Redundant Glycerol-3-Phosphate Acyltransferases Regulate Thermo-Sensitive Genic Male Sterility in Rice

Thermo-sensitive genic male sterility (TGMS) lines play a pivotal role in two-line hybrid rice breeding. However, the availability of elite TGMS germplasm remains limited, and the molecular mechanisms underlying fertility transition of TGMS lines are still poorly understood. Here, we identified a novel TGMS line, Osgpat6.1, which exhibits temperature-dependent fertility transition, with male sterility under high-temperature (HT) and restored fertility under low-temperature (LT). Cytological observations revealed that the TGMS line exhibits disrupted programmed cell death (PCD) of the tapetum under HT conditions. This disruption leads to delayed tapetum degradation and defective pollen exine formation. Map-based cloning identified a premature termination mutation in glycerol-3-phosphate acyltransferase 6.1 (OsGPAT6.1), which is highly expressed in anthers and essential for lipid biosynthesis. Lipidomic profiling revealed that disruption of OsGPAT6.1 perturbed lipid metabolism during pollen development. Genetic studies further revealed that male sterility in Osgpat6.1 mutant was restored through functional redundancy with its homologues OsGPAT6.2 and OsGPAT6.3 under LT. Overexpression of OsGPAT6.2 or OsGPAT6.3 partially restored the fertility of Osgpat6.1 mutant under HT. However, simultaneous disruption of all three homologues abolished fertility restoration even under LT. Notably, introgression of the Osgpat6.1 allele into diverse rice cultivars produced stable TGMS lines without compromising hybrid vigor. Collectively, these findings demonstrate that dysfunction of GPAT confers TGMS in rice by disrupting glycerolipid metabolism and provides TGMS germplasm to facilitate two-line hybrid rice breeding.