Repression of alpha-actinin SM exon splicing by assisted binding of PTB to the polypyrimidine tract

Polypyrimidine tract binding protein (PTB) acts as a regulatory repressor of a large number of alternatively spliced exons, often requiring multiple binding sites in order to repress splicing. In one case, cooperative binding of PTB has been shown to accompany repression. The SM exon of the alpha-ac...

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Bibliographic Details
Published in:RNA (Cambridge) 2007-08, Vol.13 (8), p.1214-1223
Main Authors: Matlin, Arianne J, Southby, Justine, Gooding, Clare, Smith, Christopher W J
Format: Article
Language:English
Subjects:
Actinin - genetics
Actinin - metabolism
Base Sequence
Exons
HeLa Cells
Humans
Introns
Molecular Sequence Data
Nuclear Proteins - metabolism
Polypyrimidine Tract-Binding Protein - metabolism
Ribonucleoproteins - metabolism
RNA Splicing
Splicing Factor U2AF
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Summary:Polypyrimidine tract binding protein (PTB) acts as a regulatory repressor of a large number of alternatively spliced exons, often requiring multiple binding sites in order to repress splicing. In one case, cooperative binding of PTB has been shown to accompany repression. The SM exon of the alpha-actinin pre-mRNA is also repressed by PTB, leading to inclusion of the alternative upstream NM exon. The SM exon has a distant branch point located 386 nt upstream of the exon with an adjacent 26 nucleotide pyrimidine tract. Here we have analyzed PTB binding to the NM and SM exon region of the alpha-actinin pre-mRNA. We find that three regions of the intron bind PTB, including the 3' end of the polypyrimidine tract (PPT) and two additional regions between the PPT and the SM exon. The downstream PTB binding sites are essential for full repression and promote binding of PTB to the PPT with a consequent reduction in U2AF(65) binding. Our results are consistent with a repressive mechanism in which cooperative binding of PTB to the PPT competes with binding of U2AF(65), thereby specifically blocking splicing of the SM exon.
ISSN:1355-8382
1469-9001
DOI:10.1261/rna.219607