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Genes Dev. 2002 Nov 1;16(21):2778-91.

5' exon interactions within the human spliceosome establish a framework for exon junction complex structure and assembly.

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  • 1Howard Hughes Medical Institute, Department of Biochemistry, Brandeis University, Massachusetts 02454, USA.

Abstract

A general consequence of pre-mRNA splicing is the stable deposition of several proteins 20-24 nucleotides (nt) upstream of exon-exon junctions on spliced mRNAs. This exon junction complex (EJC) contains factors involved in mRNA export, cytoplasmic localization, and nonsense-mediated mRNA decay. Here we probed the mechanism and timing of EJC assembly. Over the course of splicing, the 5' exon is subject to numerous dynamic protein-RNA interactions involving at least nine distinct polypeptides. Within the fully assembled spliceosome, these interactions afford protection to the last 25-27 nt of the 5' exon intermediate. Coincident with exon ligation, interactions at the 3' end of the 5' exon disappear, and new species associate with position -24. Mass spectrometry and Western blotting of purified H, C, and mRNP complexes revealed that at least one EJC component, REF/Aly, can interact with pre-mRNA prior to spliceosome assembly, whereas Y14, Magoh, RNPS1, UAP56, and SRm160 are found in intermediate-containing spliceosomes. Upon exon ligation, association of RNPS1, UAP56, and SRm160 is destabilized. In contrast, REF/Aly, Y14, and Magoh remain stably bound to spliced mRNA, indicating that these three proteins are components of the EJC core.

PMID:
12414731
[PubMed - indexed for MEDLINE]
PMCID:
PMC187475
Free PMC Article
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