Identification and functional validation of <italic>osg1-7</italic>, a <italic>G1</italic> mutant associated with the sterile lemma identity gene in rice

Rice (Oryza sativa L.) is one of the most important cereal crops worldwide and a model organism for monocotyledonous plants. Yield-related traits in rice are a major focus in crop genetics and molecular breeding, with the morphology and structure of floral organs playing a critical role in determining yield. The sterile lemma, a unique organ of the rice spikelet, serves as a key entry point for understanding the mechanisms of floral development. The LONG STERILE LEMMA1 (G1) gene is widely recognized as a pivotal regulator of sterile lemma identity in rice. In this study, we identified and characterized a new strong mutant of the G1 gene, designated as g1-7, which exhibits distinct morphological and molecular phenotypes, providing deeper insights into the role of G1 in rice development and yield formation.The g1-7 mutant was derived from the Oryza sativa subsp. Indica cv. “Huanghuazhan” through gamma-ray irradiation. Phenotypic analysis revealed that the sterile lemmas of g1-7 were significantly elongated and widened, resembling the lemma in structure. Microscopic observations, including scanning electron microscopy and paraffin sectioning, demonstrated that the mutant sterile lemmas exhibited lemma-like cellular arrangements and vascular bundle patterns, indicating a homeotic transformation. Compared to the wild type, g1-7 showed a significant reduction in secondary branch number and thousand-grain weight, along with a prolonged growth duration from sowing to heading, suggesting that G1 may regulate these agronomic traits either directly or indirectly.Genetic analysis confirmed that the long sterile lemma phenotype of g1-7 was controlled by a single recessive gene. Map-based cloning localized the candidate gene to a 24-kb region on chromosome 7, where G1/Os07g0139300 was identified as the causative gene. Sequencing revealed a single-base deletion (G) at position 575 in the coding sequence of G1 in the mutant, leading to a frameshift mutation and premature termination of translation. This mutation resulted in the loss of 85 amino acids at the carboxyl terminus, including a predicted intrinsically disordered region (IDR), while the conserved ALOG domain remained intact. Subcellular localization confirmed that the G1 protein was nuclear-localized, consistent with its putative role as a transcription factor.Functional validation through complementation and CRISPR/Cas9-mediated knockout experiments further confirmed the role of G1 in sterile lemma development. Complementation of g1-7 with the wild-type G1 gene restored the normal sterile lemma phenotype, while knockout of G1 in the wild-type background recapitulated the g1-7 mutant phenotype. Additionally, quantitative RT-PCR analysis revealed upregulation of lemma-related genes, such as DROOPING LEAF (DL) and OsMADS1, in the sterile lemmas of g1-7, further supporting the lemma-like identity of the mutant organs.These findings highlight the critical role of G1 in maintaining sterile lemma identity and its broader impact on yield-related traits. The loss of the carboxyl-terminal IDR in g1-7 suggests that this region, along with the ALOG domain, is essential for G1 function. The study also provides evidence that G1 and its orthologs, such as TAWAWA1, may share conserved roles in regulating inflorescence architecture. However, whether the effects on thousand-grain weight and growth duration are direct or secondary consequences of sterile lemma transformation requires further investigation.In summary, the results underscore the importance of G1 in sterile lemma specification and its potential applications in rice breeding for yield improvement. The molecular and phenotypic characterization of g1-7 offers valuable insights into the genetic control of floral organ development and provides a foundation for future studies on the ALOG gene family in plants.