INTRODUCTION:Achieving both high yield and superior grain quality remains a major challenge in rice breeding due to the long-standing trade-off between these traits. Enhancing vascular transport efficiency may provide a strategy to overcome this constraint, yet the genetic basis linking peduncle vascular architecture with yield-quality coordination remains poorly understood. OBJECTIVE:This study aimed to develop a flow-centered molecular design framework targeting vascular transport capacity to reconcile yield and quality in Oryza sativa. METHODS:Using 248 accessions from the 3 K Rice Genomes panel, 14 traits related to peduncle vascular bundles, yield, and quality were phenotyped, and 31 cloned genes were haplotyped. Haplotype validity was confirmed by functional verification using near-isogenic or transgenic lines. Trait correlations, genetic effects, and pyramiding interactions of key genes were assessed. Superior haplotypes were converted into KASP markers and tested across 221 released cultivars. A breeding strategy was proposed and validated using introgression lines. RESULTS:The peduncle vascular bundles play a crucial role in simultaneously enhancing single-panicle weight and grain appearance quality in japonica/geng rice. Five key genes (GL3.1, GW5, FLO2, LVPA4, and RST1) were identified as synergistic regulators enhancing vascular development, panicle weight, and grain quality without compromising yield. A pyramiding-effect network of genes to guide the simultaneous improvement of yield and quality were constructed. Based on the uneven distribution of superior alleles among modern cultivars, a flow-centered molecular design breeding strategy was subsequently proposed and validated through the development of introgression lines, confirming that optimizing vascular systems can simultaneously improve yield and quality. CONCLUSIONS:This study establishes a flow-centered genetic and conceptual framework linking vascular bundle architecture to yield-quality coordination and provides practical molecular tools for next-generation high-yield, high-quality rice breeding, while also offering a strategic reference for similar improvements in other crops such as wheat and maize.

Pokkah boeng disease (PBD), caused by Fusarium sacchari, is a devastating fungal disease that threatens global sugarcane production. Understanding the genetic basis of PBD resistance is crucial for molecular breeding under sugarcane’s complex autopolyploidy. Here, we evaluated an F1 population of 201 individuals derived from a cross between ‘ROC22’ (resistant) and ‘Zhongzhe 1’ (susceptible). Based on multi-year field trials and artificial inoculations, we established a robust phenotypic evaluation system using Best Linear Unbiased Prediction (BLUP) values. A k-mer-based genome-wide association study (GWAS) identified 759 associated loci, highlighting 22 stable major-effect loci detected across multiple environments. Time-series RNA-seq analysis revealed the transcriptional dynamics of infection, pinpointing 48 h post-inoculation (hpi) as the critical time point for resistance-susceptibility divergence. By integrating physical mapping with temporal expression patterns, we identified four key candidate genes: the sensor DUF4220, the signaling switch Ras-family protein, the defense executioner DDE superfamily endonuclease, and the homeostasis regulator T-complex protein. This study outlines a four-dimensional (perception-transduction-execution-monitoring) immune framework, revealing the molecular mechanisms by which resistant genotypes construct multiple defense barriers through precise temporal coordination. The identified major-effect loci and core genes provide pivotal targets and a scientific foundation for marker-assisted and genomic selection in sugarcane disease-resistance breeding.

Cadmium (Cd) contamination in agricultural soil poses a severe threat to rice growth and food safety worldwide. Seedling-stage Cd tolerance directly determines rice establishment and subsequent yield under Cd stress, but its genetic basis remains largely unclear. In this study, a genome-wide association study (GWAS) was conducted using 490 diverse accessions from the 3000 Rice Genome Project (3K RGP). Three biomass-related traits, shoot height (SH), shoot dry weight (SDW), and root dry weight (RDW), were measured under control and Cd stress conditions, along with their relative values. A total of 3,196,134 high-quality SNPs were used for genetic analysis, and population structure was corrected by principal component analysis (PCA) and kinship matrix. In total, 39 stable QTLs were detected, including 19 for RDW, 18 for SDW, and 2 for SH, most of which were specifically identified under Cd stress. Three major QTLs (qSDW1.1, qRDW3.2, qRDW5.2) were prioritized for candidate gene mining. Combining LD block analysis, gene annotation, Cd-responsive transcriptome data, and haplotype analysis, OsAKR2 (LOC_Os01g62870), OsAS1 (LOC_Os03g18130), and LOC_Os05g11320 were identified as key candidate genes regulating seedling Cd tolerance, and superior haplotypes of these genes were identified. This study reveals the genetic architecture of rice seedling Cd tolerance and provides elite QTLs, genes, and haplotypes for molecular breeding of Cd-resilient rice varieties.

