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oai:recercat.cat:10230/237222025-12-12T01:40:57Zcom_2072_6col_2072_452952

Regulation of FAS exon definition and apoptosis by the ewing sarcoma protein Paronetto, Maria Paola Bernardis, Isabella Volpe, Elisabetta Bechara, Elias Sebestyén, Endre Eyras Jiménez, Eduardo Valcárcel, J. (Juan) Proteïnes Antígens CD RNA The Ewing sarcoma protein EWS is an RNA and DNA binding protein implicated in transcription, pre-mRNA splicing, and DNA damage response. Using CLIP-seq, we identified EWS RNA binding sites in exonic regions near 5′ splice sites. A prominent target was exon 6 of the FAS/CD95 receptor, which is alternatively spliced to generate isoforms with opposing activities in programmed cell death. Depletion and overexpression experiments showed that EWS promotes exon 6 inclusion and consequently the synthesis of the proapoptotic FAS/CD95 isoform, whereas an EWS-FLI1 fusion protein characteristic of Ewing sarcomas shows decreased activity. Biochemical analyses revealed that EWS binding promotes the recruitment of U1snRNP and U2AF65 to the splice sites flanking exon 6 and therefore exon definition. Consistent with a role for EWS in the regulation of programmed cell death, cells depleted of EWS show decreased sensitivity to FAS-induced apoptosis, and elevated EWS expression enhances apoptosis in EWS-haploinsufficient Ewing sarcoma cells. M.P.P. was supported by a fellowship from the HFSP. This project was supported by Fundación Botín, Fundación Sandra Ibarra (FSI2013), the European Union Sixth Framework Programme under grant agreement Nr. LSHG-CT-2005-518238-V (EURASNET), the Spanish Ministry of Economy and Competitiveness (grant no. CSD2009-00080, Consolider RNAREG/BFU2011-29583 / BIO2011-23920), and AIRC (Grant MFAG 11658). We also acknowledge support of the Spanish Ministry of Economy and Competitiveness, “Centro de Excelencia Severo Ochoa 2013-2017” (SEV-2012-0208) 2015-06-04T06:56:36Z 2015-06-04T06:56:36Z 2014 info:eu-repo/semantics/article info:eu-repo/semantics/acceptedVersion Cell Reports. 2014; 7: 1211-1226 info:eu-repo/grantAgreement/EC/FP6/518238 info:eu-repo/grantAgreement/ES/3PN/CSD2009-00080 info:eu-repo/grantAgreement/ES/3PN/BFU2011-29583 info:eu-repo/grantAgreement/ES/3PN/BIO2011-23920 © 2014, Elsevier. Licensed under the Creative Commons Attribution-NonCommercial-NoDerivatives 3.0 International http://creativecommons.org/licenses/by-nc-nd/3.0/ http://dx.doi.org/10.1016/j.celrep.2014.03.077 http://creativecommons.org/licenses/by-nc-nd/3.0/ info:eu-repo/semantics/openAccess Elsevier