The transcribed centromeric gene OsMRPL15 is essential for pollen development in rice.

Centromeres consist of highly repetitive sequences that are challenging to map, clone, and sequence. Active genes exist in centromeric regions, but their biological functions are difficult to explore owing to extreme suppression of recombination in these regions. In this study, we used the CRISPR/Cas9 system to knock out the transcribed gene Mitochondrial Ribosomal Protein L15 (OsMRPL15), located in the centromeric region of rice (Oryza sativa) chromosome 8, resulting in gametophyte sterility. Osmrpl15 pollen was completely sterile, with abnormalities appearing at the tricellular stage including the absence of starch granules and disrupted mitochondrial structure. Loss of OsMRPL15 caused abnormal accumulation of mitoribosomal proteins and large subunit rRNA in pollen mitochondria. Moreover, the biosynthesis of several proteins in mitochondria was defective, and expression of mitochondrial genes was upregulated at the mRNA level. Osmrpl15 pollen contained smaller amounts of intermediates related to starch metabolism than wild-type pollen, while biosynthesis of several amino acids was upregulated, possibly to compensate for defective mitochondrial protein biosynthesis and initiate consumption of carbohydrates necessary for starch biosynthesis. These results provide further insight into how defects in mitoribosome development cause gametophyte male sterility.

MUS81 is required for atypical recombination intermediate resolution but not crossover designation in rice.

The endonuclease methyl methanesulfonate and UV-sensitive protein 81 (MUS81) has been reported to participate in DNA repair during mitosis and meiosis. However, the exact meiotic function of MUS81 in rice remains unclear. Here, we use a combination of physiological, cytological, and genetic approaches to provide evidence that MUS81 functions in atypical recombination intermediate resolution rather than crossover designation in rice. Cytological and genetic analysis revealed that the total chiasma numbers in mus81 mutants were indistinguishable from wild-type. The numbers of HEI10 foci (the sites of interference-sensitive crossovers) in mus81 were also similar to that of wild-type. Moreover, disruption of MUS81 in msh5 or msh4 msh5 background did not further decrease chiasmata frequency, suggesting that rice MUS81 did not function in crossover designation. Mutation of FANCM and ZEP1 could enhance recombination frequency. Unexpectedly, chromosome fragments and bridges were frequently observed in mus81 zep1 and mus81 fancm, illustrating that MUS81 may resolve atypical recombination intermediates. Taken together, our data suggest that MUS81 contributes little to crossover designation but plays a crucial role in the resolution of atypical meiotic intermediates by working together with other anti-crossover factors.

OsRAD51 Plays a Vital Role in Promoting Homologous Recombination in Rice Meiosis.

Meiotic recombination plays a pivotal role in achieving accurate chromosomal segregation and increasing genetic diversity. In the homologous recombination pathway, the detailed mechanisms of how OsRAD51 and OsDMC1 work in rice meiosis remain to be explored. Here, we obtained different types of mutants for Osrad51a1, Osrad51a2, Osdmc1a, and Osdmc1b through CRISPR/Cas9. Both Osrad51a1 and Osrad51a2 exhibited normal vegetative growth and fertility. Osrad51 (Osrad51a1 Osrad51a2) mutant plants show normal vegetative growth but exhibit complete sterility, indicating that OsRAD51A1 and OsRAD51A2 are functionally redundant in rice fertility. In contrast to the wild type, Osrad51 chromosomes are not paired perfectly at pachytene and synaptonemal complex (SC) formation is deficient. Moreover, univalents and multivalent associations were observed at metaphase I, chromosome fragments presented at anaphase I, and crossover formation is basically suppressed in Osrad51 pollen mother cells (PMCs). OsRAD51 foci emerge at leptotene and disappear from late pachytene and chromosome localization of OsRAD51 depends on the formation of double-strand breaks (DSBs). Most OsRAD51 foci can co-localize with OsDMC1 signals. OsRAD51 is essential for the loading of OsDMC1 onto chromosomes, and vice versa. In addition, both OsRAD51 and OsDMC1 can interact with OsFIGL1 and OsBRCA2, two important components in rice meiosis. Moreover, the Osrad51 Osdmc1 (Osrad51a1 Osrad51a2 Osdmc1a Osdmc1b) quadruple mutant PMCs exhibited similar defective phenotypes as Osrad51 in homologous pairing, synapsis, and DSB repair. Taken together, our results suggest that the recombinases DMC1 and RAD51 may functionally depend on each other and play important roles in meiotic recombination during meiosis in rice.