Great advances have been made in rice functional genomics, with thousands of rice genes cloned and characterized. However, few of these genes have been successfully utilized in rice improvement, largely due to the missing link between gene function and the quantitative/population genetic parameters underlying complex traits. To address this challenge, we cloned three related small-effect QTLs-qPHDF11.2, qPHDF12, and qPHDF10-to investigate the genetic control of drought tolerance (DT) in rice using an integrated strategy. This strategy integrates genome-wide association studies (GWAS) with the identification and functional validation/characterization of candidate QTL genes supported by multi-omics analyses and AlphaFold3 structural prediction. Compelling evidence demonstrates that OsGH18, OsMYB2, and OsCAD3 correspond to qPHDF11.2, qPHDF12, and qPHDF10, respectively. OsMYB2 functions as a drought-induced transcription factor that regulates OsGH18 and OsCAD3, both of which are directly involved in lignin biosynthesis. Haplotype analyses identified two and five alleles in the promoter regions of OsMYB2 and OsGH18, respectively, as well as four alleles in the coding region of OsCAD3. Comprehensive genetic analyses revealed strong epistatic interactions among alleles at these three loci, leading to a generalized model in which specific allelic combinations at OsMYB2, OsGH18, and OsCAD3 function as a single regulatory module to modulate lignin biosynthesis and cell-wall lignin, thereby improving DT in rice through tri-genic epistasis. This epistatic model fits well with five DT-related traits measured at the seedling, vegetative, and reproductive stages and shows strong predictive power (∼6%-35%) for field performance across diverse rice accessions from three populations. Moreover, it enables the identification of tri-genic genotypes associated with extreme DT phenotypes as promising donors for DT improvement. Our study implicates that once functional relationships among genes within a complex regulatory network are experimentally established, their epistatic interactions across multiple alleles can be computationally resolved using quantitative genetic models. This strategy will facilitate the development of innovative breeding approaches for improving complex traits by integrating rapidly expanding functional genomics data with advances in AI technologies.

Artificial selection has greatly shaped crop agronomic traits1-3; however, the mechanistic basis of how immunity is selected remains unclear. Here we identify the Oryza sativa nucleotide-binding site and leucine-rich repeat (NLR) receptor XA48 and downstream transcription factors OsVOZ1 and OsVOZ2 (OsVOZ1/2), which confer resistance to bacterial blight. XA48 perceives the ancient pathogen effector XopG, activating effector-triggered immunity by degrading the negative regulator OsVOZ1/2. The XA48-OsVOZ1 module has undergone subspecies-specific selection: Xa48 is retained only in Oryza sativa indica and was lost in Oryza sativa japonica. By contrast, OsVOZ1 has diverged into two haplotypes-O. s. indica retains both OsVOZ1A/S alleles compatible with Xa48, whereas O. s. japonica has only OsVOZ1A. Reintroducing Xa48 into O. s. japonica severely compromises yield owing to the XA48-OsVOZ1A-mediated immune incompatibility. Stacking XA48-mediated effector-triggered immunity with XA21-mediated pattern-triggered immunity reconstitutes the broad-spectrum resistance from wild rice. Our study therefore reveals how asymmetric selection of an NLR-transcription factor module shapes disease resistance and reproductive development, providing a strategy for breeding crops by harnessing the relative immunity of wild rice.

