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Title: The organization of RNA contacts by PTB for regulation of FAS splicing
Authors: Mickleburgh, I.
Kafasla, P.
Cherny, Dmitry
Llorian, M.
Curry, S.
Jackson, R. J.
Smith, C. W.
First Published: 23-Jun-2014
Publisher: Oxford University Press (OUP)
Citation: Nucleic Acids Research, 2014, 42 (13), pp. 8605-8620
Abstract: Post-transcriptional steps of gene expression are regulated by RNA binding proteins. Major progress has been made in characterizing RNA-protein interactions, from high resolution structures to transcriptome-wide profiling. Due to the inherent technical challenges, less attention has been paid to the way in which proteins with multiple RNA binding domains engage with target RNAs. We have investigated how the four RNA recognition motif (RRM) domains of Polypyrimidine tract binding (PTB) protein, a major splicing regulator, interact with FAS pre-mRNA under conditions in which PTB represses FAS exon 6 splicing. A combination of tethered hydroxyl radical probing, targeted inactivation of individual RRMs and single molecule analyses revealed an unequal division of labour between the four RRMs of PTB. RNA binding by RRM4 is the most important for function despite the low intrinsic binding specificity and the complete lack of effect of disrupting individual RRM4 contact points on the RNA. The ordered RRM3-4 di-domain packing provides an extended binding surface for RNA interacting at RRM4, via basic residues in the preceding linker. Our results illustrate how multiple alternative low-specificity binding configurations of RRM4 are consistent with repressor function as long as the overall ribonucleoprotein architecture provided by appropriate di-domain packing is maintained.
DOI Link: 10.1093/nar/gku519
ISSN: 0305-1048
eISSN: 1362-4962
Version: Publisher Version
Status: Peer-reviewed
Type: Journal Article
Rights: © The Author(s) 2014. 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 (CC BY 3.0) (, which permits unrestricted reuse, distribution, and reproduction in any medium, provided the original work is properly cited.
Description: PMCID: PMC4117754
Appears in Collections:Published Articles, Dept. of Biochemistry

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