The E3 ubiquitin ligase CSIT1 regulates critical sterility-inducing temperature by ribosome-associated quality control to safeguard two-line hybrid breeding in rice

Two-line hybrid breeding can fully utilize heterosis in crops. In thermo-sensitive genic male sterile (TGMS) lines, low critical sterility-inducing temperature (CSIT) is vital to safeguard the production of two-line hybrid seeds in rice (Oryza sativa), but the molecular mechanism determining CSIT is unclear. Here, we report the cloning of CSIT1, which encodes an E3 ubiquitin ligase, and show that CSIT1 modulates the CSIT of thermo-sensitive genic male sterility 5 (tms5)-based TGMS lines through ribosome-associated quality control (RQC). Biochemical assays demonstrated that CSIT1 binds to the 80S ribosomes and ubiquitinates abnormal nascent polypeptides for degradation in the RQC process. Loss of CSIT1 function inhibits the possible damage of tms5 to the ubiquitination system and protein translation, resulting in enhanced accumulation of anther-related proteins such as catalase to suppress abnormal accumulation of reactive oxygen species and premature programmed cell death in the tapetum, thereby leading to a much higher CSIT in the tms5-based TGMS lines. Taken together, our findings reveal a regulatory mechanism of CSIT, providing new insights into RQC and potential targets for future two-line hybrid breeding. This study shows that the E3 ubiquitin ligase CSIT1 ubiquitinates abnormal proteins by ribosome-associated quality control and affects the programmed cell death in the tapetum to regulate critical sterility-inducing temperature of thermo-sensitive genic male sterility in rice. The results reveals a regulatory mechanism of critical sterility-inducing temperature, providing new insights into ribosome-associated quality control and potential targets for two-line hybrid breeding.