Rice stands as one of the world’s most essential cereal crops, underpinning global food security and economic stability. Salinity-alkalinity stress represents a major environmental constraint that severely impairs rice growth, with mesocotyl elongation being particularly vulnerable. Despite its agronomic importance, the genetic basis of mesocotyl tolerance to combined saline-alkaline stress remains largely unexplored. In this study, we measured mesocotyl length (ML) and relative ML traits under salt-alkali stress and control conditions in a 148-line doubled haploid (DH) population constructed from Sea Rice 86 and Nipponbare (Nip) plants. By constructing a high-density genetic map, we identified two loci significantly associated with mesocotyl elongation under stress, one governing salt tolerance and the other conferring alkali tolerance. Through integrative gene functional annotation and haplotype analysis, we identified two key candidate genes (LOC_Os03g49260 and LOC_Os03g49500) regulating ML, three genes (LOC_Os04g52479, LOC_Os04g52510, and LOC_Os04g52725) linked to salt tolerance, and one pivotal gene (LOC_Os03g01410) associated with alkali tolerance. An effective strategy for enhancing rice ML under salt-alkali stress could be the pyramiding of favorable haplotypes from multiple candidate genes to achieve an optimal haplotype combination. These findings not only provide critical insights into the genetic mechanisms of salinity‒alkalinity tolerance in rice but also provides a functional roadmap for developing resilient rice varieties equipped with superior mesocotyl traits and improved stress adaptability.

In rice (Oryza sativa L.), the vascular bundle tissue in the panicle neck plays a critical role in transporting photosynthetic products, nutrients, and water from the roots and leaves to the panicles, and is positively associated with grain yield. However, the genetic mechanisms regulating vascular bundle number in the panicle neck remain poorly understood. Here, we report the map-based cloning and functional characterization of LVN3, a quantitative trait locus controlling the number of large vascular bundles in rice (Oryza sativa L.) panicle neck. LVN3 encodes OsGA20ox1, an enzyme in gibberellin (GA) biosynthesis. It is expressed in developing panicles and vascular bundles, and positively regulates both the number and cross-sectional area of vascular bundles in the panicle neck and the second internode, as well as flag leaf size and grain number per panicle. Compared with the near-isogenic line NIL-LVN3LT, NIL-LVN3TQ showed significantly increased vascular bundle number and area, enlarged flag leaves, and improved panicle morphology, ultimately leading to a 6.6% increase in grain yield. RNA-seq analysis revealed that LVN3 affects the expression of auxin-related genes in the shoot apex, suggesting that LVN3-mediated vascular development is partly driven by crosstalk between GA and auxin signaling. Cultivars carrying the LVN3-2 haplotype exhibited moderate effective panicle number per plant, grain number, and grain weight, yet achieved the highest grain yield among six haplotypes, identifying LVN3-2 as a superior haplotype for optimizing grain yield components. Moreover, natural variation at position rs3:36150392 in the LVN3 promoter may represent a functional site associated with vascular bundle development and grain number per panicle in the japonica (Geng) subpopulation. Together, these findings demonstrate that LVN3 regulates source capacity, sink size, and transport efficiency, showing potential for application in rice breeding programs aimed at increasing grain yield.

While high-density SNP genotyping enables genome-wide assays in rice (Oryza sativa L.) and other crops, PCR-based markers--particularly those derived from insertion-deletion (InDel) variations-remain crucial for fine-mapping. Currently, readily available primer information for high-density InDel rice markers is still limited. We present the first version of PrimeInDel (https://rfgb.rmbreeding.cn/search/variation/primeIndel), an online tool integrating three sets of InDel markers: an 8K-set and a 316K-set from the 3K Rice Genomes (3K-RG) project, and a 22K-set compiled from published literatures. In user cases, primers from PrimeInDel proved highly effective, narrowing a cold-tolerance QTL (qSR2) from 2.7 Mb to 200 kb and a heading-date QTL (qDeh1.1) from 239.5 kb to 83.6 kb. Our work provides the community with a robust, PCR-based InDel primer, accessible via an online data set, which will facilitate germplasm genotyping and gene identification in rice breeding. ### Competing Interest Statement The authors have declared no competing interest. Biological Breeding-National Science and Technology Major Project, 2022ZD0400404 Bill & Melinda Gates Foundation, OPP1130530

