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.
