Investigating amyotrophic lateral sclerosis candidate genes
thesisposted on 29.03.2022, 03:43 by Sandrine Kim Kiow Chan Moi Fat
Amyotrophic lateral sclerosis (ALS) is a fatal neurodegenerative disease of motor neurons.Approximately 10% of cases are classiffed as familial and 90% sporadic, although a proportion of sporadic cases have an unrecognised family history. This thesis aimed o identify novel ALS genes from familial and sporadic cases. Whole-exome and whole genome sequencing analysis identified 22 candidate gene variants in an ALS family,which were investigated using an in silico analysis pipeline including protein predictions,conservation, genic tolerance, tissue expression and gene function. Three top priority variants, in CYB5R3, DCAF7, and SAV1, were assessed using in vitro toxicity and localisation assays, with one candidate, CYB5R3, displaying a differential cytoplasmic protein expression pattern. Whole-genome data from 635 sporadic ALS patients were interrogated for ALS candidate genes (n = 21), and ten novel variants were identified, including a novel CYB5R3 variant, which rated strongly from in silico tools. Lastly, this project sought to develop a CRISPR-Cas9 protocol to generate ALS cell models for future use in the in vitro analysis pipeline. Overall, a combination of genetic, in silico and in vitro pipelines were used to identify candidate genes in an ALS kindred and apparently sporadic patients. Novel ALS genes will further our understanding of disease biology and contribute to development of diagnostics and treatments.