Inheritance, mapping and investigation of the causative mutation for equine cerebellar abiotrophy

Equine cerebellar abiotrophy (CA) is a neurodegenerative disease that is found almost exclusively in the Arabian breed. Horses affected with CA generally begin to exhibit signs of the disease between six weeks and four months of age, although onset is variable. These signs include progressive ataxia, intention head tremor, and a lack of menace response. While the disease itself is not fatal, most affected horses are euthanized, as they are unusable as performance or breeding animals, and often uncoordinated enough to be handled safely.;CA is caused by a post-natal degeneration of the Purkinje cells, the largest neurons of the cerebellum, which control motor coordination and form connections between the molecular and granular layers. The degeneration of Purkinje cells is usually followed by a concurrent loss of granular neurons, leading to a disorganization of the molecular and granular layers.;Case reports of CA in horses have been published as early as the 1960's. While CA has long been suspected to be inherited as an autosomal recessive trait, the rarity of cases made genetic analysis difficult. Therefore, no research exists that attempts to identify the mode of inheritance of the disease or to elucidate the genetic mechanisms that cause it.;The goals of this research were to identify the mode of inheritance of CA, map the trait to a region of the equine genome, and to use sequencing and gene expression techniques to find the likely causative mutation of the disease. To establish the mode of inheritance of CA, complex segregation analysis was performed on 804 Arabian horses segregating for the disease. This analysis demonstrated with statistical significance that CA is influence by a single locus of large effect, and estimated transmission probabilities of the CA allele fell within the expected values for an autosomal recessive locus.;Linkage mapping of CA was performed using a whole genome scan of 335 microsatellite markers on 32 horses from four paternal half-sibling families segregating for CA. Linkage was identified between CA and microsatellite markers on ECA2. Haplotyping and homozygosity analysis of affected horses further refined the map position of CA to a 142 Kb region which contains four annotated genes in the horse.;To identify a possible causative mutation for CA in this region, all four genes were completely sequenced in affected and unaffected horses. A total of 22 polymorphisms were identified that segregated with CA. One of these, the ECA2:13074277 SNP, was found exclusively in Arabian horses. The ECA2:13074277 SNP is located in an exon of the TOE1 gene but causes a relatively innocuous amino acid change. However, TOE1 overlaps at its 5' end with the 5' end of the MUTYH gene, a DNA glycosylase responsible for oxidative damage repair in Purkinje cells. The ECA2:13074277 SNP is approximately 1200 base pairs upstream of MUTYH and immediately adjacent to a possible binding site for the transcription factor GATA-2, suggesting a possible role in regulation of MUTYH. A qPCR analysis of MUTYH in affected and control horses showed that MUTYH expression is negatively altered in affected horses. The ECA2:13074277 SNP is therefore the most likely causative mutation for CA, and acts by down-regulating the expression of a DNA repair gene, leading to apoptosis of Purkinje cells.