Isolation of the canine α- L -fucosidase cDNA and definition of the fucosidosis mutation in English Springer Spaniels

Fucosidosis is a lysosomal storage disorder caused by deficiency of α-L-fucosidase. A biochemically and clinically well characterized canine model of fucosidosis exists in a colony of English Springer Spaniels. To facilitate its use as a model for gene therapy and enzyme replacement therapy in lysosomal storage disorders displaying neurological symptoms, isolation of the canine a-L-fucosidase cDNA was undertaken. Both the nucleotide sequence and the predicted amino acid sequence of canine fucosidase show high levels of identity with the human and rat sequences. Fucosidosis dogs were found to have a greatly reduced level of α-L-fucosidase mRNA when compared with normal dogs by Northern blot analysis. Direct PCR sequencing of products generated from cDNA demonstrated a 14-bp deletion in mRNA from affected dogs. This deletion creates a frameshift mutation and introduces a premature translation termination codon at amino acid position 152 and was shown to correspond to a deletion of the last 14 base pairs of exon 1 of the canine α-L-fucosidase gene. Rapid PCR-based screening for the mutation has now been performed on genomic DNA from dogs within the colony, enabling detection of both carriers and homozygotes.