A possible explanation for these observations
A possible explanation for these observations has been provided by a recent study showing that EWS-FLI-1 increases the Skp2-mediated 26S proteasome degradation, decreasing p27 protein stability and preventing cell senescence .
Mechanism of action of EWS-FLI-1 Molecular analysis has revealed that several EWS-ETS target gene promoters contain tandem atp 4 for Ets and AP-1 proteins. Ets family members that form fusion proteins with EWS, including FLI-1, ERG and ETV1, were found to cooperatively bind these tandem elements with Fos-Jun whereas other Ets family members were not. C-terminal domain mutants of EWS-FLI-1 that cannot bind DNA together with Fos-Jun were reported to lose the ability to transform 3T3 fibroblasts. These observations suggest that the cooperation between EWS-FLI-1 and Fos-Jun is essential for at least some of the biological activities of the fusion protein . In addition, recent evidence suggests that EWS-FLI-1 associates with other proteins that may influence its function. Thus, a phage display library screen uncovered a EWS-FLI-1 binding peptide containing homology to RNA helicase A (RHA). Subsequent experiments identified RHA in a protein complex with EWS-FLI-1 in ESFT cell lines and showed that RHA binds to known EWS-FLI-1 target gene promoters, enhancing EWS-FLI-1 function .
The potential origin of Ewing sarcoma From the preceding discussion, it appears obvious that at least two key issues still remain to be addressed in order to understand Ewing’s sarcoma biology: the identity of the cells from which ESFT originate, and which presumably display permissiveness for EWS-FLI-1-mediated transformation, and the possibility that EWS-FLI-1 is the unique initiating event in ESFT development. Although mutation of p53 and loss of p16INK4A/p14ARF have been documented in ESFT, they occur in a minority of tumors, and in about 15–20% of ESFT, the only detected genetic event appears to be the t(11;22) chromosomal translocation leading to EWS-FLI-1 expression. These observations are consistent with the existence of a primary cell that can be transformed by EWS-FLI-1. Until recently, there has been no indication as to the possible nature of such a cell, other than the widely held view that sarcomas in general originate from undefined mesenchymal stem/progenitor cells . The histological features of Ewing’s sarcoma suggest a poorly differentiated tumor that has both mesenchymal and neuroectodermal features . This has led to an as yet unresolved debate as to the neuroectodermal or mesenchymal origin of these tumors. Numerous efforts have been made to identify primary cells that might undergo transformation as a result of EWS-FLI-1 expression. However, initial results were somewhat surprising. Thus, introduction of EWS-FLI-1 into mouse embryonic fibroblasts (MEFs) resulted in cell cycle arrest and cell death, with the surviving MEFs losing EWS-FLI-1 expression . MEFs from p19ARF−/− mice transfected with EWS-FLI-1 were observed to maintain EWS-FLI-1 expression but did not form tumors in vivo. Loss of p53 also failed to induce tumorigenesis by MEFs expressing EWS-FLI-1 . Only upon transformation with SV40-T antigen could MEFs lacking p19ARF or p53 and expressing EWS-FLI-1 form tumors in vivo with histological features that resemble the human Ewing sarcoma phenotype . Similar observations were made in hTERT-immortalized human primary fibroblasts where EWS-FLI-1 expression induced p53-mediated growth arrest and apoptosis . Together, these studies suggest that fibroblasts are unlikely to provide an origin of Ewing sarcoma. Moreover, as discussed above, most Ewing’s sarcomas appear to have a functional p53 pathway and to retain p19ARF expression. In an effort to identify a candidate primary cell from which ESFT originates, we expressed EWS-FLI-1 in murine cells with a variable spectrum of plasticity, ranging from embryonic stem (ES) cells and primary mesenchymal progenitor cells (MPC) to embryonic fibroblasts. Whereas ES cells and embryonic fibroblasts lost EWS-FLI-1 expression at the protein level by 14 days following infection with a retrovirus containing the fusion gene, bone marrow-derived MPCs maintained EWS-FLI-1 expression for several months . Mesenchymal progenitor cells expressing EWS-FLI-1 displayed robust upregulation of insulin-like growth factor 1 (IGF-1) and the corresponding binding proteins 3 and 5 (IGFBP-3 and IGFBP-5). Upon injection into mice, these cells formed tumors composed predominantly of sheets of small round cells. Gene expression profile analysis of tumor-derived cells revealed upregulation of known EWS-FLI-1 targets, including MYC and ID-2 and repression of p21 and TGFβRII both of which have been shown to be downregulated in Ewing sarcoma cells. Furthermore, these tumors displayed high sensitivity to IGF-1R inhibition, a hallmark of Ewing sarcoma, as well as expression of Ewing sarcoma-associated markers, including NSE and CD99. Importantly, MPC used in this study had not been immortalized prior to EWS-FLI-1 introduction and expressed functional p53 in addition to retaining the p16INK4A/p19ARF gene .