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PMID: 10567551 Published · ppublish English Journal Article

Evidence for substrate-specific requirement of the splicing factor U2AF(35) and for its function after polypyrimidine tract recognition by U2AF(65).

Molecular and cellular biology ·Vol. 19 ·No. 12 ·1999-12-00 ·Pages 8263-71

Guth S, Martínez C, Gaur RK, Valcárcel J

Abstract

U2 snRNP auxiliary factor (U2AF) promotes U2 snRNP binding to pre-mRNAs and consists of two subunits of 65 and 35 kDa, U2AF(65) and U2AF(35). U2AF(65) binds to the polypyrimidine (Py) tract upstream from the 3' splice site and plays a key role in assisting U2 snRNP recruitment. It has been proposed that U2AF(35) facilitates U2AF(65) binding through a network of protein-protein interactions with other splicing factors, but the requirement and function of U2AF(35) remain controversial. Here we show that recombinant U2AF(65) is sufficient to activate the splicing of two constitutively spliced pre-mRNAs in extracts that were chromatographically depleted of U2AF. In contrast, U2AF(65), U2AF(35), and the interaction between them are required for splicing of an immunoglobulin micro; pre-RNA containing an intron with a weak Py tract and a purine-rich exonic splicing enhancer. Remarkably, splicing activation by U2AF(35) occurs without changes in U2AF(65) cross-linking to the Py tract. These results reveal substrate-specific requirements for U2AF(35) and a novel function for this factor in pre-mRNA splicing.

MeSH Terms
Animals Beta-Globulins/genetics Cell Extracts DNA-Binding Proteins/genetics HeLa Cells Humans Immunoglobulin M/genetics Mice Nuclear Proteins Pyrimidines/metabolism RNA Precursors RNA Splicing Recombinant Fusion Proteins/metabolism Ribonucleoproteins/metabolism Splicing Factor U2AF Substrate Specificity Transcription Factors/genetics Viral Proteins
Chemicals
Beta-Globulins Cell Extracts DNA-Binding Proteins Immunoglobulin M MLTF protein, adenovirus Nuclear Proteins Pyrimidines RNA Precursors Recombinant Fusion Proteins Ribonucleoproteins Splicing Factor U2AF Transcription Factors U2AF1 protein, human U2AF2 protein, human Viral Proteins Zrsr1 protein, mouse
Authors & Affiliations
4 authors, click to expand affiliations / ORCID
Guth S
Gene Expression Programme, European Molecular Biology Laboratory, D-69117 Heidelberg, Germany.
Martínez C
Gaur R K
Valcárcel J
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42 references, click to expand
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Article Info
Journal
Molecular and cellular biology
Abbr.
Mol Cell Biol
ISSN
0270-7306
DOI
10.1128/MCB.19.12.8263
Published
1999-12-00
Pages
8263-71
Language
English
Region
United States
NLM ID
8109087
PMCID
PMC84910
Subset
IM
External Links
PubMed source DOI: 10.1128/MCB.19.12.8263 PMC Full Text View journal details
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