Multiple domains mediate transformation by the Ewing's sarcoma EWS/FLI-1 fusion gene.
The (11;22) chromosomal translocation found in Ewing's sarcoma and related tumors fuses the amino terminus of the EWS protein to the DNA-binding domain of the FLI-1 transcription factor. In contrast to normal FLI-1, the EWS/FLI-1 fusion transforms NIH3T3 cells and this activity requires both EWS and FLI-1 sequences. Reporter gene assays showed that the portion of EWS fused to FLI-1 encodes a strong transcriptional activation domain. To determine whether this function is necessary for transformation by EWS/FLI-1, deletion analysis of EWS was performed. We found that the EWS domain could be functionally subdivided into two regions: (i) an amino terminal domain (domain A) which transforms efficiently when fused to FLI-1 but has little transactivation activity in a model system and (ii) a distal region (domain B) which transactivates efficiently but transforms less efficiently when fused to FLI-1. Replacement of the EWS domain with known heterologous transcriptional activation domains yielded chimeric FLI-1 fusions that in some instances could transform NIH3T3 cells. Finally we demonstrate that EWS/FLI-1 and related FLI-1 chimeras are able to cooperate with another transcription factor to activate a model reporter gene. These results further demonstrate that EWS/FLI-1 is an aberrant transcription factor and suggest that the EWS domain mediates important protein-protein interactions with other factors resulting in the transcriptional modulation of target genes.
Lessnick
S L
SL
Molecular Biology Institute, Gwynne Hazen Cherry Memorial Laboratories, University of California, Los Angeles 90024.
Braun
B S
BS
Denny
C T
CT
May
W A
WA
D016475:3T3 Cells; D000818:Animals; D001483:Base Sequence; D002471:Cell Transformation, Neoplastic / Q000235:genetics*; D003001:Cloning, Molecular; D004268:DNA-Binding Proteins / Q000235:genetics*; D015321:Gene Rearrangement; D034441:Heterogeneous-Nuclear Ribonucleoproteins; D051379:Mice; D008969:Molecular Sequence Data; D009363:Neoplasm Proteins / Q000235:genetics*; D009687:Nuclear Proteins / Q000235:genetics*; D051789:Proto-Oncogene Protein c-fli-1; D011518:Proto-Oncogene Proteins; D034802:RNA-Binding Protein EWS; D016601:RNA-Binding Proteins / Q000235:genetics*; D011993:Recombinant Fusion Proteins / Q000235:genetics; D012261:Ribonucleoproteins / Q000235:genetics*; D012512:Sarcoma, Ewing / Q000235:genetics*; D017384:Sequence Deletion; D015534:Trans-Activators / Q000235:genetics*
D004268:DNA-Binding Proteins; C497426:Fli1 protein, mouse; D034441:Heterogeneous-Nuclear Ribonucleoproteins; D009363:Neoplasm Proteins; D009687:Nuclear Proteins; D051789:Proto-Oncogene Protein c-fli-1; D011518:Proto-Oncogene Proteins; D034802:RNA-Binding Protein EWS; D016601:RNA-Binding Proteins; D011993:Recombinant Fusion Proteins; D012261:Ribonucleoproteins; D015534:Trans-Activators