|dc.description.abstract||Sex determination in mammals is regulated by the sex-determining region on the Y chromosome (SRY); the presence of SRY activates the male developmental pathway and suppresses the gene network necessary for female gonad development. Mutations in sex determination genes lead to various abnormal sexual phenotypes, including sex reversal syndrome in which the genetic and phenotypic sex do not match. Sex reversal syndrome has been reported in humans, mouse, and several domestic species. In horses, SRY-negative XY sex reversal syndrome has been well described and is caused by deletions on the Y chromosome. However, the molecular causes of the SRY-positive condition in horses and other mammals are not known.
This research investigated five horses affected with SRY-positive XY sex reversal syndrome. Sequencing of the coding exon region of the SRY gene in the five cases showed 99-100% alignment with the sequences of normal males. Genotyping of two closely related individuals with 46 normal male controls on an equine SNP50 Beadchip identified two statistically significant SNPs in a ~16 Mb region on the long arm of horse chromosome 3 (ECA3q). The region was analyzed using Gene Ontology (GO) and Gene Relationships Across Implicated Loci (GRAIL) to select functionally relevant candidate genes for sequencing. Further analysis of the entire horse genome was done through array comparative genomic hybridization (aCGH), which investigated possible structural rearrangements, such as copy number variants (CNVs). Deletions of olfactory receptor genes were detected on multiple chromosomes and confirmed through quantitative real-time PCR (qPCR). A homozygous deletion on ECA29 in a region containing genes of the aldo-keto reductase gene family, known to play a role in interconverting sex hormones between active forms and inactive forms, was discovered in two sex reversed animals. The findings were confirmed through qPCR and fluorescence in situ hybridization (FISH), and experiments to define the specific breakpoints of the deletion through PCR have been initiated.
This research represents the first systematic search in the horse genome for mutations and CNVs related to sex determination. The findings contribute to better understanding of the molecular mechanisms of sex determination in horses and other mammals, including humans.||en