title: Junction region of EWS-FLI1 fusion protein has a dominant negative effect in Ewing's sarcoma in vitro.
abstract: BACKGROUND
  Ewing's sarcoma is a malignancy characterized by a specific 11:22 chromosomal translocation which generates a novel EWS-FLI1 fusion protein functioning as an aberrant transcription factor. In the present study, we have further characterized the junction region of the EWS-FLI1 fusion protein.
  
  
  METHODS
  In-silico model of EWS-FLI1 fusion protein was analysed for ligand binding sites, and a putative region (amino acid (aa) 251-343 of the type 1 fusion protein) in the vicinity of the fusion junction was cloned and expressed using bacterial expression. The recombinant protein was characterized by Circular Dichroism (CD). We then expressed aa 251-280 ectopically in Ewing's sarcoma cell-line and its effect on cell proliferation, tumorigenicity and expression of EWS-FLI1 target genes were analysed.
  
  
  RESULTS
  Our modelling analysis indicated that Junction region (aa 251-343) encompasses potential ligand biding sites in the EWS-FLI1 protein and when expressed in bacteria was present as soluble form. Ectopically expressing this region in Ewing's sarcoma cells inhibited tumorigenicity, and EWS-FLI1 target genes indicating a dominant negative biological effect.
  
  
  CONCLUSIONS
  Junction region can be exploited further as target for drug development in future to specifically target EWS-FLI1 in Ewing's Sarcoma.
authors.affiliation: Department of Molecular Oncology, Cancer Institute (WIA), 38, Sardar Patel Road, Chennai, 600036, India.
authors.forename: Babu
authors.initials: B
authors.lastname: Jully
authors.forename: Ramshankar
authors.initials: R
authors.lastname: Vijayalakshmi
authors.forename: Gopisetty
authors.initials: G
authors.lastname: Gopal
authors.forename: Kesavan
authors.initials: K
authors.lastname: Sabitha
authors.forename: Thangarajan
authors.initials: T
authors.lastname: Rajkumar
mesh_terms: D000595:Amino Acid Sequence; D045744:Cell Line, Tumor; D049109:Cell Proliferation; D002942:Circular Dichroism; D019938:Cyclin D1 / Q000235:genetics / Q000378:metabolism; D000071221:Enhancer of Zeste Homolog 2 Protein; D058750:Epithelial-Mesenchymal Transition / Q000235:genetics; D015972:Gene Expression Regulation, Neoplastic; D000090322:Homeobox Protein Nkx-2.2; D018398:Homeodomain Proteins / Q000235:genetics / Q000378:metabolism; D006801:Humans; D015151:Immunoblotting; D061986:MCF-7 Cells; D008969:Molecular Sequence Data; D015514:Oncogene Proteins, Fusion / Q000737:chemistry / Q000235:genetics* / Q000378:metabolism; D010455:Peptides / Q000235:genetics / Q000378:metabolism; D063151:Polycomb Repressive Complex 2 / Q000235:genetics / Q000378:metabolism; D051789:Proto-Oncogene Protein c-fli-1 / Q000737:chemistry / Q000235:genetics* / Q000378:metabolism; D016271:Proto-Oncogene Proteins c-myc / Q000235:genetics / Q000378:metabolism; D034802:RNA-Binding Protein EWS / Q000737:chemistry / Q000235:genetics* / Q000378:metabolism; D011994:Recombinant Proteins / Q000737:chemistry / Q000378:metabolism; D020133:Reverse Transcriptase Polymerase Chain Reaction; D012512:Sarcoma, Ewing / Q000235:genetics* / Q000378:metabolism / Q000473:pathology; D014157:Transcription Factors / Q000235:genetics / Q000378:metabolism; D014162:Transfection; D029961:Zebrafish Proteins
chemical_list: C106031:EWS-FLI fusion protein; D000090322:Homeobox Protein Nkx-2.2; D018398:Homeodomain Proteins; C489427:MYC protein, human; D015514:Oncogene Proteins, Fusion; D010455:Peptides; D051789:Proto-Oncogene Protein c-fli-1; D016271:Proto-Oncogene Proteins c-myc; D034802:RNA-Binding Protein EWS; D011994:Recombinant Proteins; D014157:Transcription Factors; D029961:Zebrafish Proteins; D019938:Cyclin D1; C498409:EZH2 protein, human; D000071221:Enhancer of Zeste Homolog 2 Protein; D063151:Polycomb Repressive Complex 2