The OsWRKY15-OsSUT3 Regulatory Module Enhances Rice Pollen Fertility by Mediating Sucrose Allocation to Anthers.

Sucrose transport and starch accumulation are crucial carbohydrate metabolic processes in rice, playing essential roles during late pollen development to ensure pollen maturation, fertility and high grain production. However, the molecular mechanisms regulating sucrose transport and starch accumulation during rice pollen development remain elusive. In this study, we bred and characterized a semi-fertile rice pollen variant, sfp10, which is derived from highland terraced red rice from Yunnan, China. This variant showed insufficient starch accumulation in pollen, reduced pollen fertility and decreased seed setting compared to the wild type. Using map-based cloning, CRISPR/Cas9 mutagenesis and gene complementation, the OsSUT3 gene, which causes the sfp10 variant, was shown to encode a sucrose transporter (SUT) that transports sucrose into pollen for starch synthesis, ensuring pollen viability and fertility. Additionally, transcriptome analysis identified OsWRKY15, a WRKY transcription factor that exhibits a highly similar spatiotemporal expression pattern to OsSUT3 in late-developing panicles. Yeast one-hybrid, EMSA, dual-luciferase reporter and ChIP-seq assays demonstrated that OsWRKY15 directly binds to the promoter region of OsSUT3 and upregulates its expression. Mutation of OsWRKY15 significantly reduced OsSUT3 expression and impaired pollen fertility and seed setting and these defects were further exacerbated in the wrky15/OsSUT3-KO double-knockout lines, while OsSUT3 overexpression in the wrky15 background significantly rescued pollen fertility and seed setting. These findings demonstrate that the OsWRKY15-OsSUT3 regulatory module is essential for sucrose transport and starch accumulation during late pollen development in rice, providing new insights into the transcriptional regulation of carbon allocation to anthers for rice fertility and grain production.