Quantitative dissection of sexual dimorphism in mice through Y-linked gene knockouts and multivariate phenotyping

Sexual dimorphism (SDM) is regulated by sex chromosomes, yet the specific contribution of individual genes remains unclear. To address this, we conducted a comprehensive phenotyping analysis to investigate the roles of Y chromosome-linked genes in SDM formation in mice. Using C57BL/6J mice, we identified 49 SDM traits across 14 biological systems. To assess gene-specific effects, we generated knockout (KO) mice for 10 unique Y-linked genes using CRISPR/Cas9. As expected, Sry KO resulted in feminization of most SDM traits, including gonadal sex. However, certain body size-related traits remained male-like, suggesting the involvement of additional Y-linked genes. Consistently, KOs of Uty and Usp9y significantly altered traits related to body and organ size. We further applied a dimensionality reduction approach to quantitatively capture SDM variation at the individual level, enabling visualization of phenotypic shifts in each KO mouse. Our findings demonstrate that non-Sry Y-linked genes contribute to SDM and introduce a generalizable framework for quantifying sex differences across individuals and species.