Gramineae and highlighted GAox genes that may play a role in abiotic stress. Full article

Salinity reduces agricultural productivity majorly by inhibiting seed germination. Exogenous salicylic acid (SA) can prevent the harm caused to rice by salinity, but the mechanisms by which it promotes rice seed germination under salt stress are unclear. In this study, the inhibition of germination in salt-sensitive Nipponbare under salt stress was greater than that in salt-tolerant Huaidao 5. Treatment with exogenous SA significantly improved germination of Nipponbare, but had little effect on Huaidao 5. The effects of exogenous SA on ion balance, metabolism of reactive oxygen species (ROS), hormone homeostasis, starch hydrolysis, and other physiological processes involved in seed germination of rice under salt stress were investigated. Under salt stress, Na⁺ content and the Na⁺/K⁺ ratio in rice seeds increased sharply. Seeds were subjected to ion pressure, which led to massive accumulation of H₂O₂, O₂⁻, and malonaldehyde (MDA); imbalanced endogenous hormone homeostasis; decreased gibberellic acid (GA1 and GA4) content; increased abscisic acid (ABA) content; inhibition of α-amylase (EC 3.2.1.1) activity; and slowed starch hydrolysis rate, all which eventually led to the inhibition of the germination of rice seeds. Exogenous SA could effectively enhance the expression of OsHKT1;1, OsHKT1;5, OsHKT2;1 and OsSOS1 to reduce the absorption of Na+ by seeds; reduce the Na⁺/K⁺ ratio; improve the activities of SOD, POD, and CAT; reduce the accumulation of H₂O₂, O₂⁻, and MDA; enhance the expression of the GA biosynthetic genes OsGA20ox1 and OsGA3ox2; inhibit the expression of the ABA biosynthetic gene OsNCED5; increase GA1 and GA4 content; reduce ABA content; improve α-amylase activity, and increase the content of soluble sugars. In summary, exogenous SA can alleviate ion toxicity by reducing Na⁺ content, thereby helping to maintain ROS and hormone homeostasis, promote starch hydrolysis, and provide sufficient energy for seed germination, all of which ultimately improves rice seed germination under salt stress. This study presents a feasible means for improving the germination of direct-seeded rice in saline soil. Full article

Gibberellin 2-oxidase (GA2ox) plays an important role in the GA catabolic pathway and the molecular function of the OsGA2ox genes in plant abiotic stress tolerance remains largely unknown. In this study, we functionally characterized the rice gibberellin 2-oxidase 8 (OsGA2ox8) gene. The OsGA2ox8 protein was localized in the nucleus, cell membrane, and cytoplasm, and was induced in response to various abiotic stresses and phytohormones. The overexpression of OsGA2ox8 significantly enhanced the osmotic stress tolerance of transgenic rice plants by increasing the number of osmotic regulators and antioxidants. OsGA2ox8 was differentially expressed in the shoots and roots to cope with osmotic stress. The plants overexpressing OsGA2ox8 showed reduced lengths of shoots and roots at the seedling stage, but no difference in plant height at the heading stage was observed, which may be due to the interaction of OsGA2ox8 and OsGA20ox1, implying a complex feedback regulation between GA biosynthesis and metabolism in rice. Importantly, OsGA2ox8 was able to indirectly regulate several genes associated with the anthocyanin and flavonoid biosynthetic pathway and the jasmonic acid (JA) and abscisic acid (ABA) biosynthetic pathway, and overexpression of OsGA2ox8 activated JA signal transduction by inhibiting the expression of jasmonate ZIM domain-containing proteins. These results provide a basis for a future understanding of the networks and respective phenotypic effects associated with OsGA2ox8. Full article

Semi-dwarfism is a main agronomic trait in crop breeding. In this study, we performed genome-wide association study (GWAS) and identified a new quantitative trait nucleotide (QTN) for rice shoot length. The peak QTN (C/T) was located in the first coding region of a group III WRKY transcription factor OsWRKY21 (LOC_Os01g60640). Interestingly, further haplotype analysis showed that C/T difference only existed in the indica group but not in the japonica group, resulting in significant differences in plant height among the different indica rice varieties. OsWRKY21 was expressed in embryo, radicle, shoots, leaves, and stems. Most notably, overexpressing OsWRKY21 resulted in the semi-dwarf phenotype, early heading date and short internodes compared to the wild type, while the knockout mutant plants by CRISPR/Cas9 technology yielded the opposite. The overexpressing lines exhibited the decreased length of the cells near sclerenchyma epidermis, accompanied with the lower levels of indole-3-acetic acid (IAA) and gibberellin 3 (GA₃), but increased levels of the abscisic acid (ABA) and salicylic acid (SA) in the internodes at heading stage. Moreover, the semi-dwarf phenotype could be fully rescued by exogenous GA₃ application at seedling stage. The RNA-seq and qRT-PCR analysis confirmed the differential expression levels of genes in development and the stress responses in rice, including GA metabolism (GA20ox2, GA2ox6, and YABY1) and cell wall biosynthesis (CesA4, 7, and 9) and regulation (MYB103L). These data suggest the essential role of OsWRKY21 in regulation of internode elongation and plant height in rice. Full article

