Assembly and dynamics of the U4/U6 di-snRNP by single-molecule FRET
File(s)
Published version
Author(s)
Hardin, JW
Warnasooriya, C
Kondo, Y
Nagai, K
Rueda, DS
Type
Journal Article
Abstract
In large ribonucleoprotein machines, such as ribosomes and spliceosomes, RNA functions as an assembly scaffold as well as a critical catalytic component. Protein binding to the RNA scaffold can induce structural changes, which in turn modulate subsequent binding of other components. The spliceosomal U4/U6 di-snRNP contains extensively base paired U4 and U6 snRNAs, Snu13, Prp31, Prp3 and Prp4, seven Sm and seven LSm proteins. We have studied successive binding of all protein components to the snRNA duplex during di-snRNP assembly by electrophoretic mobility shift assay and accompanying conformational changes in the U4/U6 RNA 3-way junction by single-molecule FRET. Stems I and II of the duplex were found to co-axially stack in free RNA and function as a rigid scaffold during the entire assembly, but the U4 snRNA 5′ stem-loop adopts alternative orientations each stabilized by Prp31 and Prp3/4 binding accounting for altered Prp3/4 binding affinities in presence of Prp31.
Date Issued
2015-12-15
Date Acceptance
2015-09-24
Citation
Nucleic Acids Research, 2015, 43 (22), pp.10963-10974
ISSN
1362-4962
Publisher
Oxford University Press (OUP): Policy C - Option B
Start Page
10963
End Page
10974
Journal / Book Title
Nucleic Acids Research
Volume
43
Issue
22
Copyright Statement
© The Author(s) 2015. Published by Oxford University Press on behalf of Nucleic Acids Research. This is an Open Access article distributed under the terms of the Creative Commons Attribution License (http://creativecommons.org/licenses/by/4.0/), which permits unrestricted reuse, distribution, and reproduction in any medium, provided the original work is properly cited.
License URL
Publication Status
Published
Date Publish Online
2015-10-25