Occhiodoro T, Anson D S
Lysosomal Diseases Research Unit, Department of Chemical Pathology, Women's and Children's Hospital, 72 King William Road, North Adelaide, South Australia, 5006, Australia.
Mamm Genome. 1996 Apr;7(4):271-4. doi: 10.1007/s003359900081.
Fucosidosis is a lysosomal storage disorder caused by deficiency of alpha-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 alpha-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 alpha-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 alpha-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.
岩藻糖苷贮积症是一种由α-L-岩藻糖苷酶缺乏引起的溶酶体贮积病。在英国激飞猎犬群体中存在一种生化和临床特征明确的岩藻糖苷贮积症犬模型。为了便于将其用作显示神经症状的溶酶体贮积病基因治疗和酶替代疗法的模型，开展了犬α-L-岩藻糖苷酶cDNA的分离工作。犬岩藻糖苷酶的核苷酸序列和预测的氨基酸序列与人类和大鼠序列均显示出高度的同一性。通过Northern印迹分析发现，与正常犬相比，患岩藻糖苷贮积症的犬α-L-岩藻糖苷酶mRNA水平大幅降低。对cDNA产生的产物进行直接PCR测序表明，患病犬的mRNA中有一个14 bp的缺失。这种缺失产生了移码突变，并在氨基酸位置152处引入了一个过早的翻译终止密码子，且已证明其对应于犬α-L-岩藻糖苷酶基因外显子1最后14个碱基对的缺失。现已对该群体中犬的基因组DNA进行了基于PCR的快速突变筛查，能够检测出携带者和纯合子。