FIGNL1 Inhibits Non-homologous Chromosome Association and Crossover Formation.

Meiotic crossovers (COs) not only generate genetic diversity but also ensure the accuracy of homologous chromosome segregation. Here, we identified FIGNL1 as a new inhibitor for extra crossover formation in rice. The fignl1 mutant displays abnormal interactions between non-homologous chromosomes at diakinesis, and chromosome bridges and fragmentation at subsequent stages of meiosis, but shows normal homologous chromosome pairing and synapsis during early prophase I. FIGNL1 participates in homologous chromosome recombination and functions downstream of DMC1. Mutation of FIGNL1 increases the number of bivalents in zip4 mutants, but does not change the number of HEI10 foci, indicating that FIGNL1 functions in limiting class II CO formation. FIGNL1 interacts with MEICA1, and colocalizes with MEICA1 in a dynamic pattern as punctate foci located between two linear homologous chromosomes. The localization of FIGNL1 depends on ZEP1-mediated assembly of the synaptonemal complex. Based on these results, we propose that FIGNL1 inhibits non-homologous chromosome interaction and CO formation during rice meiosis.

Nitrogen nutrition contributes to plant fertility by affecting meiosis initiation.

Nitrogen (N), one of the most important plant nutrients, plays crucial roles in multiple plant developmental processes. Spikelets are the primary sink tissues during reproductive growth, and N deficiency can cause floral abortion. However, the roles of N nutrition in meiosis, the crucial step in plant sexual reproduction, are poorly understood. Here, we identified an N-dependent meiotic entrance mutant with loss of function of ELECTRON TRANSFER FLAVOPROTEIN SUBUNIT ß (ETFß) in rice (Oryza sativa). etfß displayed meiosis initiation defects, excessive accumulation of branched-chain amino acids (BCAAs) and decrease in total N contents in spikelets under N starvation, which were rescued by applying excess exogenous inorganic N. Under N starvation, ETFß, through its involvement in BCAA catabolism, promotes N reutilization and contributes to meeting N demands of spikelets, highlighting the impact of N nutrition on meiosis initiation. We conclude that N nutrition contributes to plant fertility by affecting meiosis initiation.

The E3 ubiquitin ligase DESYNAPSIS1 regulates synapsis and recombination in rice meiosis.

Synaptonemal complex (SC) assembly and homologous recombination, the most critical events during prophase I, are the prerequisite for faithful meiotic chromosome segregation. However, the underlying regulatory mechanism remains largely unknown. Here, we reveal that a functional RING finger E3 ubiquitin ligase, DESYNAPSIS1 (DSNP1), plays significant roles in SC assembly and homologous recombination during rice meiosis. In the dsnp1 mutant, homologous synapsis is discontinuous and aberrant SC-like polycomplexes occur independent of coaligned homologous chromosomes. Accompanying the decreased foci of HEI10, ZIP4, and MER3 on meiotic chromosomes, the number of crossovers (COs) decreases dramatically in dsnp1 meiocytes. Furthermore, the absence of central elements largely restores the localization of non-ZEP1 ZMM proteins and the number of COs in the dsnp1 background. Collectively, DSNP1 stabilizes the canonical tripartite SC structure along paired homologous chromosomes and further promotes the formation of COs.

Concurrent Disruption of Genetic Interference and Increase of Genetic Recombination Frequency in Hybrid Rice Using CRISPR/Cas9.

Manipulation of the distribution and frequency of meiotic recombination events to increase genetic diversity and disrupting genetic interference are long-standing goals in crop breeding. However, attenuation of genetic interference is usually accompanied by a reduction in recombination frequency and subsequent loss of plant fertility. In the present study, we generated null mutants of the ZEP1 gene, which encodes the central component of the meiotic synaptonemal complex (SC), in a hybrid rice using CRISPR/Cas9. The null mutants exhibited absolute male sterility but maintained nearly unaffected female fertility. By pollinating the zep1 null mutants with pollen from indica rice variety 93-11, we successfully conducted genetic analysis and found that genetic recombination frequency was greatly increased and genetic interference was completely eliminated in the absence of ZEP1. The findings provided direct evidence to support the controversial hypothesis that SC is involved in mediating interference. Additionally, the remained female fertility of the null mutants makes it possible to break linkage drag. Our study provides a potential approach to increase genetic diversity and fully eliminate genetic interference in rice breeding.