Rice (Oryza sativa L.) leaf rolling enhances canopy architecture and photosynthetic efficiency, thereby improving yield potential. Through genome-wide association studies (GWAS) of 802 Xian and Geng rice accessions, we identified 32 loci associated with the leaf rolling index (LRI) and characterized OUTWARD ROLLED LEAF 4 (ORL4) as a regulatory gene from natural germplasm. CRISPR/Cas9-mediated knockout of ORL4 resulted in abaxially rolled leaves, driven by excessive bulliform cell proliferation and disrupted vascular patterning. A promoter variant (Chr04_19562737-C/T) modulates ORL4 expression by altering the binding affinity of the transcription factor RICE OUTERMOST CELL-SPECIFIC 8 (ROC8). Carriers of the ORL4C allele exhibit stronger ROC8 binding, higher ORL4 expression, and adaxially rolled or flat leaves; ORL4T carriers show the opposite phenotype. ORL4 forms a ternary complex with ROC8 and TOPLESS-RELATED PROTEIN 2(TPL2) to suppress ABAXIALLY CURLED LEAF 1(ACL1), a positive regulator of bulliform cell differentiation. Evolutionary analyses indicated that the ORL4C and ORL4T alleles originated from Oryza nivara and Oryza rufipogon, with ORL4C favored in Geng and ORL4T in Xian during domestication. Promoter editing of ORL4 in the elite cultivar "Shennong 89-366" generated lines with enhanced LRI, achieving a significant yield increase in field trials. This study elucidates the genetic and molecular basis of leaf rolling and provides a promising strategy for rice yield improvement via precise promoter editing.

Rice (Oryza sativa L.), a staple food for more than half of the global population, is moderately salt-sensitive and increasingly threatened by soil salinization. Deciphering the genetic mechanisms of salt tolerance (ST) is pivotal for accelerating genetic improvement of ST by rice molecular breeding and safeguarding global food security. A quantitative trait locus for rice ST, qRLS8, was mapped using a BC2F7 backcross introgression line population derived from the cross between Minghui 63 and 02428. Combined with transcriptome analysis, Lhca4, encoding a subunit of the light-harvesting complex, was identified as the candidate gene of qRLS8. Lhca4 positively regulates rice ST at seedling stage based on the phenotypic verification using knockout and overexpression transgenic lines of Lhca4. Overexpression of Lhca4 increased the activities of superoxide dismutase and peroxidase, and decreased the accumulation of H2O2 and O2−, holding a high photochemical efficiency under salt stress conditions. In contrast, knockout of Lhca4 increased accumulation of reactive oxygen species (ROS) in rice, resulting in disruption of the chloroplast lamellae and a decrease in photosynthetic efficiency. Transcriptome analysis revealed that Lhca4 mediated salt stress response pathway involved in metabolic regulation, enzyme activity regulation, and antioxidant regulation. Lhca4 confers rice ST by preserving chloroplast integrity, maintaining photochemical efficiency, and systemically modulating ROS homeostasis through enhancing antioxidant defense system. These findings provide a valuable gene target for the development of salt-tolerant rice varieties without compromising photosynthetic capacity.

Pentatricopeptide repeat (PPR) proteins are key regulators of organelle RNA metabolism in plants, yet their precise mechanisms in chloroplast RNA processing remain unclear. Here, we identify WPR, a unique P-type PPR protein in rice (Oryza sativa L.), as a critical factor in chloroplast RNA splicing and editing. A ~112-kb chromosomal inversion upstream of WPR causes an albino panicle rachis phenotype (wpr mutant), while complete loss of WPR function leads to seedling lethality. WPR deficiency disrupts the splicing of multiple group II introns (atpF, ndhA, ndhB, petB, rpl2, and rps12) and impairs RNA editing in transcripts such as ndhA, ndhB, ndhG, rps14, and ycf3. Electrophoretic mobility shift assay (EMSA) data confirm that WPR directly binds to precursor mRNAs of atpF, ndhA, petB, rpl2, and rps12. Strikingly, WPR interacts with both RNA editing factors (MORF1, MORF8b) and the splicing factor CAF2, but not with other PPR proteins targeting the same transcripts. Unlike most PPR proteins, WPR contains only six PPR repeats, which is the fewest among all functionally characterized rice PPR proteins. With few informative repeats, WPR likely possesses a broad, low-specificity RNA-binding activity. Moreover, WPR may act on chloroplast RNA maturation by recruiting MORFs and CAF2 rather than other PPR proteins, highlighting a novel regulatory mode in which P-type PPR protein may act as an RNA-binding scaffold to integrate diverse RNA-processing machineries. This study advances the understanding of PPR protein diversity and provides new insights into the molecular mechanisms of chloroplast RNA processing in rice.

