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PMID: 7935481 Published · ppublish English Journal Article

SR proteins promote the first specific recognition of Pre-mRNA and are present together with the U1 small nuclear ribonucleoprotein particle in a general splicing enhancer complex.

Molecular and cellular biology ·Vol. 14 ·No. 11 ·1994-11-00 ·Pages 7670-82

Staknis D, Reed R

Abstract

We show that addition of SR proteins to in vitro splicing extracts results in a significant increase in assembly of the earliest prespliceosomal complex E and a corresponding decrease in assembly of the heterogeneous nuclear ribonucleoprotein (hnRNP) complex H. In addition, SR proteins promote formation of the E5' and E3' complexes that assemble on RNAs containing only 5' and 3' splice sites, respectively. We conclude that SR proteins promote the earliest specific recognition of both the 5' and 3' splice sites and are limiting for this function in HeLa nuclear extracts. Using UV cross-linking, we demonstrate specific, splice site-dependent RNA-protein interactions of SR proteins in the E, E5', and E3' complexes. SR proteins do not UV cross-link in the H complex, and conversely, hnRNP cross-linking is largely excluded from the E-type complexes. We also show that a discrete complex resembling the E5' complex assembles on both purine-rich and non-purine-rich exonic splicing enhancers. This complex, which we have designated the Enhancer complex, contains U1 small nuclear RNP (snRNP) and is associated with different SR protein family members, depending on the sequence of the enhancer. We propose that both downstream 5' splice site enhancers and exonic enhancers function by establishing a network of pre-mRNA-protein and protein-protein interactions involving U1 snRNP, SR proteins, and U2AF that is similar to the interactions that bring the 5' and 3' splice sites together in the E complex.

MeSH Terms
Base Sequence Binding Sites Cross-Linking Reagents DNA/genetics Enhancer Elements, Genetic HeLa Cells Humans Models, Biological Molecular Sequence Data RNA Precursors/genetics,metabolism,radiation effects RNA Splicing/genetics RNA-Binding Proteins/metabolism,radiation effects Ribonucleoprotein, U1 Small Nuclear/genetics,metabolism Spliceosomes/metabolism Ultraviolet Rays
Chemicals
Cross-Linking Reagents RNA Precursors RNA-Binding Proteins Ribonucleoprotein, U1 Small Nuclear DNA
Authors & Affiliations
2 authors, click to expand affiliations / ORCID
Staknis D
Department of Cell Biology, Harvard Medical School, Boston, Massachusetts 02115.
Reed R
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Article Info
Journal
Molecular and cellular biology
Abbr.
Mol Cell Biol
ISSN
0270-7306
DOI
10.1128/mcb.14.11.7670-7682.1994
Published
1994-11-00
Pages
7670-82
Language
English
Region
United States
NLM ID
8109087
PMCID
PMC359303
Subset
IM
External Links
PubMed source DOI: 10.1128/mcb.14.11.7670-7682.1994 PMC Full Text View journal details
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