title: Allele-specific silencing of mutant Ataxin-7 in SCA7 patient-derived fibroblasts.
abstract: Polyglutamine (polyQ) disorders are inherited neurodegenerative conditions defined by a common pathogenic CAG repeat expansion leading to a toxic gain-of-function of the mutant protein. Consequences of this toxicity include activation of heat-shock proteins (HSPs), impairment of the ubiquitin-proteasome pathway and transcriptional dysregulation. Several studies in animal models have shown that reducing levels of toxic protein using small RNAs would be an ideal therapeutic approach for such disorders, including spinocerebellar ataxia-7 (SCA7). However, testing such RNA interference (RNAi) effectors in genetically appropriate patient cell lines with a disease-relevant phenotype has yet to be explored. Here, we have used primary adult dermal fibroblasts from SCA7 patients and controls to assess the endogenous allele-specific silencing of ataxin-7 by two distinct siRNAs. We further identified altered expression of two disease-relevant transcripts in SCA7 patient cells: a twofold increase in levels of the HSP DNAJA1 and a twofold decrease in levels of the de-ubiquitinating enzyme, UCHL1. After siRNA treatment, the expression of both genes was restored towards normal levels. To our knowledge, this is the first time that allele-specific silencing of mutant ataxin-7, targeting a common SNP, has been demonstrated in patient cells. These findings highlight the advantage of an allele-specific RNAi-based therapeutic approach, and indicate the value of primary patient-derived cells as useful models for mechanistic studies and for measuring efficacy of RNAi effectors on a patient-to-patient basis in the polyQ diseases.
authors.affiliation: Department of Physiology, Anatomy and Genetics, University of Oxford, Oxford, UK.
authors.forename: Janine
authors.initials: J
authors.lastname: Scholefield
authors.affiliation: 1] Department of Physiology, Anatomy and Genetics, University of Oxford, Oxford, UK [2] Division of Human Genetics, Department of Clinical Laboratory Sciences, Institute of Infectious Disease and Molecular Medicine, University of Cape Town, Cape Town, South Africa.
authors.forename: Lauren
authors.initials: L
authors.lastname: Watson
authors.affiliation: Division of Human Genetics, Department of Clinical Laboratory Sciences, Institute of Infectious Disease and Molecular Medicine, University of Cape Town, Cape Town, South Africa.
authors.forename: Danielle
authors.initials: D
authors.lastname: Smith
authors.affiliation: Division of Human Genetics, Department of Clinical Laboratory Sciences, Institute of Infectious Disease and Molecular Medicine, University of Cape Town, Cape Town, South Africa.
authors.forename: Jacquie
authors.initials: J
authors.lastname: Greenberg
authors.affiliation: 1] Department of Physiology, Anatomy and Genetics, University of Oxford, Oxford, UK [2] Division of Human Genetics, Department of Clinical Laboratory Sciences, Institute of Infectious Disease and Molecular Medicine, University of Cape Town, Cape Town, South Africa.
authors.forename: Matthew J A
authors.initials: MJ
authors.lastname: Wood
mesh_terms: D000483:Alleles; D000067719:Ataxin-7; D002460:Cell Line; D005347:Fibroblasts / Q000378:metabolism*; D020868:Gene Silencing; D060005:Genotyping Techniques; D050956:HSP40 Heat-Shock Proteins / Q000235:genetics / Q000378:metabolism; D006801:Humans; D009419:Nerve Tissue Proteins / Q000235:genetics* / Q000378:metabolism; D019636:Neurodegenerative Diseases / Q000235:genetics / Q000628:therapy; D010641:Phenotype; D020641:Polymorphism, Single Nucleotide; D034622:RNA Interference; D034741:RNA, Small Interfering / Q000235:genetics / Q000378:metabolism; D043222:Ubiquitin Thiolesterase / Q000235:genetics / Q000378:metabolism
chemical_list: C000593829:ATXN7 protein, human; D000067719:Ataxin-7; C107481:DNAJA1 protein, human; D050956:HSP40 Heat-Shock Proteins; D009419:Nerve Tissue Proteins; D034741:RNA, Small Interfering; C482860:UCHL1 protein, human; D043222:Ubiquitin Thiolesterase