Elevated cholesterol in ATAD3 mutants is a compensatory mechanism that leads to membrane cholesterol aggregation.

Aberrant cholesterol metabolism causes neurological disease and neurodegeneration, and mitochondria have been linked to perturbed cholesterol homeostasis via the study of pathological mutations in the ATAD3 gene cluster. However, whether the cholesterol changes were compensatory or contributory to the disorder was unclear, and the effects on cell membranes and the wider cell were also unknown. Using patient-derived cells, we show that cholesterol perturbation is a conserved feature of pathological ATAD3 variants that is accompanied by an expanded lysosome population containing membrane whorls characteristic of lysosomal storage diseases.

CRISPR/Cas9-mediated tissue-specific knockout and cDNA rescue using sgRNAs that target exon-intron junctions in .

In this protocol, we take CRISPR/Cas9 and Gal4/UAS approaches to achieve tissue-specific knockout in parallel with rescue of the knockout by cDNA expression in . We demonstrate that guide RNAs targeting the exon-intron junction of target genes cleave the genomic locus of the genes, but not transgenes, in a tissue where Gal4 drives Cas9 expression. The efficiency of this approach enables the determination of pathogenicity of disease-associated variants in human genes in a tissue-specific manner in .

Bi-allelic variants in OGDHL cause a neurodevelopmental spectrum disease featuring epilepsy, hearing loss, visual impairment, and ataxia.

The 2-oxoglutarate dehydrogenase-like (OGDHL) protein is a rate-limiting enzyme in the Krebs cycle that plays a pivotal role in mitochondrial metabolism. OGDHL expression is restricted mainly to the brain in humans. Here, we report nine individuals from eight unrelated families carrying bi-allelic variants in OGDHL with a range of neurological and neurodevelopmental phenotypes including epilepsy, hearing loss, visual impairment, gait ataxia, microcephaly, and hypoplastic corpus callosum.