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Tandem RNA isolation reveals functional rearrangement of RNA-binding proteins on CDKN1B/p27Kip1 3'UTRs in cisplatin treated cells

Iadevaia, Valentina, Wouters, Maikel D., Kanitz, Alexander, Matia-González, Ana M., Laing, Emma E. and Gerber, André P. (2020) Tandem RNA isolation reveals functional rearrangement of RNA-binding proteins on CDKN1B/p27Kip1 3'UTRs in cisplatin treated cells RNA Biology, 17 (1). pp. 33-46.

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Abstract

Post-transcriptional control of gene expression is mediated via RNA-binding proteins (RBPs) that interact with mRNAs in a combinatorial fashion. While recent global RNA interactome capture experiments expanded the repertoire of cellular RBPs quiet dramatically, little is known about the assembly of RBPs on particular mRNAs; and how these associations change and control the fate of the mRNA in drug-treatment conditions. Here we introduce a novel biochemical approach, termed tobramycinbased tandem RNA isolation procedure (tobTRIP), to quantify proteins associated with the 3'UTRs of cyclin-dependent kinase inhibitor 1B (CDKN1B/p27Kip1) mRNAs in vivo. P27Kip1 plays an important role in mediating a cell's response to cisplatin (CP), a widely used chemotherapeutic cancer drug that induces DNA damage and cell cycle arrest. We found that p27Kip1 mRNA is stabilized upon CP treatment of HEK293 cells through elements in its 3'UTR. Applying tobTRIP, we further compared the associated proteins in CP and non-treated cells, and identified more than fifty interacting RBPs, many functionally related and evoking a coordinated response. Knock-downs of several of the identified RBPs in HEK293 cells confirmed their involvement in CP-induced p27 mRNA regulation; while knock-down of the KH-type splicing regulatory protein (KHSRP) further enhanced the sensitivity of MCF7 adenocarcinoma cancer cells to CP treatment. Our results highlight the benefit of specific in vivo mRNA-protein interactome capture to reveal post-transcriptional regulatory networks implicated in cellular drug response and adaptation.

Item Type: Article
Divisions : Faculty of Health and Medical Sciences > School of Biosciences and Medicine
Authors :
NameEmailORCID
Iadevaia, Valentinav.iadevaia@surrey.ac.uk
Wouters, Maikel D.
Kanitz, Alexander
Matia-González, Ana M.
Laing, Emma E.E.Laing@surrey.ac.uk
Gerber, André P.a.gerber@surrey.ac.uk
Date : January 2020
Funders : Biotechnology and Biological Sciences Research Council (BBSRC), Leverhulme Trust, Royal Society
DOI : /10.1080/15476286.2019.1662268
Copyright Disclaimer : © 2019 The Author(s). Published by Informa UK Limited, trading as Taylor & Francis Group. 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 use, distribution, and reproduction in any medium, provided the original work is properly cited. This is an Accepted Manuscript of an article published by Taylor & Francis in RNA Biology on 16 September 2019 available online: http://www.tandfonline.com/10.1080/15476286.2019.1662268
Uncontrolled Keywords : Post-transcriptional gene regulation; RNA-binding protein; mRNA stability; Drug response; p27; KHSRP
Related URLs :
Depositing User : Clive Harris
Date Deposited : 04 Sep 2019 10:56
Last Modified : 27 Feb 2020 15:47
URI: http://epubs.surrey.ac.uk/id/eprint/852541

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