title: Insertion of expanded CAG trinucleotide repeat motifs into a yeast artificial chromosome containing the human Machado-Joseph disease gene.
abstract: Machado-Joseph disease or spinocerebellar ataxia 3 (SCA3) is a progressive neurodegenerative disorder caused by pathological expansion of a trinucleotide repeat motif present within exon 4 of the MJD1 gene. Previous attempts to create a transgenic animal model have failed to produce a neurological deficit truly representative of the disease phenotype. This appears to be the result of inappropriate expression of the mutant protein in neuronal populations generally spared in the disease state. Introduction of a human disease gene in the context of a yeast artificial chromosome clone containing endogenous regulatory elements would enhance the potential for correct tissue/cell-specific expression at physiological levels. We report the introduction of expanded CAG repeat motifs into a 250kb yeast artificial chromosome clone spanning the MJD1 locus using two rounds of homologous recombination. Transformants exhibited both expansions and contractions of the motif with alleles ranging in size from 48 to 84 repeat units. The availability of these clones for modelling of the disease in transgenic animals should allow elucidation of the role of repeat length in the phenotypic spectrum of the disease.
authors.affiliation: Hereditary Ataxia Research Group, Molecular Genetics, Division of Biomedical Sciences, Imperial College of Science Technology and Medicine, Sir Alexander Fleming Building, South Kensington Campus, London SW7 2AZ, UK. c.cemal@ic.ac.uk
authors.forename: C K
authors.initials: CK
authors.lastname: Cemal
authors.forename: C
authors.initials: C
authors.lastname: Huxley
authors.forename: S
authors.initials: S
authors.lastname: Chamberlain
mesh_terms: D000067699:Ataxin-3; D002874:Chromosome Mapping; D018244:Chromosomes, Artificial, Yeast; D004262:DNA Restriction Enzymes / Q000378:metabolism; D005819:Genetic Markers; D006801:Humans; D017827:Machado-Joseph Disease / Q000235:genetics; D008957:Models, Genetic; D016254:Mutagenesis, Insertional; D009419:Nerve Tissue Proteins / Q000235:genetics*; D009687:Nuclear Proteins; D010641:Phenotype; D011995:Recombination, Genetic; D012097:Repressor Proteins; D019680:Trinucleotide Repeat Expansion
chemical_list: D005819:Genetic Markers; D009419:Nerve Tissue Proteins; D009687:Nuclear Proteins; D012097:Repressor Proteins; D004262:DNA Restriction Enzymes; C092341:ATXN3 protein, human; D000067699:Ataxin-3