Multi-locus allelic architecture underlying natural variation in leaf rolling in japonica rice

Leaf rolling is a key component of rice canopy architecture that affects light interception, microclimate formation, and planting density. The contribution of naturally occurring allelic variation to quantitative variation in leaf rolling within cultivated rice remains poorly understood, while extreme leaf rolling caused by loss-of-function mutations often results in detrimental pleiotropic effects. Herein, we examined how multi-locus allelic variation contributes to natural variation in leaf rolling within japonica rice. Leaf rolling was quantified based on the leaf rolling index (LRI) using a panel of 201 japonica accessions. The phenotype was transformed using the Yeo-Johnson method to reduce strong right skewness and improve the distributional properties of the data, thereby facilitating subsequent regression modeling. Haplotype analyses were performed for previously reported leaf rolling-associated genes and genome-wide association study (GWAS) lead loci, leading to the identification of five loci exhibiting substantial haplotype-dependent phenotypic variation. Phenotypically defined allelic groups represented these loci were subsequently evaluated using multiple linear regression (MLR), with the first two principal components derived from genome-wide SNP data included as covariates to account for population structure. The final MLR model identified four loci (qALR1, OsYABBY1, OsSLL2, and OsSRL10) as the independent contributors to leaf rolling variation, collectively explaining 21% of the variance in the transformed phenotype after accounting for population structure. Model diagnostics and ten-fold cross-validation supported the statistical validity of the framework and indicated stable model performance across validation folds. Analysis of multi-locus allelic combinations showed 13 distinct configurations that clustered into three phenotypically differentiated groups. This reflected the cumulative dosage of high-leaf rolling alleles. Thus, the natural variation in leaf rolling in japonica rice is governed by the additive effects of multiple moderate-impact loci. The multi-locus allelic framework established here provides a statistically sound and biologically interpretable basis for dissecting polygenic canopy traits and practical guidance for developing genetic materials aimed at optimizing rice plant architecture.