Grain number per panicle critically determines rice yield. Although many underlying genes have been reported, yet the molecular mechanisms linking ethylene to panicle development remain unclear. Here, we identify GRAIN NUMBER PER PANICLE 3 (GNP3) as a regulator of GNP through genome-wide association study (GWAS) combined with map-based cloning. GNP3 encodes a MITOGEN-ACTIVATED PROTEIN KINASE KINASE KINASE 22 (OsMKKK22) that phosphorylates S-adenosyl-L-methionine synthetase 1 (SAMS1), triggering its degradation to suppress ethylene biosynthesis. Ethylene overaccumulation in gnp3-1 mutants reduces grain number, while GNP3 overexpression enhances panicle branching and grain yield by lowering ethylene levels. We demonstrate that a natural haplotype GNP3Hap-Tprevalent in indica subspecies strengthens GNP3-SAMS1 interaction, accelerating SAMS1 degradation and improving grain number. Furthermore, overexpressing GNP3 increases grain yield by approximately 20% in field plot conditions. Our findings unveil a MAPK-ethylene regulatory module and highlight GNP3Hap-T as a valuable genetic resource for breeding high-yield rice.

Nitrogen is a crucial element that impacts rice yield and its constituent factors. The effects of reduced nitrogen levels on yield constitute is a complex quantitative trait that is controlled by multiple genes, and its genetic basis requires further exploration. In this study, 562 MAGIC line population and 284 germplasm varieties were used for genome-wide association analysis (GWAS) and haplotype analysis, aiming to detect quantitative trait loci (QTL) and candidate genes associated with tolerance to low nitrogen levels. The ratio of effective panicle number per plant (REPN), total number of grains per panicle (RTGN), seed setting rate (RSSR), thousand grain weight (RTGW), biomass (RBM), harvest index (RHI), and grain yield per plant (RGY) of low to normal nitrogen conditions were measured in this study. The RBM and RHI were directly closely related to RGY, while the RSSR indirectly and positively affected RGY through RHI, and the REPN and RTGN mainly indirectly and positively affected RGY through RBM. LOC_Os06g06440 was the most likely gene affecting low-nitrogen-tolerance-related traits in rice within the region, ranging from 2.898 Mb to 3.046 Mb (148 kb) on chromosome 6, and the haplotype AA, with a significantly larger mean RGY of 0.95 and 1.53 in the MAGIC and germplasm varieties, respectively, was the advanced allele of LOC_Os06g06440. Nine xian (indica) varieties (IRIS_313-11624, IRIS_313-10932, CX382, B067, B249, IRIS_313-8215, IRIS_313-10544, B052, and B233) carrying the superior haplotype (AA) of LOC_Os06g06440 and having a higher RGY were selected for the molecular marker-assisted selection of low nitrogen tolerance in rice. These results will enhance our knowledge of the genetic basis of tolerance to low levels of nitrogen and provide valuable information for improving tolerance to low levels of nitrogen in rice-breeding programs.

