title: Splice isoform-specific suppression of the Cav2.1 variant underlying spinocerebellar ataxia type 6.
abstract: 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.
authors.affiliation: Institute of Neuroscience, School of Life Sciences, National Yang-Ming University, Taipei, Taiwan.
authors.forename: Wei-Ling
authors.initials: WL
authors.lastname: Tsou
authors.forename: Bing-Wen
authors.initials: BW
authors.lastname: Soong
authors.forename: Henry L
authors.initials: HL
authors.lastname: Paulson
authors.forename: Edgardo
authors.initials: E
authors.lastname: Rodríguez-Lebrón
mesh_terms: D017398:Alternative Splicing / Q000235:genetics*; D015220:Calcium Channels / Q000235:genetics*; D020864:Calcium Channels, N-Type / Q000235:genetics*; D045744:Cell Line, Tumor; D015316:Genetic Therapy / Q000379:methods*; D014644:Genetic Variation / Q000235:genetics*; D006801:Humans; D009447:Neuroblastoma; D020033:Protein Isoforms / Q000096:biosynthesis / Q000235:genetics; D012326:RNA Splicing / Q000235:genetics; D012333:RNA, Messenger / Q000096:biosynthesis; D012091:Repetitive Sequences, Nucleic Acid / Q000235:genetics; D020754:Spinocerebellar Ataxias / Q000235:genetics* / Q000628:therapy*; D014158:Transcription, Genetic / Q000235:genetics
chemical_list: D015220:Calcium Channels; D020864:Calcium Channels, N-Type; D020033:Protein Isoforms; D012333:RNA, Messenger; C105656:CACNA1A protein, human