Sex-aware genome-wide assessment of de novo variants in autism across coding and noncoding regions.

backgroundAutism spectrum disorder (ASD) shows a consistent sex bias, yet how sex shapes de novo variant (DNV) risk across coding and noncoding sequence remains unclear. I analyzed DNVs in > 41,000 parent–child sequenced trios from three ASD family-based cohorts and compared DNV characteristics and enrichment patterns in males and females. Notably, these trios included individuals with ASD as well as those without ASD. I developed a new sex-aware DNV caller (HAT-FLEX) and thoroughly evaluated each candidate DNV using an additional tool introduced in this study, SNOW, to generate a high-confidence callset.

resultsI identified enrichment of missense and loss-of-function (LOF) DNVs both overall and within known ASD-related genes (i.e., SFARI genes). Gene-specific enrichment analyses revealed twelve genes that were exome-wide significant and specific to males, for significance, including FOXP1, SMAD6, AUTS2, CCDC168, PIEZO1, EML6, ZNF84, IGSF23, OTOG, NHSL1, ADNP, and FREM3 and three genes that were specific to females, for significance, including TAOK1, MECP2, and DDX3X within a variant class. Direct comparisons of DNVs in males and females revealed MECP2 as the only exome-wide significant gene; however, GABBR2 was also trending toward enrichment in the direct males with ASD comparison to females with ASD. Furthermore, probit analyses support a female protective effect and demonstrates that damaging DNVs, particularly LOF variants, including potentially stronger X-linked effects in females, are associated with increased ASD liability, whereas synonymous variants are not. Finally, I analyzed promoters and identified a single significant promoter region (p = 3.8 × 10−13), associated with the WDR74 gene, with the signal driven by DNVs observed in males with ASD. Surprisingly, the noncoding RNA gene RNU2-2 lies within this significant WDR74 promoter and accounted for most of the DNVs in the region. RNU2-2 DNVs were present in 0.2% of individuals with ASD in comparison to 0.05% of individuals without ASD.

conclusionsThese findings show that ASD DNV risk differs by sex at both the gene and liability levels, supporting a female protective effect while highlighting sex-specific patterns in coding variation. They also identify RNU2-2 as a noncoding contributor to ASD risk, expanding the landscape of ASD-associated variation beyond protein-coding genes.