Huntington’s disease investigated using sheep as a model

Abstract

Huntington’s disease (HD) is a debilitating late-onset neurodegenerative disorder characterised by symptoms crossing motor, cognitive and behavioural faculties. Its cause is an expanded polyglutamine coding repeat within exon 1 of the IT15 gene. Despite isolation of the mutation more than 20 years ago, the molecular mechanisms underlying pathogenesis in HD remain poorly understood and there is no effective treatment that can prevent or delay its onset or progression. The aim of the research described in this thesis was to investigate HD using sheep as a model. Our laboratory previously generated a transgenic HD sheep (OVT73) in order to provide a large mammalian model for investigating the natural history of HD including its presymptomatic phase and for therapeutic testing. The results of this thesis describe molecular analyses of the OVT73 sheep. Candidate gene expression studies revealed changes in mRNA levels of several end-stage HD marker genes in the 6 month old striatum. In addition, RNA-Seq analysis of motor cortex tissue led to the generation of an annotated sheep brain transcriptome. Metabolite studies also revealed OVT73 specific differences. A number of fatty acids and amino acids were differentially abundant in cerebellum and liver tissue from 5 year old OVT73 sheep. Moreover, striking transgene-specific differences in the correlation of abundances for metabolite pairs were identified which suggested tightened co-regulation of metabolite abundance in the 5 year old OVT73 samples. Metabolite signatures were identified which discriminated between OVT73 and wild-type samples of 5 year old cerebellum and liver. Together these findings provide insight into metabolic dysfunction in HD. Finally in this research, the insertion of expanded CAG repeats into the endogenous huntingtin gene of a sheep fibroblast cell line mediated by zinc-finger nucleases constituted the first step towards making a second generation HD sheep; a knock-in model. The ability to investigate HD in large animals which accurately recapitulate the genetic basis of HD may provide new insight into the molecular mechanism of HD, particularly the early presymptomatic stage for which little is currently known. In turn, this may provide unique opportunities for the development of disease-modifying therapies and their progress to the clinic.