A dose-sensitive miR164e-WRKY50-JAZ7 module fine-tunes grain size in rice

Polyploidization drives evolutionary innovation and crop domestication by leveraging novel gene combinations and increased genetic variation. Although tetraploid rice shows promise for yield enhancement due to its larger grains, the underlying regulatory mechanisms remain unclear. In this study, using tetraploid rice CX35-4x and its diploid counterpart CX35-2x, we constructed the first comprehensive mRNA and microRNA atlas during grain development. Transcriptomic analysis revealed that during key stages of hull development, transcription factors and genes involved in plant hormone signal transduction exhibited pronounced dose-sensitive effects in tetraploid rice, forming a core regulatory network for polyploid grain development. Integrated analysis revealed that dose-sensitive miRNAs regulate these key genes, enabling construction of a miRNA-target network controlling grain size. Functional validation of key dose-sensitive miR164e and WRKY50 transcription factors confirmed that miR164e inhibit WRKY50 expression via 3'UTR targeting, negatively regulating grain development. miR164e mutation increased thousand-grain weight by 25%, while WRKY50 mutation caused 16% reduction. Further DAP-seq analysis identified JAZ7 as direct WRKY50 targets. As dose-sensitive hormone signaling components, JAZ7 expression is activated by WRKY50, and JAZ7 loss-of-function reduced thousand-grain weight by 18%. Our study systematically reveals the dose-sensitive regulatory network controlling polyploid rice grain size, and for the first time, establishes the novel miR164e-WRKY50-JAZ7 dose-sensitive module as a key regulatory mechanism. These findings provide new insights into polyploidy advantage and valuable genetic resources for further rice yield improvement.