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

Caceres, J. F., Krainer, A. R. (December 1993) Functional analysis of pre-mRNA splicing factor SF2/ASF structural domains. EMBO Journal, 12 (12). pp. 4715-4726. ISSN 0261-4189

Abstract

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 contacts 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.

Item Type:	Paper
Uncontrolled Keywords:	alternative splicing premessenger rna splicing rna-protein interactions rs domain sf2 asf nuclear ribonucleoprotein binding domain conserved family site selection poly(a)-binding protein alternative 5' polymerase-ii snrnp protein 70k protein factor sc35
Subjects:	bioinformatics > genomics and proteomics > genetics & nucleic acid processing > DNA, RNA structure, function, modification bioinformatics > genomics and proteomics > genetics & nucleic acid processing bioinformatics > genomics and proteomics > genetics & nucleic acid processing > DNA, RNA structure, function, modification > nuclear ribonucleoprotein bioinformatics > genomics and proteomics > genetics & nucleic acid processing > DNA, RNA structure, function, modification > pre-mRNA bioinformatics > genomics and proteomics > genetics & nucleic acid processing > DNA, RNA structure, function, modification > RNA splicing
CSHL Authors:	Caceres, Javier Krainer, Adrian R.
Communities:	CSHL labs > Krainer lab
Depositing User:	Matt Covey
Date:	December 1993
Date Deposited:	11 Dec 2013 14:58
Last Modified:	11 Dec 2013 14:58
PMCID:	PMC413916
URI:	https://repository.cshl.edu/id/eprint/28954

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