Isolation of an active step I spliceosome and composition of its RNP core
- 5 March 2008
- journal article
- research article
- Published by Springer Science and Business Media LLC in Nature
- Vol. 452 (7189), 846-850
- https://doi.org/10.1038/nature06842
Abstract
Formation of catalytically active RNA structures within the spliceosome requires the assistance of proteins. However, little is known about the number and nature of proteins needed to establish and maintain the spliceosome’s active site. Here we affinity-purified human spliceosomal C complexes and show that they catalyse exon ligation in the absence of added factors. Comparisons of the composition of the precatalytic versus the catalytic spliceosome revealed a marked exchange of proteins during the transition from the B to the C complex, with apparent stabilization of Prp19–CDC5 complex proteins and destabilization of SF3a/b proteins. Disruption of purified C complexes led to the isolation of a salt-stable ribonucleoprotein (RNP) core that contained both splicing intermediates and U2, U5 and U6 small nuclear RNA plus predominantly U5 and human Prp19–CDC5 proteins and Prp19-related factors. Our data provide insights into the spliceosome’s catalytic RNP domain and indicate a central role for the aforementioned proteins in sustaining its catalytically active structure.This publication has 23 references indexed in Scilit:
- Repositioning of the Reaction Intermediate within the Catalytic Center of the SpliceosomeMolecular Cell, 2006
- Prp8 protein: At the heart of the spliceosomeRNA, 2005
- A subset of human 35S U5 proteins, including Prp19, function prior to catalytic step 1 of splicingThe EMBO Journal, 2004
- The Prp19p-Associated Complex in Spliceosome ActivationScience, 2003
- Splicing-related catalysis by protein-free snRNAsNature, 2001
- Functional analysis of the human CDC5L complex and identification of its components by mass spectrometryThe EMBO Journal, 2000
- Mechanical Devices of the Spliceosome: Motors, Clocks, Springs, and ThingsCell, 1998
- Protein functions in pre-mRNA splicingCurrent Opinion in Cell Biology, 1997
- Evidence that sequence-independent binding of highly conserved U2 snRNP proteins upstream of the branch site is required for assembly of spliceosomal complex A.Genes & Development, 1996
- Yeast precursor mRNA processing protein PRP19 associates with the spliceosome concomitant with or just after dissociation of U4 small nuclear RNA.Proceedings of the National Academy of Sciences of the United States of America, 1993