Replication protein A large subunit (RPA1a) limits chiasma formation during rice meiosis.

Replication protein A (RPA), a single-stranded DNA-binding protein, plays essential role in homologous recombination. However, because deletion of RPA causes embryonic lethality in mammals, the exact function of RPA in meiosis remains unclear. In this study, we generated an rpa1a mutant using CRISPR/Cas9 technology and explored its function in rice (Oryza sativa) meiosis. In rpa1a, 12 bivalents were formed at metaphase I, just like in wild-type, but chromosome fragmentations were consistently observed at anaphase I. Fluorescence in situ hybridization assays indicated that these fragmentations were due to the failure of the recombination intermediates to resolve. Importantly, the mutant had a highly elevated chiasma number, and loss of RPA1a could completely restore the 12 bivalent formations in the zmm (for ZIP1-4, MSH4/5, and MER3) mutant background. Protein-protein interaction assays showed that RPA1a formed a complex with the methyl methansulfonate and UV sensitive 81 (and the Fanconi anemia complementation group M-Bloom syndrome protein homologs (RECQ4A)-Topoisomerase3α-RecQ-mediated genome instability 1 complex to regulate chiasma formation and processing of the recombination intermediates. Thus, our data establish a pivotal role for RPA1a in promoting the accurate resolution of recombination intermediates and in limiting redundant chiasma formation during rice meiosis.

Reproductive cells and peripheral parietal cells collaboratively participate in meiotic fate acquisition in rice anthers.

In flowering plants, the transition from mitosis to meiosis is the precondition for gametogenesis, which is the most crucial event during sexual reproduction. Here, we report an intriguing mechanism whereby germ cells and surrounding somatic cells cooperatively involve in the meiotic switch during anther development in rice (Oryza sativa). In double mutants with loss function of both leptotene chromosome establishment- and somatic cell layer differentiation-associated genes, chromosome morphology in the reproductive cells remains the same as that in somatic cells, and sporogenous cells fail to differentiate into pollen mother cells. OsSPOROCYTELESS and MICROSPORELESS1, two pivotal genes involved in meiosis entry, are prominently downregulated in anthers of plants with mutations in both MULTIPLE SPOROCYTE1 and LEPTOTENE 1. In addition, the transcription of redox-related genes is also affected. Therefore, germ cells and the surrounding somatic cells collaboratively participate in meiosis initiation in rice.

PRD1, a homologous recombination initiation factor, is involved in spindle assembly in rice meiosis.

The bipolar spindle structure in meiosis is essential for faithful chromosome segregation. PUTATIVE RECOMBINATION INITIATION DEFECT 1 (PRD1) previously has been shown to participate in the formation of DNA double strand breaks (DSBs). However, the role of PRD1 in meiotic spindle assembly has not been elucidated. Here, we reveal by both genetic analysis and immunostaining technology that PRD1 is involved in spindle assembly in rice (Oryza sativa) meiosis. We show that DSB formation and bipolar spindle assembly are disturbed in prd1 meiocytes. PRD1 signals display a dynamic pattern of localization from covering entire chromosomes at leptotene to congregating at the centromere region after leptotene. Centromeric localization of PRD1 signals depends on the organization of leptotene chromosomes, but not on DSB formation and axis establishment. PRD1 exhibits interaction and co-localization with several kinetochore components. We also find that bi-orientation of sister kinetochores within a univalent induced by mutation of REC8 can restore bipolarity in prd1. Furthermore, PRD1 directly interacts with REC8 and SGO1, suggesting that PRD1 may play a role in regulating the orientation of sister kinetochores. Taken together, we speculate that PRD1 promotes bipolar spindle assembly, presumably by modulating the orientation of sister kinetochores in rice meiosis.

Efficacy and safety of tratinterol hydrochloride tablets in bronchial asthma: a randomized double-blind and multicenter clinical trial.

