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PMID: 8112302 Published · ppublish English Journal Article Research Support, Non-U.S. Gov't

The splicing factor PRP2, a putative RNA helicase, interacts directly with pre-mRNA.

The EMBO journal ·Vol. 13 ·No. 4 ·1994-02-15 ·Pages 888-97

Teigelkamp S, McGarvey M, Plumpton M, Beggs JD

Abstract

The RNA helicase-like splicing factor PRP2 interacts only transiently with spliceosomes. To facilitate analysis of interactions of PRP2 with spliceosomal components, PRP2 protein was stalled in splicing complexes by two different methods. A dominant negative mutant form of PRP2 protein, which associates stably with spliceosomes, was found to interact directly with pre-mRNAs, as demonstrated by UV-crosslinking experiments. The use of various mutant and truncated pre-mRNAs revealed that this interaction requires a spliceable pre-mRNA and an assembled spliceosome; a 3' splice site is not required. To extend these observations to the wild-type PRP2 protein, spliceosomes were depleted of ATP; PRP2 protein interacts with pre-mRNA in these spliceosomes in an ATP-independent fashion. Comparison of RNA binding by PRP2 protein in the presence of ATP or gamma S-ATP showed that ATP hydrolysis rather than mere ATP binding is required to release PRP2 protein from pre-mRNA. As PRP2 is an RNA-stimulated ATPase, these experiments strongly suggest that the pre-mRNA is the native co-factor stimulating ATP hydrolysis by PRP2 protein in spliceosomes. Since PRP2 is a putative RNA helicase, we propose that the pre-mRNA is the target of RNA displacement activity of PRP2 protein, promoting the first step of splicing.

MeSH Terms
Adenosine Triphosphate/metabolism Base Sequence DEAD-box RNA Helicases Fungal Proteins/metabolism,radiation effects Hydrolysis Molecular Sequence Data RNA Helicases RNA Nucleotidyltransferases/metabolism RNA Precursors/metabolism RNA Splicing RNA, Fungal/metabolism RNA, Messenger/metabolism Saccharomyces cerevisiae/genetics Saccharomyces cerevisiae Proteins Spliceosomes/metabolism Ultraviolet Rays
Chemicals
Fungal Proteins RNA Precursors RNA, Fungal RNA, Messenger Saccharomyces cerevisiae Proteins Adenosine Triphosphate RNA Nucleotidyltransferases PRP2 protein, S cerevisiae DEAD-box RNA Helicases RNA Helicases
Authors & Affiliations
4 authors, click to expand affiliations / ORCID
Teigelkamp S
Institute of Cell and Molecular Biology, University of Edinburgh, UK.
McGarvey M
Plumpton M
Beggs J D
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Article Info
Journal
The EMBO journal
Abbr.
EMBO J
ISSN
0261-4189
Published
1994-02-15
Pages
888-97
Language
English
Region
England
NLM ID
8208664
PMCID
PMC394888
Subset
IM
External Links
PubMed source PMC Full Text View journal details
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