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EWS/FLI1 up regulates mE2-C, a cyclin-selective ubiquitin conjugating enzyme involved in cyclin B destruction
A Arvand1, H Bastians, S M Welford
1Department of Experimental Pathology/Laboratory Medicine, University of California, Los Angeles, USA.
Abstract:
The EWS/FLI1 fusion gene found in Ewing's sarcoma and primitive neuroectodermal tumor, is able to transform certain cell lines by acting as an aberrant transcription factor. The ability of EWS/FLI1 to modulate gene expression in cells transformed and resistant to transformation by EWS/FLI1, was assessed by Representational Difference Analysis (RDA). We found that the cyclin selective ubiquitin conjugase murine E2-C, was up regulated in NIH3T3 cells transformed by EWS/FLI1 but not in a nontransformed NIH3T3 clone expressing EWS/FLI1. We also found that mE2-C is upregulated in NIH3T3 cells transformed by other genes including activated cdc42, v-ABL and c-myc. We demonstrated that expression of mE2-C in both the EWS/FLI1 transformed and parent NIH3T3 lines varies with the cell cycle. Finally, dominant-negative mE2-C, created by changing a catalytic cysteine to serine, inhibits the in vitro ubiquitination and degradation of cyclin B in human HeLa cell extracts. These data suggest that part of the biologic effect of EWS/FLI1 could be to transcriptionally modulate genes involved in cell cycle regulation.
Insights
The EWS/FLI1 fusion gene upregulates murine E2-C, a protein involved in cell cycle regulation. This suggests EWS/FLI1 contributes to cancer development by altering cell cycle control.
Area of Science:
- Oncology
- Molecular Biology
- Cell Biology
Background:
- The EWS/FLI1 fusion gene is a key driver in Ewing's sarcoma and primitive neuroectodermal tumors.
- EWS/FLI1 functions as an aberrant transcription factor, driving cellular transformation.
Purpose of the Study:
- To investigate how EWS/FLI1 modulates gene expression in transformed and non-transformed cells.
- To identify specific genes regulated by EWS/FLI1 that contribute to its oncogenic effects.
Main Methods:
- Representational Difference Analysis (RDA) was employed to compare gene expression profiles.
- Cell cycle analysis and in vitro ubiquitination assays were performed.
Main Results:
- Murine E2-C (mE2-C), a cyclin-selective ubiquitin conjugase, was found to be upregulated in EWS/FLI1-transformed NIH3T3 cells.
- mE2-C upregulation was also observed in cells transformed by other oncogenes (cdc42, v-ABL, c-myc) and varied with the cell cycle.
- A dominant-negative form of mE2-C inhibited cyclin B ubiquitination and degradation.
Conclusions:
- EWS/FLI1 transcriptionally modulates genes involved in cell cycle regulation, such as mE2-C.
- Altered cell cycle control via mE2-C is a potential mechanism contributing to the biological effects of EWS/FLI1 in sarcomagenesis.