The domestication of rice is a cornerstone of human agriculture, with *Oryza sativa* feeding over half the world's population. Increasing rice yields while enhancing environmental resilience is crucial, but challenging due to the negative correlation between yield and stress tolerance. Grain size and shape are key yield determinants, controlled by quantitative trait loci (QTLs) affecting various genetic pathways. Many genes like *GW2*, *GL3.1*, *GS5*, *GW8*, *GS2/GL2*, *GS3*, *GL7/GW7*, and *GS9*, have been identified, but the underlying mechanisms remain complex. Wild rice (*Oryza rufipogon*) offers valuable genetic resources for improving yield and stress tolerance in cultivated rice. While wild rice generally has inferior yield traits, it possesses genes for environmental adaptation lost during domestication. This study focuses on a QTL, *qGL12*, from wild rice that enhances grain length, aiming to elucidate the mechanisms involved and determine if improvements in yield and stress tolerance are synergistic.

Extensive research has focused on identifying genes affecting grain size in rice, revealing several key players involved in hormone signaling, G-protein pathways, and transcriptional regulation. *GS3*, a transmembrane protein, contributes significantly to grain length differences between indica and japonica varieties. *GL3.1*, a serine/threonine phosphatase, regulates grain length through cell cycle control. Transcription factors like OsMADSI and OsGRF4 (encoded by *GS2*) also play crucial roles. OsGRF4's interaction with OsGIF1/2/3 coactivators further impacts grain size. While progress has been made, the complex interplay of these factors and the mechanisms underlying yield-stress tolerance trade-offs require further investigation. Wild rice has emerged as a potential source of beneficial genes for rice improvement, but the effective utilization of these resources requires a deeper understanding of their function and interaction with existing cultivated rice genes.

To identify genes influencing grain length and 1000-grain weight (TGW), chromosome segment substitution lines (CSSLs) were constructed using an indica variety ('9311') and *O. rufipogon*. A major QTL, *qGL12*, was identified and fine-mapped to a 16-kb interval. Candidate genes were identified, and their expression levels were compared between a near-isogenic line (NIL) and the recurrent parent. Overexpression (OE) and knockout (KO) lines were generated using CRISPR/Cas9 technology to confirm candidate gene function. RNA sequencing was performed to identify downstream genes regulated by *GL12*. Subcellular localization studies were conducted to determine GL12 protein location. Transient expression assays in *Nicotiana benthamiana* were used to assess promoter activity. Yeast one-hybrid, yeast two-hybrid, and electrophoretic mobility shift assays (EMSA) examined protein-protein and protein-DNA interactions. Salt tolerance assays were performed to assess the effect of *GL12* on stress response. Finally, haplotype and evolutionary analyses were conducted to determine the role of *GL12* in indica and japonica subspecies divergence using data from the 3K Rice Genomes Project and the Rice Super Pangenome Information Resource Database.

The wild rice *GL12^W* allele significantly increased grain length and 1000-grain weight in both indica and japonica backgrounds. *GL12^W* overexpression altered the expression of multiple grain size-related genes, including *GS2*, primarily by promoting cell elongation in the spikelet hull. RNA sequencing revealed that *GL12^W* regulated the expression of 8079 genes, many involved in metabolism and genetic information processing. A G/T SNP in the *GL12* promoter was identified as crucial for its interaction with OsGIF1, a coactivator that positively regulates *GL12^W* expression in young panicles. The same SNP also modulates the interaction with WRKY53, a transcription factor that represses *GL12^W* expression under salt stress. *GL12^W* overexpression enhanced salt tolerance by upregulating salt-tolerance related genes, including *NCED3* and *NACs*. The G/T variation was found to be strongly associated with the divergence between indica and japonica subspecies, suggesting artificial selection during rice domestication. *GL12^W* improved plant height and panicle length but reduced the number of grains per panicle.

This study successfully identified a wild rice gene, *GL12^W*, that improves both grain length and salt tolerance in cultivated rice. The synergistic effect is mediated by the interplay between the G/T SNP in the *GL12* promoter and the transcription coactivator GIF1 and the transcription factor WRKY53. GIF1 enhances *GL12^W* expression during early panicle development, promoting grain size, while WRKY53 negatively regulates *GL12^W* expression during salt stress, promoting tolerance. This research highlights the potential of wild rice genes for enhancing both yield and stress resistance, and provides a valuable resource for rice improvement programs. However, the negative effect on grain number per panicle observed with *GL12^W* suggests further research is needed to optimize its use in breeding programs.

The wild rice *GL12^W* gene presents a promising target for improving both grain length and salt tolerance in rice. Its regulation by GIF1 and WRKY53, controlled by a G/T SNP in its promoter region, offers a compelling mechanism for synergistic trait enhancement. While the observed reduction in grain number per panicle needs addressing, future research could focus on refining gene editing strategies or combining *GL12^W* with other genes to optimize yield and stress resistance simultaneously. The findings underscore the potential of wild rice genetic resources for addressing critical challenges in rice breeding.

The study primarily focused on the effects of *GL12^W* on grain length and salt tolerance. While the negative effect on grain number per panicle was noted, more detailed investigations into the broader impact on overall yield are needed. Additionally, the study's findings might not be fully generalizable to all rice varieties or environmental conditions. Further research should investigate the gene's interactions with other QTLs and its performance under diverse environmental stresses.