In vivo selection reveals combinatorial controls that define a critical exon in the spinal muscular atrophy genes
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TLDR
The results suggest that the evolutionary conserved weak 5' ss may serve as a mechanism to regulate exon 7 splicing under different physiological contexts, and are the first report in which a functional selection method has been applied to analyze the entire exon.Abstract:
Humans have two near identical copies of the survival of motor neuron (SMN) gene, SMN1 and SMN2. In spinal muscular atrophy (SMA), SMN2 is not able to compensate for the loss of SMN1 due to an inhibitory mutation at position 6 (C6U mutation in transcript) of exon 7. We have recently shown that C6U creates an extended inhibitory context (Exinct) that causes skipping of exon 7 in SMN2. Previous studies have shown that an exonic splicing enhancer associated with Tra2 (Tra2-ESE) is required for exon 7 inclusion in both SMN1 and SMN2. Here we describe the method of in vivo selection that determined the position-specific role of wild-type nucleotides within the entire exon 7. Our results confirmed the existence of Exinct and revealed the presence of an additional inhibitory tract (3'-Cluster) near the 3'-end of exon 7. We also demonstrate that a single nucleotide substitution at the last position of exon 7 improves the 5' splice site (ss) such that the presence of inhibitory elements (Exinct as well as the 3'-Cluster) and the absence of Tra2-ESE no longer determined exon 7 usage. Our results suggest that the evolutionary conserved weak 5' ss may serve as a mechanism to regulate exon 7 splicing under different physiological contexts. This is the first report in which a functional selection method has been applied to analyze the entire exon. This method offers unparallel advantage for determining the relative strength of splice sites, as well as for identifying the novel exonic cis-elements.read more
Citations
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The human splicing code reveals new insights into the genetic determinants of disease
Hui Yuan Xiong,Babak Alipanahi,Babak Alipanahi,Leo J. Lee,Leo J. Lee,Hannes Bretschneider,Daniele Merico,Ryan K. C. Yuen,Yimin Hua,Serge Gueroussov,Hamed S. Najafabadi,Hamed S. Najafabadi,Timothy P. Hughes,Quaid Morris,Yoseph Barash,Adrian R. Krainer,Nebojsa Jojic,Stephen W. Scherer,Benjamin J. Blencowe,Brendan J. Frey +19 more
TL;DR: A computational model is developed that scores how strongly genetic variants affect RNA splicing, a critical step in gene expression whose disruption contributes to many diseases, including cancers and neurological disorders, and provides insights into the role of aberrant splicing in disease.
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Alternative splicing and disease.
TL;DR: Well-studied examples that show how a disturbance of a fine-tuned balance of factors regulates splice site selection can cause human disease are discussed.
Journal ArticleDOI
Antisense Masking of an hnRNP A1/A2 Intronic Splicing Silencer Corrects SMN2 Splicing in Transgenic Mice
TL;DR: The results show that the high-resolution ASO-tiling approach can identify cis-elements that modulate splicing positively or negatively and highlight the therapeutic potential of some of these ASOs in the context of SMA.
Journal ArticleDOI
Splicing of a Critical Exon of Human Survival Motor Neuron Is Regulated by a Unique Silencer Element Located in the Last Intron
TL;DR: A novel inhibitory element located immediately downstream of the 5′ splice site in intron 7 is described, which underscores for the first time the profound impact of an evolutionarily nonconserved intronic element on SMN2 exon 7 splicing.
Journal ArticleDOI
Enhancement of SMN2 exon 7 inclusion by antisense oligonucleotides targeting the exon
TL;DR: A large number of ASOs with a 2′-O-methoxy-ethyl ribose (MOE) backbone that hybridize to different positions of SMN2 exon 7 increase full-length SMN protein levels, demonstrating that they do not interfere with mRNA export or translation, despite hybridizing to an exon.
References
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Journal ArticleDOI
Identification and characterization of a spinal muscular atrophy-determining gene
Suzie Lefebvre,Lydie Burglen,Sophie Reboullet,Olivier Clermont,Philippe Burlet,Louis Viollet,Bernard Bénichou,Corinne Cruaud,Philippe Millasseau,Massimo Zeviani,Denis Le Paslier,Jean Frézal,Daniel Cohen,Jean Weissenbach,Arnold Munnich,Judith Melki +15 more
TL;DR: The inverted duplication of a 500 kb element in normal chromosomes is described and the critical region is narrowed to 140 kb within the telomeric region, suggesting that this gene, termed the survival motor neuron (SMN) gene, is an SMA-determining gene.
Journal ArticleDOI
Mechanisms of Alternative Pre-Messenger RNA Splicing
TL;DR: This review describes what is currently known of the molecular mechanisms that control changes in splice site choice and starts with the best-characterized systems from the Drosophila sex determination pathway, and then describes the regulators of other systems about whose mechanisms there is some data.
Journal ArticleDOI
Listening to silence and understanding nonsense: exonic mutations that affect splicing
TL;DR: As the splicing mechanisms that depend on exonic signals are elucidated, new therapeutic approaches to treating certain genetic diseases can begin to be explored.
Journal ArticleDOI
ESEfinder: A web resource to identify exonic splicing enhancers.
TL;DR: ESEfinder (http://exon.cshl.edu/ESE/) is a web-based resource that facilitates rapid analysis of exon sequences to identify putative ESEs responsive to the human SR proteins SF2/ASF, SC35, SRp40 and SRp55, and to predict whether exonic mutations disrupt such elements.
Journal ArticleDOI
A single nucleotide in the SMN gene regulates splicing and is responsible for spinal muscular atrophy
TL;DR: The failure of SMN2 to fully compensate for SMN1 and protect from SMA is due to a nucleotide exchange (C/T) that attenuates activity of an exonic enhancer.
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