A multi-exon-skipping detection assay reveals surprising diversity of splice isoforms of spinal muscular atrophy genes
Humans have two near identical copies of Survival Motor Neuron gene: SMN1 and SMN2. Loss of SMN1 coupled with the predominant skipping of SMN2 exon 7 causes spinal muscular atrophy (SMA), a neurodegenerative disease. SMA patient cells devoid of SMN1 provide a powerful system to examine splicing patt...
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| Published in: | PloS one Vol. 7; no. 11; p. e49595 |
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| Main Authors: | , , , |
| Format: | Journal Article |
| Language: | English |
| Published: |
United States
Public Library of Science
19-11-2012
Public Library of Science (PLoS) |
| Subjects: | |
| Online Access: | Get full text |
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| Summary: | Humans have two near identical copies of Survival Motor Neuron gene: SMN1 and SMN2. Loss of SMN1 coupled with the predominant skipping of SMN2 exon 7 causes spinal muscular atrophy (SMA), a neurodegenerative disease. SMA patient cells devoid of SMN1 provide a powerful system to examine splicing pattern of various SMN2 exons. Until now, similar system to examine splicing of SMN1 exons was unavailable. We have recently screened several patient cell lines derived from various diseases, including SMA, Alzheimer's disease, Parkinson's disease and Batten disease. Here we report a Batten disease cell line that lacks functional SMN2, as an ideal system to examine pre-mRNA splicing of SMN1. We employ a multiple-exon-skipping detection assay (MESDA) to capture simultaneously skipping of multiple exons. Our results show surprising diversity of splice isoforms and reveal novel splicing events that include skipping of exon 4 and co-skipping of three adjacent exons of SMN. Contrary to the general belief, MESDA captured oxidative-stress induced skipping of SMN1 exon 5 in several cell types, including non-neuronal cells. We further demonstrate that the predominant SMN2 exon 7 skipping induced by oxidative stress is modulated by a combinatorial control that includes promoter sequence, endogenous context, and the weak splice sites. We also show that an 8-mer antisense oligonucleotide blocking a recently described GC-rich sequence prevents SMN2 exon 7 skipping under the conditions of oxidative stress. Our findings bring new insight into splicing regulation of an essential housekeeping gene linked to neurodegeneration and infant mortality. |
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| Bibliography: | ObjectType-Article-1 SourceType-Scholarly Journals-1 ObjectType-Feature-2 content type line 23 Competing Interests: The authors have the following interests. Iowa State University holds the intellectual property rights on ASO (3UP8) described in this study (Patent# US 20110269820 A1). Therefore, authors (RNS and NNS) and Iowa State University could potentially benefit from any future commercial exploitation of the above-mentioned ASO. There are no further patents, products in development or marketed products to declare. This does not alter the authors' adherence to all the PLOS ONE policies on sharing data and materials. Conceived and designed the experiments: RNS NNS JS SJR. Performed the experiments: NNS JS SJR. Analyzed the data: RNS NNS JS SJR. Contributed reagents/materials/analysis tools: RNS. Wrote the paper: RNS NNS. |
| ISSN: | 1932-6203 1932-6203 |
| DOI: | 10.1371/journal.pone.0049595 |