Increasing crop yield is one of the most important goals of plant science research. Grain size is a major determinant of grain yield in cereals and is a target trait for both domestication and artificial breeding(1). We showed that the quantitative trait locus (QTL) GS5 in rice controls grain size by regulating grain width, filling and weight. GS5 encodes a putative serine carboxypeptidase and functions as a positive regulator of grain size, such that higher expression of GS5 is correlated with larger grain size. Sequencing of the promoter region in 51 rice accessions from a wide geographic range identified three haplotypes that seem to be associated with grain width. The results suggest that natural variation in GS5 contributes to grain size diversity in rice and may be useful in improving yield in rice and, potentially, other crops(2).

HighlightTwo SNPs in the promoter of GS5 are responsible for expression variation controlling grain size. Enhanced expression of GS5 competitively inhibits the interaction between OsBAK1 and OsMSBP1, promoting grain size.

Calcium-dependent protein kinases (CPKs) are serine/threonine protein kinases that function in plant stress responses. Although CPKs are recognized as key messengers in signal transduction, the specific roles of CPKs and the molecular mechanisms underlying their activity remain largely unknown. Here, we characterized the function of OsCPK24, a cytosol-localized calciumdependent protein kinase in rice. OsCPK24 was universally and highly expressed in rice plants and was induced by cold treatment. Whereas OsCPK24 knockdown plants exhibited increased sensitivity to cold compared to wild type (WT), OsCPK24-overexpressing plants exhibited increased cold tolerance. Plants overexpressing OsCPK24 exhibited increased accumulation of proline (an osmoprotectant) and glutathione (an antioxidant) and maintained a higher GSH/GSSG (reduced glutathione to oxidized glutathione) ratio during cold stress compared to WT. In addition to these effects in response to cold stress, we observed the kinase activity of OsCPK24 varied under different calcium concentrations. Further, OsCPK24 phosphorylated OsGrx10, a glutathionedependent thioltransferase, at rates modulated by changes in calcium concentration. Together, our results support the hypothesis that OsCPK24 functions as a positive regulator of cold stress tolerance in rice, a process mediated by calcium signaling and involving phosphorylation and the inhibition of OsGrx10 to sustain higher glutathione levels.

Pollen development is critical to the reproductive success of flowering plants, but how it is regulated is not well understood. Here, we isolated two allelic male-sterile mutants of OsMYB80 and investigated how OsMYB80 regulates male fertility in rice. OsMYB80 was barely expressed in tissues other than anthers, where it initiated the expression during meiosis, reached the peak at the tetrad-releasing stage and then quickly declined afterward. The osmyb80 mutants exhibited premature tapetum cell death, lack of Ubisch bodies, no exine and microspore degeneration. To understand how OsMYB80 regulates anther development, RNA-seq analysis was conducted to identify genes differentially regulated by OsMYB80 in rice anthers. In addition, DNA affinity purification sequencing (DAP-seq) analysis was performed to identify DNA fragments interacting with OsMYB80 in vitro. Overlap of the genes identified by RNA-seq and DAP-seq revealed 188 genes that were differentially regulated by OsMYB80 and also carried an OsMYB80-interacting DNA element in the promoter. Ten of these promoter elements were randomly selected for gel shift assay and yeast one-hybrid assay, and all showed OsMYB80 binding. The 10 promoters also showed OsMYB80-dependent induction when co-expressed in rice protoplast. Functional annotation of the 188 genes suggested that OsMYB80 regulates male fertility by directly targeting multiple biological processes. The identification of these genes significantly enriched the gene networks governing anther development and provided much new information for the understanding of pollen development and male fertility.

Meiotic recombination plays a central role in maintaining genome stability and increasing genetic diversity. Although meiotic progression and core components are widely conserved across kingdoms, significant differences remain among species. Here we identify a rice gene ABERRANT GAMETOGENESIS 1 (AGG1) that controls both male and female gametogenesis. Cytological and immunostaining analysis showed that in the osagg1 mutant the early recombination processes and synapsis occurred normally, but the chiasma number was dramatically reduced. Moreover, OsAGG1 was found to interact with ZMM proteins OsHEI10, OsZIP4, and OsMSH5. These results suggested that OsAGG1 plays an important role in crossover formation. Phylogenetic analysis showed that OsAGG1 is a plant-specific protein with a highly conserved N-terminal region. Further genetic and protein interaction analyses revealed that the conserved N-terminus was essential for the function of the OsAGG1 protein. Overall, our work demonstrates that OsAGG1 is a novel and critical component in rice meiotic crossover formation, expanding our understanding of meiotic progression. This study identified a plant-specific gene ABERRANT GAMETOGENESIS 1 that is required for meiotic crossover formation in rice. The conserved N-terminus of the AGG1 protein was found to be essential for its function.

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Large‐scale production of male sterile seeds can be achieved by introducing a fertility‐restoration gene linked with a pollen‐killer gene into a recessive male sterile mutant. We attempted to construct this system in rice by using a late‐stage pollen‐specific (LSP) promoter driving the expression of maize α‐amylase gene ZM‐AA1. To obtain such promoters in rice, we conducted comparative RNA‐seq analysis of mature pollen with meiosis anther, and compared this with the transcriptomic data of various tissues in the Rice Expression Database, resulting in 269 candidate LSP genes. Initial test of nine LSP genes showed that only the most active OsLSP3 promoter could drive ZM‐AA1 to disrupt pollen. We then analyzed an additional 22 LSP genes and found 12 genes stronger than OsLSP3 in late‐stage anthers. The promoters of OsLSP5 and OsLSP6 showing higher expression than OsLSP3 at stages 11 and 12 could drive ZM‐AA1 to inactivate pollen, while the promoter of OsLSP4 showing higher expression at stage 12 only could not drive ZM‐AA1 to disrupt pollen, suggesting that strong promoter activity at stage 11 was critical for pollen inactivation. The strong pollen‐specific promoters identified in this study provided valuable tools for genetic engineering of rice male sterile system for hybrid rice production.

Pollen grains are covered by exine that protects the pollen from stress and facilitates pollination. Here we isolated a male sterile mutant s13283 in rice exhibiting aborted pollen with abnormal exine and defective aperture. The mutant gene encodes a novel plasma membrane‐localized legume‐lectin receptor kinase that we named OsLecRK‐S.7. OsLecRK‐S.7 was expressed at different levels in all tested tissues and throughout anther development. In vitro kinase assay showed OsLecRK‐S.7 capable of autophosporylation. Mutation in s13283 (E560K) and mutation of the conserved ATP binding site (K418E) both knocked out the kinase activity. Mass spectrometry showed Thr376, Ser378, Thr386, Thr403, and Thr657 to be the autophosphorylation sites. Mutation of individual autophosphorylation site affected the in vitro kinase activity to different degrees, but did not abolish the gene function in fertility complementation. oslecrk‐s.7 mutant plant overexpressing OsLecRK‐S.7 recovered male fertility but showed severe growth retardation with reduced number of tillers, and these phenotypes were abolished by E560K or K418E mutation. The results indicated that OsLecRK‐S.7 was a key regulator of pollen development.