PURPOSE: The aim of this study was to investigate the efficacy and safety of tratinterol hydrochloride in bronchial asthma (BA) treatment. METHODS: Patients enrolled in this study were distributed randomly into a treatment group (tratinterol hydrochloride) and an active control group (procaterol hydrochloride) and were treated for 2 weeks after running-in. The end points were changes in pulmonary function and clinical symptoms after administration. Safety indices were physical examinations, laboratory testing and spontaneous reporting. FINDINGS: We enrolled 732 subjects, -365 in the treatment group and 367 in the active control group. Forced expiratory volume (FEV1), significantly increased in both group after treatment (P < 0.05). Least-squares (LS) means were -0.03/in the full-analysis set (FAS) and -0.02 in the per-protocol set (PPS) set, and 95% confidence intervals (CIs) for these sets were -0.09 to 0.03 and -0.08 to 0.04, respectively. Forced expiratory volume (FVC), morning peak expiratory flow (PEF) and asthma scores were significantly different with pretreatment (P < 0.05). There was no difference in asymptomatic days or frequency of relief medicine use (P > 0.05). No serious adverse events occurred. IMPLICATIONS: Tratinterol hydrochloride was effective, safe and not inferior to procaterol hydrochloride in treating BA.

Oryza sativa RNA-Dependent RNA Polymerase 6 Contributes to Double-Strand Break Formation in Meiosis.

RNA-dependent RNA polymerase 6 (RDR6) is a core component of the small RNA biogenesis pathway, but its function in meiosis is unclear. Here, we report a new allele of OsRDR6 (Osrdr6-meiosis [Osrdr6-mei]), which causes meiosis-specific phenotypes in rice (Oryza sativa). In Osrdr6-mei, meiotic double-strand break (DSB) formation is partially blocked. We created a biallelic mutant with more severe phenotypes, Osrdr6-bi, by crossing Osrdr6-mei with a knockout mutant, Osrdr6-edit In Osrdr6-bi meiocytes, 24 univalents were observed, and no histone H2AX phosphorylation foci were detected. Compared with the wild type, the number of 21-nucleotide small RNAs in Osrdr6-mei was dramatically lower, while the number of 24-nucleotide small RNAs was significantly higher. Thousands of differentially methylated regions (DMRs) were discovered in Osrdr6-mei, implying that OsRDR6 plays an important role in DNA methylation. There were 457 genes downregulated in Osrdr6-mei, including three genes, CENTRAL REGION COMPONENT1, P31 comet , and O. sativa SOLO DANCERS, related to DSB formation. Interestingly, the downregulated genes were associated with a high level of 24-nucleotide small RNAs but less strongly associated with DMRs. Therefore, we speculate that the alteration in expression of small RNAs in Osrdr6 mutants leads to the defects in DSB formation during meiosis, which might not be directly dependent on RNA-directed DNA methylation.

The SUN Domain Proteins OsSUN1 and OsSUN2 Play Critical but Partially Redundant Roles in Meiosis.

During meiosis, Sad1/UNC-84 (SUN) domain proteins play conserved roles in promoting telomere bouquet formation and homologous pairing across species. Arabidopsis (Arabidopsis thaliana) AtSUN1 and AtSUN2 have been shown to have overlapping functions in meiosis. However, the role of SUN proteins in rice (Oryza sativa) meiosis and the extent of functional redundancy between them remain elusive. Here, we generated single and double mutants of OsSUN1 and OsSUN2 in rice using genome editing. The Ossun1 Ossun2 double mutant showed severe defects in telomere clustering, homologous pairing, and crossover formation, suggesting that OsSUN1 and OsSUN2 are essential for rice meiosis. When introducing a mutant allele of O. sativa SPORULATION11-1 (OsSPO11-1), which encodes a topoisomerase initiating homologous recombination, into the Ossun1 Ossun2 mutant, we observed a combined Osspo11-1- and Ossun1 Ossun2-like phenotype, demonstrating that OsSUN1 and OsSUN2 promote bouquet formation independent of OsSPO11-1 but regulate pairing and crossover formation downstream of OsSPO11-1. Importantly, the Ossun1 single mutant had a normal phenotype, but meiosis was disrupted in the Ossun2 mutant, indicating that OsSUN1 and OsSUN2 are not completely redundant in rice. Further analyses revealed a genetic dosage-dependent effect and an evolutionary differentiation between OsSUN1 and OsSUN2 These results suggested that OsSUN2 plays a more critical role than OsSUN1 in rice meiosis. Taken together, this work reveals the essential but partially redundant roles of OsSUN1 and OsSUN2 in rice meiosis and demonstrates that functional divergence of SUN proteins has taken place during evolution.

