Chew, S. L., Liu, H. X., Mayeda, A., Krainer, A. R. (September 1999) Evidence for the function of an exonic splicing enhancer after the first catalytic step of pre-mRNA splicing. Proceedings of the National Academy of Sciences of the United States of America, 96 (19). pp. 10655-10660. ISSN 0027-8424
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Abstract
Exonic splicing enhancers (ESEs) activate pre-mRNA splicing by promoting the use of the flanking splice sites, They are recognized by members of the serine/arginine-rich (SR) family of proteins, such as splicing factor 2/alternative splicing factor (SF2/ASF), which recruit basal splicing factors to form the initial complexes during spliceosome assembly. The in vitro splicing kinetics of an ESE-dependent IgM pre-mRNA suggested that an SF2/ASF-specific ESE has additional functions later in the splicing reaction, after the completion of the first catalytic step. A bimolecular exon ligation assay, which physically uncouples the first and second catalytic steps of splicing in a trans-splicing reaction, was adapted to test the function of the ESE after the first step. A 3' exon containing the SF2/ASF-specific ESE underwent bimolecular exon ligation, whereas 3' exons without the ESE or with control sequences did not. The ESE-dependent trans-splicing reaction occurred after inactivation of U1 or U2 small nuclear ribonucleoprotein particles, compatible with a functional assay for events after the first step of splicing. The ESE-dependent step appears to take place before the ATP-independent part of the second catalytic step. Bimolecular exon ligation also occurred in an S100 cytosolic extract, requiring both the SF2/ASF-dependent ESE and complementation with SF2/ASF, These data suggest that some ESEs can act late in the splicing reaction, together with appropriate SR proteins, to enhance the second catalytic step of splicing.
| Item Type: | Paper |
|---|---|
| Uncontrolled Keywords: | messenger-rna sr proteins expression spliceosome mechanism sequences subset sites gene |
| Subjects: | bioinformatics > genomics and proteomics > genetics & nucleic acid processing > DNA, RNA structure, function, modification bioinformatics > genomics and proteomics > genetics & nucleic acid processing > DNA, RNA structure, function, modification > exons > exon splicing 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: | |
| Communities: | CSHL labs > Krainer lab |
| Depositing User: | Matt Covey |
| Date: | September 1999 |
| Date Deposited: | 11 Dec 2013 19:29 |
| Last Modified: | 10 Sep 2019 19:39 |
| PMCID: | PMC17938 |
| Related URLs: | |
| URI: | https://repository.cshl.edu/id/eprint/28929 |
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