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- Title
Homotypic and heterotypic interactions of EWS, FLI1 and their oncogenic fusion protein.
- Authors
Spahn, Laura; Siligan, Christine; Bachmaier, Radostina; Schmid, Johannes A; Aryee, Dave N T; Kovar, Heinrich
- Abstract
In Ewing's sarcoma family tumors, the ets transcription factor gene FLI1 is rearranged with one EWS allele resulting in coexpression of germline EWS and chimeric EWS-FLI1 proteins. Here, we investigated the potential of germline EWS, FLI1 and EWS-FLI1 to oligomerize. In two functional in vivo tests, fluorescence resonance energy transfer (FRET) and the mammalian two-hybrid (MTH) assay, self-association of EWS and EWS-FLI1, but not of FLI1 was detected. In addition, interaction of EWS-FLI1 with EWS and FLI1 was observed. GST pull-down assays and immunoprecipitation experiments largely confirmed these results. The EWS N-terminal domain present in both EWS and EWS-FLI1 was found to contribute to homotypic and heterotypic interactions of these proteins. However, in the context of germline EWS, the presence of the whole or part of the C-terminal RNA-binding domain greatly supported the self-association potential of the protein. Involvement of an RNA component in EWS oligomerization was confirmed by sensitivity of the corresponding GST pull-down assay to RNaseA treatment. In contrast, EWS-FLI1 was able to self-associate and also bind to FLI1 via its C-terminal domain, which comprises the FLI1 DNA-binding motif. Accordingly, the EWS-FLI1 interaction was not disrupted by RNaseA treatment. Despite its potential to oligomerize, EWS-FLI1 bound to a tandem ets-binding site of the TGFß type II receptor promoter as a monomer. Therefore, the functional consequences of homo- and hetero-oligomerization of EWS and EWS-FLI1 proteins remain to be elucidated.Oncogene (2003) 22, 6819-6829. doi:10.1038/sj.onc.1206810
- Subjects
SARCOMA; TUMORS; PROTEINS; EWING'S sarcoma; MONOMERS
- Publication
Oncogene, 2003, Vol 22, Issue 44, p6819
- ISSN
0950-9232
- Publication type
Article
- DOI
10.1038/sj.onc.1206810