BACKGROUND:Rice, being a thermophilic crop, exhibits high sensitivity to low-temperature stress throughout its growth and development. Consequently, enhancing cold tolerance (CT) has been a paramount objective in rice breeding programs. The budding and seedling stages are particularly susceptible to low-temperature damage, making it crucial to improve CT during these stages to ensure the stable establishment and development of the rice population. RESULTS:In this study, we exposed the parental lines Nipponbare (NIP) and Searice 86 (SR86), along with their derived 170 doubled-haploid (DH) population lines, to cold treatments during both the budding and seedling stages. Quantitative trait locus (QTL) mapping was performed using statistical indices such as the survival rate at the budding stage (SRBS), severity of damage at the budding stage (SDBS), survival rate at the seedling stage (SRSS), and wilting degree at the seedling stage (WDSS). This analysis identified four QTLs at the budding stage and eight QTLs at the seedling stage. Furthermore, by integrating differentially expressed genes (DEGs) from transcriptomic data with genes located within the QTL regions, we identified 10 candidate genes for the budding stage and 11 candidate genes for the seedling stage. Based on DNA sequence variations between the parental lines, changes in gene expression under cold treatment, and haplotype analyses, the key candidate genes were ultimately determined to be Os02g0250600 for the budding stage and Os06g0696600 for the seedling stage. Additionally, Gene Ontology (GO) and Kyoto Encyclopedia of Genes and Genomes (KEGG) enrichment analyses of transcriptomic data from both stages revealed significant differences in the regulatory pathways involved in CT between the budding and seedling stages. CONCLUSION:The results indicate that Os02g0250600 is the pivotal gene responsible for CT at the budding stage, with haplotype 4 exhibiting the highest level of CT. Meanwhile, Os06g0696600 plays a crucial role in CT at the seedling stage, where haplotypes 2 and 4 have been identified as advantageous. A comprehensive analysis integrating QTL and transcriptome data from both stages revealed distinct differences in CT mechanisms, highlighting stage-specific variations. This study provides valuable theoretical insights and practical references for the cloning of CT genes and the development of cold-tolerant rice varieties during the budding and seedling stages.

Alkaline soil is characterized by high soluble salt content, elevated pH levels, and ionic imbalance, all of which collectively intensify the harmful effects of alkaline stress on plants. To gain molecular insights into alkaline tolerance (AT), we evaluated 13 AT-related traits in 508 diverse rice accessions from the 3K Rice Germplasm Project at the seedling stage. A total of 2 929 764, 2 059 114, and 1 365 868 single nucleotide polymorphisms were used to identify alkaline-tolerance QTLs via genome-wide association studies (GWAS) in the entire population as well as in the xian and geng subpopulations, respectively. Candidate genes and their superior haplotypes were further identified through gene-based association, haplotype analysis, and gene function annotation. In total, 99 QTLs were identified for AT by GWAS, and three genes (LOC_Os03g49050 for qSSD3.1, LOC_Os05g48760 for qSKC5, and LOC_Os12g01922 for qSNC12) were selected as the most promising candidate genes. Furthermore, we successfully mined superior alleles of key candidate genes from natural variants associated with AT-related traits. This study identified crucial candidate genes and their favorable alleles for AT traits, laying a foundation for further gene cloning and the development of AT rice varieties via marker-assisted selection.

Artificial selection has greatly shaped crop agronomic traits; however, the mechanistic basis of immunity selection has remained elusive. This study identifies a new rice NLR XA48 and its downstream transcription factor OsVOZ1, which confer bacterial blight resistance. XA48 perceives an ancient pathogen effector, XopG, to activates effector-triggered immunity (ETI). The XA48-OsVOZ1 module has undergone subspecies-specific selection. Xa48 is retained in indica but functionally lost in japonica rice. OsVOZ1 has also diverged into two haplotypes, indica kept both OsVOZ1A/S alleles that match XA48; while japonica only inherited OsVOZ1A that greatly decreases yield when Xa48 is reintroduced into japonica, mechanistically explaining the Xa48 loss in japonica. We resurrected wild rice broad-spectrum resistance by stacking XA48-mediated ETI with XA21-mediated pattern-triggered immunity (PTI). Thus, our study reveals that the asymmetric selection of an NLR-TF module shapes both disease resistance and reproduction, and provides a paradigm for breeding crops by harnessing the immunity of wild relatives. ### Competing Interest Statement The authors have declared no competing interest.