Defective Microspore Development 1 is required for microspore cell integrity and pollen wall formation in rice.

Highly coordinated pollen wall patterning is essential for male reproductive development. Here, we report the identification of Defective Microspore Development 1 (DMD1), which encodes a nuclear-localized protein possessing transactivation activity. DMD1 is preferentially expressed in the tapetum and microspores during post-meiotic development. Mutations in DMD1 cause a male-sterile phenotype with impaired microspore cell integrity. The mutants display abnormal callose degradation, accompanied by inhibited primexine thickening in the newly released microspores. Several genes associated with callose degradation and primexine formation are downregulated in dmd1 anthers. In addition, irregular Ubisch body morphology and discontinuous endexine occur, and the baculum is completely absent in dmd1. DMD1 interacts with Tapetum Degeneration Retardation (TDR), a basic helix-loop-helix transcription factor required for exine formation. Taken together, our results suggest that DMD1 is responsible for microspore cell integrity, primexine formation and exine pattern formation during Oryza sativa (rice) microspore development.

OsRAD51D promotes homologous pairing and recombination by preventing nonhomologous interactions in rice meiosis.

Homologous recombination is carefully orchestrated to maintain genome integrity. RAD51D has been previously shown to be essential for double-strand break repair in mammalian somatic cells. However, the function of RAD51D during meiosis is largely unknown. Here, through detailed analyses of Osrad51d single and double mutants, we pinpoint the specific function of OsRAD51D in coordinating homologous pairing and recombination by preventing nonhomologous interactions during meiosis. OsRAD51D is associated with telomeres in both meiocytes and somatic cells. Loss of OsRAD51D leads to significant induction of nonhomologous pairing and chromosome entanglements, suggesting its role in suppressing nonhomologous interactions. The failed localization of OsRAD51 and OsDMC1 in Osrad51d, together with the genetic analysis of Osrad51d Osdmc1a Osdmc1b, indicates that OsRAD51D acts at a very early stage of homologous recombination. Observations from the Osrad51d pair1 and Osrad51d ku70 double mutants further demonstrate that nonhomologous interactions require double-strand break formation but do not depend on the KU70-mediated repair pathway. Moreover, the interplay between OsRAD51D and OsRAD51C indicates both conservation and divergence of their functions in meiosis. Altogether, this work reveals that OsRAD51D plays an essential role in the inhibition of nonhomologous connections, thus guaranteeing faithful pairing and recombination during meiosis.

OsHOP2 regulates the maturation of crossovers by promoting homologous pairing and synapsis in rice meiosis.

Meiotic recombination is closely linked with homologous pairing and synapsis. Previous studies have shown that HOMOLOGOUS PAIRING PROTEIN2 (HOP2), plays an essential role in homologous pairing and synapsis. However, the mechanism by which HOP2 regulates crossover (CO) formation has not been elucidated. Here, we show that OsHOP2 mediates the maturation of COs by promoting homologous pairing and synapsis in rice (Oryza sativa) meiosis. We used a combination of genetic analysis, immunolocalization and super-resolution imaging to analyze the function of OsHOP2 in rice meiosis. We showed that full-length pairing, synapsis and CO formation are disturbed in Oshop2 meiocytes. Moreover, structured illumination microscopy showed that OsHOP2 localized to chromatin and displayed considerable co-localization with axial elements (AEs) and central elements (CEs). Importantly, the interaction between OsHOP2 and a transverse filament protein of synaptonemal complex (ZEP1), provided further evidence that OsHOP2 was involved in assembly or stabilization of the structure of the synaptonemal complex (SC). Although the initiation of recombination and CO designation occur normally in Oshop2 mutants, mature COs were severely reduced, and human enhancer of invasion 10 (HEI10)10 foci were only present on the synapsed region. Putting the data together, we speculate that OsHOP2 may serve as a global regulator to coordinate homologous pairing, synapsis and meiotic recombination in rice meiosis.

The OsRR24/LEPTO1 Type-B Response Regulator is Essential for the Organization of Leptotene Chromosomes in Rice Meiosis.

