Attenuation of the ELAV1-like protein HuR sensitizes adenocarcinoma cells to the intrinsic apoptotic pathway by increasing the translation of caspase-2L

Caspase-2 represents the most conserved member of the caspase family, which exhibits features of both initiator and effector caspases. Using ribonucleoprotein (RNP)-immunoprecipitation assay, we identified the proapoptot
Caspase-2 represents the most conserved member of the caspase family, which exhibits features of both initiator and effector caspases. Using ribonucleoprotein (RNP)-immunoprecipitation assay, we identified the proapoptotic caspase-2L encoding mRNA as a novel target of the ubiquitous RNA-binding protein HuR in DLD-1 colon carcinoma cells. Unexpectedly, crosslinking-RNP and RNA probe pull-down experiments revealed that HuR binds exclusively to the caspase-2-5' untranslated region (UTR) despite that the 3' UTR of the mRNA bears several adenylate- and uridylate-rich elements representing the prototypical HuR binding sites. By using RNAi-mediated loss-of-function approach, we observed that HuR regulates the mRNA and in turn the protein levels of caspase-2 in a negative manner. Silencing of HuR did not affect the stability of caspase-2 mRNA but resulted in an increased redistribution of caspase-2 transcripts from RNP particles to translational active polysomes implicating that HuR exerts a direct repressive effect on caspase-2 translation. Consistently, in vitro translation of a luciferase reporter gene under the control of an upstream caspase-2-5'UTR was strongly impaired after the addition of recombinant HuR, whereas translation of caspase-2 coding region without the 5'UTR is not affected by HuR confirming the functional role of the caspase-2-5'UTR. Functionally, an elevation in caspase-2 level by HuR knockdown correlated with an increased sensitivity of cells to apoptosis induced by staurosporine- and pore-forming toxins as implicated by their significant accumulation in the sub G1 phase and an increase in caspase-2, -3 and poly ADP-ribose polymerase cleavage, respectively. Importantly, HuR knockdown cells remained insensitive toward STS-induced apoptosis if cells were additionally transfected with caspase-2-specific siRNAs. Collectively, our findings support the hypothesis that HuR by acting as an endogenous inhibitor of caspase-2-driven apoptosis may essentially contribute to the antiapoptotic program of adenocarcinoma cells by HuR. 
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Metadaten
Author:Christoph Winkler, Anke Doller, Gergely Imre, Amel Badawi, Tobias Schmid, Sebastian Schulz, Nico Steinmeyer, Josef Martin Pfeilschifter, Krishnaraj Rajalingam, Wolfgang Eberhardt
URN:urn:nbn:de:hebis:30:3-420035
DOI:http://dx.doi.org/10.1038/cddis.2014.279
Pubmed Id:http://www.ncbi.nlm.nih.gov/pubmed?term=25010987
Parent Title (English):Cell death and disease
Document Type:Article
Language:English
Date of Publication (online):2016/11/11
Date of first Publication:2014/07/10
Publishing Institution:Universitätsbibliothek Johann Christian Senckenberg
Release Date:2016/11/11
Volume:5
Issue:e1321
Pagenumber:12
HeBIS PPN:42481952X
Institutes:Pharmazie
Medizin
Zentrum für Arzneimittelforschung, Entwicklung und Sicherheit
Dewey Decimal Classification:570 Biowissenschaften; Biologie
610 Medizin und Gesundheit
Sammlungen:Universitätspublikationen
Licence (German):License LogoCreative Commons - Namensnennung 4.0

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