Paclobutrazol (PBZ) is extensively used to modulate plant architecture in rice. However, its comprehensive effects on grain yield and aroma in aromatic rice have not been thoroughly investigated. This study used the local aromatic rice cultivars (Meixiangzhan 2 and Xiangyaxiangzhan) as experimental materials to evaluate the impacts of foliar-applied PBZ at three concentrations (0 (CK), 150 (T1), and 300 (T2) mg L−1) on grain yield, photosynthetic characteristics, fragrance formation, and radiation use efficiency (RUE). Field experiments revealed that T1 significantly reduced the leaf area index (LAI) by 10.12% and intercepted photosynthetically active radiation (IPAR) by 10.74%, meanwhile significantly increasing SPAD values by 12.94% and net photosynthetic rate (Pn) by 9.95%, leading to improved RUE up to 25.21%. These changes contributed to a larger number of grains per panicle and increased 1000-grain weight, ultimately enhancing grain yield. In contrast, T2 resulted in a sharp reduction by 24.84% in IPAR and a significant decline in Pn by 10.07% during the late grain-filling stage, thus limiting the supply of photosynthetic assimilates, eventually reducing grain yield. PBZ application also significantly elevated 2-acetyl-1-pyrroline (2-AP) content by 28.74% under T1 and 17.51% under T2, compared to the control. The increase in 2-AP was mainly associated with elevated levels of key precursors, including proline, Δ1-pyrroline-5-carboxylic acid, and Δ1-pyrroline. In spite of differences in traits between cultivars, the traits responded to PBZ in the same pattern. These results indicate that foliar application of PBZ at 150 mg L−1 can effectively improve both yield and aroma of aromatic rice, offering a promising cultivation strategy for high-quality aromatic rice production.

RNA m6A methylation installed by RNA methyltransferases plays a crucial role in regulating plant growth and development and environmental stress responses. However, the underlying molecular mechanisms of m6A methylation involved in seed germination and stress responses are largely unknown. In the present study, we surveyed global m6A methylation in rice seed germination under salt stress and the control (no stress) using an osmta1 mutant and its wild type. The knockout of OsMTA1 resulted in a decreased level of m6A methylation and delayed seed germination, together with increased oxidative damage in the osmta1-1 mutant, especially under salt stress, indicating that OsMTA1 performs a crucial function in rice seed germination and salt stress response. Comparative analysis of m6A profiling using methylated RNA immunoprecipitation sequencing revealed that a unique set of genes that functioned in seed germination, cell growth, and development, including OsbZIP78 and OsA8, were hypomethylated in osmta1-1 embryos and germinating seeds. Numerous genes involved in plant growth and stress response were hypomethylated in the osmta1-1 mutant during seed germination under salt stress. Further combined analysis of the m6A methylome and transcriptome revealed that the loss of function of OsMTA1 had a more complex impact on gene expression in osmta1-1. Several hypomethylated genes with a negative role in growth and development, such as OsHsfA7 and OsHDAC3, were highly up-regulated in the osmta1-1 mutant under the control condition. In contrast, several hypomethylated genes positively associated with stress response were down-regulated, whereas a different set of hypomethylated genes that functioned as negative regulators of growth and stress response were up-regulated in the osmta1-1 mutant under salt stress. These results further demonstrated that OsMTA1-mediated m6A methylation modulated rice seed germination and salt stress response by regulating transcription of a unique set of genes with diverse functions. Our results reveal a crucial role for the m6A methyltransferase gene OsMTA1 in regulating rice seed germination and salt stress response, and provide candidate genes to assist in breeding new stress-tolerant rice varieties.

Soil alkalization is one of the most severe abiotic stresses constraining rice yields. However, the genetic basis underlying alkaline tolerance of rice remains poorly understood. Here, we used genome-wide association analysis to identify OsNPF7.3 as the candidate gene for qAT4 , which is a major locus associated with alkaline tolerance at rice seedling stage. OsNPF7.3 encodes a nitrate/oligopeptide transporter and acts as a negative regulator of rice alkaline tolerance. A natural variation of 7-bp insertion/deletion in the OsNPF7.3 promoter, affecting the binding affinity of transcription factor OsDOF11, mainly contributes to differential transcriptional levels of OsNPF7.3 , and thus leads to differential alkaline tolerance between japonica and indica subspecies. OsNPF7.3 localizes to the vacuolar membrane and mediates nitrogen transport from older to younger leaves under alkaline stress. Loss of OsNPF7.3 significantly upregulated the expression of nitrogen metabolism-related genes and metabolites, suggesting its regulatory role in nitrogen allocation. Together, these findings reveal an OsDOF11- OsNPF7.3 -nitrogen metabolism regulatory module that connects nitrogen homeostasis to alkaline tolerance, providing a promising target for the development of alkaline-tolerant rice varieties.