Recurrent de novo variants in the spliceosomal factor CRNKL1 are associated with severe microcephaly and pontocerebellar hypoplasia with seizures.

Splicing is a complex process that is required to create the transcriptomic diversity needed for specialized functions in higher eukaryotes. The spliceosome contains more than 100 proteins and RNA molecules, which coordinate this dynamic process. Despite the ubiquity of splicing, pathogenic variants in spliceosomal components often cause a tissue-specific phenotype, hinting at further complexities that are not yet fully understood. We have identified a cohort of ten families with de novo missense variants in a spliceosomal component, CRNKL1, where nine individuals harbor one of two missense variants that both affect the same amino acid, Arg267. All affected individuals share a common and specific phenotype: profound pre- and post-natal microcephaly, with pontocerebellar hypoplasia, seizures, and severe intellectual disability. Microinjection of mRNA encoding mutant Crnkl1 into a zebrafish model caused a severe lack of brain development accompanied by a significant reduction in proliferating cells and widespread cellular stress, as indicated by p53 staining. RNA sequencing analysis of injected zebrafish embryos showed broad transcriptomic changes, with altered expression of neuronal and cell cycle genes. Taking these results together, we have identified CRNKL1 as a disease-associated gene and demonstrate the requirement for this protein in brain development. Our findings contribute to a growing disease cluster, where associated components act at the same spliceosomal stage and cause a severe neurological phenotype, suggesting a more intricate role for these spliceosomal subcomplexes than previously thought.