Identification and functional analysis of novel disease genes for early-onset neurodevelopmental disorders

• Applicant: Dr. Frederike Leonie Harms
• Subject Area: Human Genetics
• Term: since 2024
• Project identifier: Deutsche Forschungsgemeinschaft (DFG) - Project number 537922834

Next-generation sequencing technologies have accelerated the discovery of novel disease genes for Mendelian disorders. However, up to 60% of patients remain genetically undiagnosed after routine diagnostics, which means that more gene-disease associations are likely to be discovered. With the identification of variants in so-called candidate genes, there is also an urgent need to distinguish disease-causing sequence variants from the many variants without any functional impact and to improve our understanding of pathophysiology.With my main research focus on the identification and validation of pathogenic variants in candidate genes for early-onset neurodevelopmental disorders, I aim to confirm the pathogenicity of variants in two candidate genes by functional studies and identify the genetic cause in patients without a molecular diagnosis using genome-wide sequencing methods within this project. I have identified CTU1, encoding the cytosolic thiouridylase subunit 1, which catalyzes thiolation of the wobble uridine of glutamine-, glutamate-, and lysine-tRNAs, as a novel autosomal recessive candidate gene for a neurodevelopmental disorder with aplasia of the olfactory bulb. To characterize the effects of the biallelic CTU1 missense variants, I will investigate the thiolation of the corresponding tRNAs and analyze affected pathways proteome-wide in patient-derived fibroblasts. To gain insight at the molecular level, I will study ATP and iron-sulfur cluster binding and protein-protein interactions of the CTU1 variant proteins. I have ascertained ten subjects with microcephaly and/or abnormalities of higher mental function carrying heterozygous (mostly de novo) loss-of-function variants in MSI1. MSI1 encodes the RNA-binding protein Musashi-1, which is important for self-renewal and proliferation of (neural) stem cells (NSCs). As MSI1 is specifically expressed in the developing brain, I will generate human induced pluripotent stem cells (hiPSCs) from two patients’ fibroblasts and correct the MSI1 variants by CRISPR/Cas9-mediated genome editing. hiNSCs will then be differentiated and used as a cell model to decipher the dysregulated pathways in wild-type and mutant hiNSCs by transcriptome- and proteome-wide analyses.To extend the knowledge of genes in which variants cause early-onset neurodevelopmental disorders, I will reanalyze existing trio whole-exome sequencing data from ten patients without a molecular diagnosis. In up to five remaining unsolved patients and healthy parents, I will then perform trio whole-genome sequencing. Variant analysis will focus on the exome, but will also include deep intronic variants, variants in the 5' and 3' untranslated regions, and genome-wide copy number and structural variants.The comprehensive data generated within this project will advance our knowledge not only of genes associated with early-onset neurodevelopmental disorders, but also of the underlying pathophysiology.

• DFG Programme: Research Grants