Functional analysis of pre-mRNA splicing factor SF2/ASF structural domains.

Human pre-mRNA splicing factor SF2/ASF has an activity required for general splicing in vitro and promotes utilization of proximal alternative 5' splice sites in a concentration-dependent manner by opposing hnRNP A1. We introduced selected mutations in the N-terminal RNA recognition motif (RRM)...

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Main Authors: Cáceres, J F, Krainer, A R
Formato: Artigo
Idioma:en
Publicado em: 1993
Assuntos:
Research Article
Acesso em linha:https://ncbi.nlm.nih.gov/pmc/articles/PMC413916/
https://ncbi.nlm.nih.gov/pubmed/8223480
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spelling pubmed-4139162005-04-27 Functional analysis of pre-mRNA splicing factor SF2/ASF structural domains. Cáceres, J F Krainer, A R EMBO J Research Article Human pre-mRNA splicing factor SF2/ASF has an activity required for general splicing in vitro and promotes utilization of proximal alternative 5' splice sites in a concentration-dependent manner by opposing hnRNP A1. We introduced selected mutations in the N-terminal RNA recognition motif (RRM) and the C-terminal Arg/Ser (RS) domain of SF2/ASF, and assayed the resulting recombinant proteins for constitutive and alternative splicing in vitro and for binding to pre-mRNA and mRNA. Mutants inactive in constitutive splicing can affect alternative splice site selection, demonstrating that these activities involve distinct molecular interactions. Specific protein-RNA contact mediated by Phe56 and Phe58 in the RNP-1 submotif of the SF2/ASF RRM are essential for constitutive splicing, although they are not required for RRM-mediated binding to pre-mRNA. The RS domain is also required for constitutive splicing activity and both Arg and Ser residues are important. Analysis of domain deletion mutants demonstrated strong synergy between the RRM and a central degenerate RRM repeat in binding to RNA. These two domains are sufficient for alternative splicing activity in the absence of an RS domain. 1993-12 /pmc/articles/PMC413916/ /pubmed/8223480 Text en
institution US National Library of Medicine
collection PubMed Central
language en
format Article
topic Research Article
spellingShingle Research Article
Cáceres, J F
Krainer, A R
Functional analysis of pre-mRNA splicing factor SF2/ASF structural domains.
description Human pre-mRNA splicing factor SF2/ASF has an activity required for general splicing in vitro and promotes utilization of proximal alternative 5' splice sites in a concentration-dependent manner by opposing hnRNP A1. We introduced selected mutations in the N-terminal RNA recognition motif (RRM) and the C-terminal Arg/Ser (RS) domain of SF2/ASF, and assayed the resulting recombinant proteins for constitutive and alternative splicing in vitro and for binding to pre-mRNA and mRNA. Mutants inactive in constitutive splicing can affect alternative splice site selection, demonstrating that these activities involve distinct molecular interactions. Specific protein-RNA contact mediated by Phe56 and Phe58 in the RNP-1 submotif of the SF2/ASF RRM are essential for constitutive splicing, although they are not required for RRM-mediated binding to pre-mRNA. The RS domain is also required for constitutive splicing activity and both Arg and Ser residues are important. Analysis of domain deletion mutants demonstrated strong synergy between the RRM and a central degenerate RRM repeat in binding to RNA. These two domains are sufficient for alternative splicing activity in the absence of an RS domain.
author Cáceres, J F
Krainer, A R
author_facet Cáceres, J F
Krainer, A R
author_sort Cáceres, J F
title Functional analysis of pre-mRNA splicing factor SF2/ASF structural domains.
title_short Functional analysis of pre-mRNA splicing factor SF2/ASF structural domains.
title_full Functional analysis of pre-mRNA splicing factor SF2/ASF structural domains.
title_fullStr Functional analysis of pre-mRNA splicing factor SF2/ASF structural domains.
title_full_unstemmed Functional analysis of pre-mRNA splicing factor SF2/ASF structural domains.
title_sort functional analysis of pre-mrna splicing factor sf2/asf structural domains.
publishDate 1993
url https://ncbi.nlm.nih.gov/pmc/articles/PMC413916/
https://ncbi.nlm.nih.gov/pubmed/8223480
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