Identifying SHROOM4 as a novel X-linked susceptibility gene for cerebral palsy in Chinese males

Cerebral palsy (CP) is a leading cause of childhood motor disability with a notable male predominance, suggesting that X-linked genetic factors may contribute to CP susceptibility. Although SHROOM4 has been implicated in several neurodevelopmental disorders, its role in CP remains unclear. This study aimed to investigate the contribution of SHROOM4 variants to male CP susceptibility. Whole-exome sequencing was performed in 1,010 Chinese male patients with sporadic CP and 1,014 male controls. Association analysis focused on common variants and haplotypes within SHROOM4 . Rare SHROOM4 variants identified in CP patients were further validated and characterized using qPCR, immunofluorescence, western blotting and CRISPR/Cas9-mediated knockout cell lines. A common T-A-G haplotype comprising rs2873098, rs2295544 and rs2295543 in SHROOM4 was significantly associated with male CP susceptibility (OR = 6.091, Pc = 3.26E-07) and was enriched in CP patients with intrauterine growth restriction. Additionally, a rare nonsense variant, c.C2050T (p.Arg684*), was identified in a patient presenting with spastic CP and intellectual disability. Functional analyses showed that p.Arg684* and a population-derived frameshift variant (p.Glu1140fs*42) were associated with reduced SHROOM4 transcript abundance, consistent with NMD-mediated transcript reduction, while residual mutant transcripts produced detectable truncated proteins with variant-specific effects on protein stability, subcellular localization, and actin cytoskeletal organization. Our findings support SHROOM4 as an X-linked susceptibility gene associated with male CP, suggesting that both common haplotypes and rare SHROOM4 variants may contribute to the genetic susceptibility to CP. These results expand our current understanding of the genetic architecture of CP and highlight cytoskeletal regulation as a potentially relevant mechanism associated with SHROOM4 variants.