Journal
NEUROBIOLOGY OF DISEASE
Volume 43, Issue 3, Pages 533-542Publisher
ACADEMIC PRESS INC ELSEVIER SCIENCE
DOI: 10.1016/j.nbd.2011.04.016
Keywords
Neurodegeneration; Autosomal dominant disorder; Spinocerebellar ataxia; Polyglutamine; Cav2.1; CACNA1A; Calcium channel; Alternative splicing; RNA interference; MicroRNA; siRNA; shRNA; Allele specific silencing; Gene therapy
Categories
Funding
- National Institutes of Health [NS-50210, NS-072229]
- Fauver Family Ataxia Research Fund
- University of Michigan Neurology Department
- National Science Council of Taiwan [NSC96-2314-B-010-036-MY3, NSC98-2917-I-010-106]
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Spinocerebellar ataxia type 6 (SCA6) is an inherited neurodegenerative disease caused by a polyglutamine (polyQ) expansion in the Ca(v)2.1 voltage-gated calcium channel subunit (CACNA1A). There is currently no treatment for this debilitating disorder and thus a pressing need to develop preventative therapies. RNA interference (RNAi) has proven effective at halting disease progression in several models of spinocerebellar ataxia (SCA), including SCA types 1 and 3. However, in SCA6 and other dominantly inherited neurodegenerative disorders, RNAi-based strategies that selectively suppress expression of mutant alleles may be required. Using a Ca(v)2.1 mini-gene reporter system, we found that pathogenic CAG expansions in Ca(v)2.1 enhance splicing activity at the 3'end of the transcript, leading to a CAG repeat length-dependent increase in the levels of a polyQ-encoding Ca(v)2.1 mRNA splice isoform and the resultant disease protein. Taking advantage of this molecular phenomenon, we developed a novel splice isoform-specific (SIS)-RNAi strategy that selectively targets the polyQ-encoding Ca(v)2.1 splice variant. Selective suppression of transiently expressed and endogenous polyQ-encoding Ca(v)2.1 splice variants was achieved in a variety of cell-based models including a human neuronal cell line, using a new artificial miRNA-like delivery system. Moreover, the efficacy of gene silencing correlated with effective intracellular recognition and processing of SIS-RNAi miRNA mimics. These results lend support to the preclinical development of SIS-RNAi as a potential therapy for SCA6 and other dominantly inherited diseases. (C) 2011 Elsevier Inc. All rights reserved.
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