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CRISPR gene editing and inducible pluripotent stem cell neuronal disease modelling for rare disease diagnosis: EMHM1 genetic variant analysis in Kleefstra Syndrome

ABSTRACT: Over 400 million people worldwide are living with a rare disease, with genetic variants determining 80% of cases. Next Generation Sequencing identifies potential disease causative genetic variants, however many of these are classified as variants of uncertain significance (VUS). Each VUS requires functional validation as pathogenic or benign in disease pathology in specialist laboratories creating major delays in patient diagnosis. In this study we test a rapid genetic variant assessment pipeline using an EHMT1 (Euchromatin histone methyltransferase 1; EHMT1 p.Gln1144Ter) genetic variant that is pathogenic for Kleefstra Syndrome. Precise CRISPR homology directed (HDR) gene editing introduced the single nucleotide genetic variant in iPS cells and EHMT1_SNV cell clones were rapidly identified with amplicon sequencing. Induction of neural differentiation and RNA sequencing determined differences in differentiation at the gene and transcription factor level. The applied CRISPR HDR methodology was rapid and reliable for the introduction of SNVs in iPSCs for subsequent neuronal cell differentiation. Key features of Kleefstra Syndrome were identified, with involvement of key transcription factors REST and SP1 in disease mechanisms. This study indicates that precise iPSC gene editing and changes in disease modelling pathways can contribute to disease diagnosis and understanding of mechanisms.

ORGANISM(S): Homo sapiens

PROVIDER: GSE178646 | GEO | 2021/06/23

REPOSITORIES: GEO

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