Title

In vivo selection reveals combinatorial controls that define a critical exon in the spinal muscular atrophy genes

UMMS Affiliation

Department of Medicine

Publication Date

7-24-2004

Document Type

Article

Subjects

Base Sequence; Cyclic AMP Response Element-Binding Protein; *Exons; Guanosine; Humans; Molecular Sequence Data; Muscular Atrophy, Spinal; Nerve Tissue Proteins; RNA-Binding Proteins; Selection (Genetics)

Disciplines

Genetics and Genomics | Life Sciences | Medicine and Health Sciences

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.

Rights and Permissions

Citation: RNA. 2004 Aug;10(8):1291-305. Link to article on publisher's site

DOI of Published Version

10.1261/rna.7580704

Related Resources

Link to Article in PubMed

Journal/Book/Conference Title

RNA (New York, N.Y.)

PubMed ID

15272122