The RING-domain E3 ubiquitin ligase OsRGLG6 regulates rice grain number and yield via ubiquitination-mediated degradation of OsOTUB1.

Rice (Oryza sativa) yield is a complex trait determined by multiple factors, including grain number per panicle (GPP), which is mainly genetically regulated. In this study, we identified the E3 RING DOMAIN LIGASE (RGLG) OsRGLG6 as a key regulator of GPP in rice. A yeast two-hybrid screen revealed that OsRGLG6 interacts with OsOTUB1, an ovarian tumor domain-containing ubiquitin-specific protease involved in panicle development. Further experiments confirmed the interaction between OsRGLG6 and OsOTUB1 in vitro and in vivo. OsRGLG6, which contains a vWA domain and a RING domain, is localized in the cytoplasm and endoplasmic reticulum and is highly expressed specifically in rice panicles. In vitro and in vivo ubiquitination assays demonstrated that OsRGLG6 ubiquitinates OsOTUB1 and promotes its degradation via the 26S proteasome pathway. CRISPR/Cas9-generated osrglg6 mutants exhibited significant reductions in GPP, indicating a crucial role for OsRGLG6 in the regulation of grain number. Transcriptome analysis revealed that OsRGLG6 regulates multiple downstream genes involved in plant hormone signaling, nitrogen utilization, and stress response, and the OsRGLG6-OsOTUB1 module specifically mediated drought-stress responses in a simulated drought experiment. This study provides insight into the molecular mechanisms that regulate rice GPP and yield, highlighting the potential of OsRGLG6 as a target for improvement of rice productivity. SUPPLEMENTARY INFORMATION: The online version contains supplementary material available at 10.1007/s42994-025-00232-5.