Response regulators play significant roles in controlling various biological processes; however, their roles in plant meiosis remain unclear. Here, we report the identification of OsRR24/LEPTOTENE1 (LEPTO1), a rice (Oryza sativa) type-B response regulator that participates in the establishment of key molecular and morphological features of chromosomes in leptotene, an early stage of prophase I in meiosis. Although meiosis initiates normally, as indicated by staining of the centromere-specific histone CENH3, the meiotic chromosomes in lepto1 mutant pollen mother cells fail to form the thin thread-like structures that are typical of leptotene chromosomes in wild-type pollen mother cells. Furthermore, lepto1 mutants fail to form chromosomal double-strand breaks, do not recruit meiosis-specific proteins to the meiotic chromosomes, and show disrupted callose deposition. LEPTO1 also is essential for programmed cell death in tapetal cells. LEPTO1 contains a conserved signal receiver domain (DDK) and a myb-like DNA binding domain at the N terminus. LEPTO1 interacts with two authentic histidine phosphotransfer (AHP) proteins, OsAHP1 and OsAHP2, via the DDK domain, and a phosphomimetic mutation of the DDK domain relieves its repression of LEPTO1 transactivation activity. Collectively, our results show that OsRR24/LEPTO1 plays a significant role in the leptotene phase of meiotic prophase I.

The zinc finger protein DCM1 is required for male meiotic cytokinesis by preserving callose in rice.

Meiotic cytokinesis influences the fertility and ploidy of gametes. However, limited information is available on the genetic control of meiotic cytokinesis in plants. Here, we identified a rice mutant with low male fertility, defective callose in meiosis 1 (dcm1). The pollen grains of dcm1 are proved to be defective in exine formation. Meiotic cytokinesis is disrupted in dcm1, resulting in disordered spindle orientation during meiosis II and formation of pollen grains with varied size and DNA content. We demonstrated that meiotic cytokinesis defect in dcm1 is caused by prematurely dissolution of callosic plates. Furthermore, peripheral callose surrounding the dcm1 pollen mother cells (PMCs) also disappeared untimely around pachytene. The DCM1 protein contains five tandem CCCH motifs and interacts with nuclear poly (A) binding proteins (PABNs) in nuclear speckles. The expression profiles of genes related to callose synthesis and degradation are significantly modified in dcm1. Together, we propose that DCM1 plays an essential role in male meiotic cytokinesis by preserving callose from prematurely dissolution in rice.

HEIP1 regulates crossover formation during meiosis in rice.

During meiosis, the number of double-strand breaks (DSBs) far exceeds the final number of crossovers (COs). Therefore, to identify proteins involved in determining which of these DSBs repaired into COs is critical in understanding the mechanism of CO control. Across species, HEI10-related proteins play important roles in CO formation. Here, through screening for HEI10-interacting proteins via a yeast two-hybrid system, we identify a CO protein HEI10 Interaction Protein 1 (HEIP1) in rice. HEIP1 colocalizes with HEI10 in a dynamic fashion along the meiotic chromosomes and specially localizes onto crossover sites from late pachytene to diplotene. Between these two proteins, HEI10 is required for the loading of HEIP1, but not vice versa. Moreover, mutations of the HEIP1 gene cause the severe reduction of chiasma frequency, whereas early homologous recombination processes are not disturbed and synapsis proceeds normally. HEIP1 interacts directly with ZIP4 and MSH5. In addition, the loading of HEIP1 depends on ZIP4, but not on MER3, MSH4, or MSH5. Together, our results suggest that HEIP1 may be a member of the ZMM group and acts as a key element regulating CO formation.

Ornithine δ-aminotransferase is critical for floret development and seed setting through mediating nitrogen reutilization in rice.

Nitrogen is one of the most important nutrient element that is essential for plant growth and development. Many genes have been reported to contribute to nitrogen absorption and transportation. However, genes involved in nitrogen reutilization are seldom reported. Ornithine δ-aminotransferase (δOAT) is the enzyme connecting arginine cycling and proline cycling. Here, we found that OsOAT, the homologue of δOAT in rice, is essential for nitrogen reutilization through mediating arginase activity. In the Osoat mutant, metabolic abnormality induced by nitrogen deficiency in floret causes malformed glumes, incapable glume opening and anther indehiscence. These defects in the mutant affect the pollination process and lead to a low seed setting rate as well as abnormal seed shape. Intriguingly, urea can rescue the phenotypes of the Osoat mutant. Therefore, OsOAT is crucial for nitrogen reutilization and plays a critical role in floret development and seed setting in rice.