Recurrent de novo variants in the spliceosomal factor CRNKL1 cause severe microcephaly and pontocerebellar hypoplasia with seizures [HeLa]

Splicing is a complex process, required to create the transcriptomic diversity needed for specialised functions in higher eukaryotes. The spliceosome contains more than 100 proteins and RNA molecules which coordinate this dynamic process, and interestingly, despite the ubiquity of splicing, pathogenic variants in spliceosomal components often cause a tissue-specific phenotype, hinting at further complexities which 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 harbour one of two missense variants that both affect the same amino acid, p.Arg267. All patients share a common and specific phenotype: profound pre- and post-natal microcephaly, with pontocerebellar hypoplasia, seizures and severe intellectual disability. Microinjection of mutant crnkl1 into a zebrafish model caused significant lack of brain development accompanied by a significant reduction in proliferating cells and widespread cellular stress, as indicated by p53 staining. RNA-seq analysis of injected zebrafish embryos showed widespread transcriptomic changes, with altered expression of neuronal and cell cycle genes. Together, we have identified CRNKL1 as a novel disease gene and demonstrate the requirement for this protein in brain development and functioning. Our findings contribute to a growing disease cluster, where components act at the same spliceosomal stage and cause a brain-specific but severe phenotype, suggesting a more intricate role for these spliceosomal subcomplexes than previously thought.