The 11;22 chromosomal translocation specifically linked to Ewing sarcoma and primitive neuroectodermal tumor results in a chimeric molecule fusing the amino-terminal-encoding portion of the EWS gene to the carboxyl-terminal DNA-binding domain encoded by the FLI1 gene.
From these results it appears that a portion of the Fli-1 ATA domain (H-L-H region) was replaced by the amino-terminal domain of EWS gene in Ewing's sarcoma cases.
EWS was first identified as forming a hybrid transcript in Ewing's sarcoma, which links its N-terminal domain to the DNA binding domain of the FLI-1 gene.
The EWS gene has been identified based on its location at the chromosome 22 breakpoint of the t(11;22)(q24;q12) translocation that characterizes Ewing sarcoma and related neuroectodermal tumors.
In the present study, we examined the efficacy of an EWS complementary DNA (cDNA) probe in detecting the t(11;22) in Southern blots of EcoRI- or HindIII-digested DNA extracted from cases of ES and PNET.
Thus, the TLS/FUS-ERG gene fusion in t(16;21) leukemia is predicted to produce a protein that is very similar to the EWS-ERG chimeric protein responsible for Ewing's sarcoma.
Although recent data have demonstrated that the chimeric EWS-FLI-1 cDNA isolated from cases of Ewing's sarcoma can transform NIH 3T3 cells, little is known about the basis for this transformation.
Significantly, the DNA binding portion of FLI-1 is the 3' part of an oncogenic fusion transcript (termed EWS-FLI) in human Ewing's sarcoma and neuroepithelioma.
Southern blot analysis revealed recurrent rearrangement of both EWS, located at 22q12, and rearranged in other tumor-specific translocations in Ewing's sarcoma and clear cell sarcoma, and of WT1, the gene at 11p13 involved in a subset of Wilms' tumor.
Further, we describe an alternative translocation in an ES cell line (#5838) in which the 5' end of the EWS gene is juxtaposed to the 3' end of the ERG gene.
These results suggest that EWS-FLI-1 contributes to the transformed phenotype of ES tumor cells by inducing the deregulated and/or unscheduled activation of genes normally responsive to FLI-1 or to other close members of the Ets family.
We have identified a second Ewing's sarcoma translocation, t(21;22)(q22;q12), that fuses EWS to a different ETS family member, the ERG gene located on band 21q22.
Recent cloning of the chromosome breakpoint regions of t(11;22)(q24;q12)Ewing's sarcoma translocation has revealed that the breakpoints were localized within the Ewing's sarcoma gene (EWS gene) on chromosome 22 and the Fli-1 gene on chromosome 11.
All 30 clinical specimens had been confirmed to contain sufficient ES/PNET DNA by the demonstration of a rearrangement of the t(11;22)-associated EWS gene using an EWS cDNA probe on the same blots.
By combining the PAX3/FKHR RT-PCR assay with primers for detection of the Ewing's sarcoma t(11;22) encoded EWS/FLI-1 chimeric transcript, we have developed a multiplex RT-PCR reaction that allows the rapid and accurate identification of either translocation in a biopsy sample.