Identification of target genes for the Ewing's sarcoma EWS/FLI fusion protein by representational difference analysis.
The EWS/FLI-1 fusion gene results from the 11;22 chromosomal translocation in Ewing's sarcoma. The product of the gene is one of a growing number of structurally altered transcription factors implicated in oncogenesis. We have employed a subtractive cloning strategy of representational difference analysis in conjunction with a model transformation system to identify genes transcribed in response to EWS/FLI. We have characterized eight transcripts that are dependent on EWS/FLI for expression and two transcripts that are repressed in response to EWS/FLI. Three of the former were identified by sequence analysis as stromelysin 1, a murine homolog of cytochrome P-450 F1 and cytokeratin 15. Stromelysin 1 is induced rapidly after expression of EWS/FLI, suggesting that the stromelysin 1 gene may be a direct target gene of EWS/FLI. These results demonstrate that expression of EWS/FLI leads to significant changes in the transcription of specific genes and that these effects are at least partially distinct from those caused by expression of germ line FLI-1. The representational difference analysis technique can potentially be applied to investigate transformation pathways activated by a broad array of genes in different tumor systems.
Braun
B S
BS
Molecular Biology Institute, University of California, Los Angeles 90024, USA.
Frieden
R
R
Lessnick
S L
SL
May
W A
WA
Denny
C T
CT
D001483:Base Sequence; D018076:DNA, Complementary / Q000235:genetics; D004268:DNA-Binding Proteins / Q000235:genetics / Q000378:metabolism*; D015972:Gene Expression Regulation, Neoplastic; D034441:Heterogeneous-Nuclear Ribonucleoproteins; D019278:Matrix Metalloproteinase 3; D008666:Metalloendopeptidases / Q000235:genetics; D008967:Molecular Biology / Q000379:methods; D008969:Molecular Sequence Data; D009363:Neoplasm Proteins / Q000235:genetics / Q000378:metabolism*; D009687:Nuclear Proteins / Q000235:genetics / Q000378:metabolism*; D051789:Proto-Oncogene Protein c-fli-1; D011518:Proto-Oncogene Proteins; D012333:RNA, Messenger / Q000235:genetics; D012334:RNA, Neoplasm / Q000235:genetics*; D034802:RNA-Binding Protein EWS; D016601:RNA-Binding Proteins / Q000235:genetics / Q000378:metabolism*; D012261:Ribonucleoproteins / Q000235:genetics / Q000378:metabolism*; D012512:Sarcoma, Ewing / Q000235:genetics*; D012641:Selection, Genetic; D015534:Trans-Activators / Q000235:genetics / Q000378:metabolism*; D015854:Up-Regulation
D018076:DNA, Complementary; D004268:DNA-Binding Proteins; D034441:Heterogeneous-Nuclear Ribonucleoproteins; D009363:Neoplasm Proteins; D009687:Nuclear Proteins; D051789:Proto-Oncogene Protein c-fli-1; D011518:Proto-Oncogene Proteins; D012333:RNA, Messenger; D012334:RNA, Neoplasm; D034802:RNA-Binding Protein EWS; D016601:RNA-Binding Proteins; D012261:Ribonucleoproteins; D015534:Trans-Activators; D008666:Metalloendopeptidases; D019278:Matrix Metalloproteinase 3