Ideal Plant Architecture 1 (IPA1) encodes SQUAMOSA PROMOTER BINDING PROTEIN-LIKE 14 (SPL14) with a pleiotropic effect on regulating rice development and biotic stress responses. To investigate the role of IPA1 in early seedling development, we developed a pair of IPA1/ipal-NILs and found that seed germination and early seedling growth were retarded in the ipa1-NIL. Analysis of the soluble sugar content, activity of amylase, and expression of the ?-amylase genes revealed that the starch metabolism was weakened in the ipa1-NIL germinating seeds. Additionally, the content of bioactive gibberellin (GA) was significantly lower than that in the IPA1-NIL seeds at 48 h of imbibition. Meanwhile, the expression of GA synthesis-related gene OsGA20ox1 was downregulated, whereas the expression of GA inactivation-related genes was upregulated in ipa1-NIL seeds. In addition, the expression of OsWRKY51 and OsWRKY71 was significantly upregulated in ipa1-NIL seeds. Using transient dual-luciferase and yeast one-hybrid assays, IPA1 was found to directly activate the expression of OsWRKY51 and OsWRKY71, which would interfere with the binding affinity of GA-induced transcription factor OsGAMYB to inhibit the expression of ?-amylase genes. In summary, our results suggest that IPA1 negatively regulates seed germination and early seedling growth by interfering with starch metabolism via the GA and WRKY pathways. Full article

The semi-dwarfing allele, sd1-d, has been widely utilized in developing high-yielding rice cultivars across the world. Originally identified from the rice cultivar Dee-Geo-Woo-Gen (DGWG), sd1-d, derived from a spontaneous mutation, has a 383-bp deletion in the SD1 gene. To date, as many as seven alleles of the SD1 gene have been identified and used in rice improvement, either with a functional single-nucleotide polymorphism (SNP), with insertion–deletions (InDels), or both. Here, we report discovery of a novel SNP in the SD1 gene from the rice genotype, Pusa 1652. Genetic analysis revealed that the inheritance of the semi-dwarfism in Pusa 1652 is monogenic and recessive, but it did not carry the sd1-d allele. However, response to exogenous gibberellic acid (GA₃) application and the subsequent bulked segregant and linkage analyses confirmed that the SD1 gene is involved in the plant height reduction in Pusa 1652. Sequencing of the SD1 gene from Pusa 1652 revealed a novel transition in exon 3 (T/A) causing a nonsense mutation at the 300th codon. The stop codon leads to premature termination, resulting in a truncated protein of OsGA20ox2 obstructing the GA₃ biosynthesis pathway. This novel recessive allele, named sd1-bm, is derived from Bindli Mutant 34 (BM34), a γ-ray induced mutant of a short-grain aromatic landrace, Bindli. BM34 is the parent of an aromatic semi-dwarf cultivar, Pusa 1176, from which Pusa 1652 is derived. The semi-dwarfing allele, sd1-bm, was further validated by developing a derived cleaved amplified polymorphic sequence (dCAPS) marker, AKS-sd1. This allele provides an alternative to the most widely used sd1-d in rice improvement programs and the functional dCAPS marker will facilitate marker-assisted introgression of the semi-dwarf trait into tall genotypes. Full article

In rice, semi-dwarfism is among the most required characteristics, as it facilitates better yields and offers lodging resistance. Here, semi-dwarf rice lines lacking any residual transgene-DNA and off-target effects were generated through CRISPR/Cas9-guided mutagenesis of the OsGA20ox2 gene in a high yielding Basmati rice line, and the isobaric tags for relative and absolute quantification (iTRAQ) strategy was utilized to elucidate the proteomic changes in mutants. The results indicated the reduced gibberellins (GA₁ and GA₄) levels, plant height (28.72%), and flag leaf length, while all the other traits remained unchanged. The OsGA20ox2 expression was highly suppressed, and the mutants exhibited decreased cell length, width, and restored their plant height by exogenous GA₃ treatment. Comparative proteomics of the wild-type and homozygous mutant line (GXU43_9) showed an altered level of 588 proteins, 273 upregulated and 315 downregulated, respectively. The identified differentially expressed proteins (DEPs) were mainly enriched in the carbon metabolism and fixation, glycolysis/gluconeogenesis, photosynthesis, and oxidative phosphorylation pathways. The proteins (Q6AWY7, Q6AWY2, Q9FRG8, Q6EPP9, Q6AWX8) associated with growth-regulating factors (GRF2, GRF7, GRF9, GRF10, and GRF11) and GA (Q8RZ73, Q9AS97, Q69VG1, Q8LNJ6, Q0JH50, and Q5MQ85) were downregulated, while the abscisic stress-ripening protein 5 (ASR5) and abscisic acid receptor (PYL5) were upregulated in mutant lines. We integrated CRISPR/Cas9 with proteomic screening as the most reliable strategy for rapid assessment of the CRISPR experiments outcomes. Full article