Ewing's sarcoma and related subtypes of primitive neuroectodermal tumours share a recurrent and specific t(11;22) (q24;q12) chromosome translocation, the breakpoints of which have recently been cloned. Phylogenetically conserved restriction fragments in the vicinity of EWSR1 and EWSR2, the genomic regions where the breakpoints of chromosome 22 and chromosome 11 are, respectively, have allowed identification of transcribed sequences from these regions and has indicated that a hybrid transcript might be generated by the translocation. Here we use these fragments to screen human complementary DNA libraries to show that the translocation alters the open reading frame of an expressed gene on chromosome 22 gene by substituting a sequence encoding a putative RNA-binding domain for that of the DNA-binding domain of the human homologue of murine Fli-1.

Balanced translocations involving band q12 of human chromosome 22 are the most frequent recurrent translocations observed in human solid tumours. It has been shown recently that this region encodes EWS, a protein with an RNA binding homologous domain. In Ewing's sarcoma and malignant melanoma of soft parts, translocations of band 22q12 to chromosome 11 and 12 result in the fusion of EWS with the transcription factors FLI‐1 and ATF1, respectively. The present analysis of 89 Ewing's sarcomas and related tumours show that in addition to the expected EWS‐FLI‐1 fusion, the EWS gene can be fused to ERG, a transcription factor closely related to FLI‐1 but located on chromosome 21. The position of the chromosome translocation breakpoints are shown to be restricted to introns 7‐10 of the EWS gene and widely dispersed within introns 3‐9 of the Ets‐related genes. This heterogeneity generates a variety of chimeric proteins that can be detected by immuno‐precipitation. On rare occasions, they may be associated with a truncated EWS protein arising from alternate splicing. All 13 different fusion proteins that were evidenced contained the N‐terminal domain of EWS and the Ets domain of FLI‐1 or ERG suggesting that oncogenic conversion is achieved by the linking of the two domains with no marked constraint on the connecting peptide.

Merkel cell carcinoma (MCC), a skin tumour with neuroendocrine features, was recently found to be associated with a new type of human polyomavirus, called Merkel cell virus (MCV). We investigated the specificity of this association as well as a causal role of MCV in oncogenesis. DNA and RNA from ten cases of MCC were analysed using PCR and RT-PCR. DNA from 1241 specimens of a wide range of human tumours was also analysed. The DIPS technique was used to identify the integration locus of viral DNA sequences. Array CGH was performed to analyse structural alterations of the cell genome. MCV DNA sequences were found in all ten cases of MCC and in none of the 1241 specimens of other tumour types. Clonal integration of MCV into the host genome was seen in all MCC cases and was checked by FISH in one case. A recurrent pattern of conserved viral sequences which encompassed the replication origin, the small tumour (ST), and the 5' part of the large tumour (LT) antigen DNA sequences was observed. Both ST and LT viral sequences were found to be significantly expressed in all MCCs. Neither recurrent site of integration nor alteration of cellular genes located near the viral sequences was observed. The tight association of MCV with MCC, the clonal pattern of MCV integration, and the expression of the viral oncoproteins strongly support a causative role for MCV in the tumour process. This information will help the development of novel approaches for the assessment and therapy of MCC and biologically related tumours.

As a result of chromosome translocations, the EWS gene is fused to a variety of transcription factors in human solid neoplasia. In Ewing tumors EWS can be fused to four dierent members of the ETS family, namely FLI-1, ERG, ETV1 and E1AF. We have identi®ed a new member of the ETS family, called FEV, which is fused to EWS in a subset of Ewing tumors. FEV encodes a 238 amino acid protein which contains an ETS DNA binding domain closely related to that of FLI-1 and ERG. However, the N-terminal portion of FEV is only 42 amino acids long which suggests that FEV is lacking important transcription regulatory domains contained in FLI-1 and ERG N-terminal parts. The C-terminal end of FEV is rich in alanine residues which may indicate that FEV is a transcription repressor. The FEV gene is encoded by three exons and is located on chromosome 2. FEV expression was only detected in adult prostate and small intestine but not in other adult nor in fetal tissues, thus indicating that FEV has a restricted expression pattern. Following a scheme similar to previously described translocations in Ewing tumors, a t(2;22) chromosome translocation fuses the N-terminal domain of EWS to the ETS DNA binding domain of FEV.