THESIS
2002
xi, 85 leaves : ill. (some col.) ; 30 cm
Abstract
The N-terminal region of the Ewings Sarcoma Oncogene (EWS-Activation-Domain, EAD) is fused to a variety of transcription factors by chromosome translocation and the fusion proteins produced (EFPs) are oncogenes. The EAD is a potent transcriptional activation domain, so study of EAD action is of significance to transcriptional mechanism and tumorigenesis. In addition, EFPs play a role in tumor maintenance, indicating that EAD inhibitors may have therapeutic potential....[
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The N-terminal region of the Ewings Sarcoma Oncogene (EWS-Activation-Domain, EAD) is fused to a variety of transcription factors by chromosome translocation and the fusion proteins produced (EFPs) are oncogenes. The EAD is a potent transcriptional activation domain, so study of EAD action is of significance to transcriptional mechanism and tumorigenesis. In addition, EFPs play a role in tumor maintenance, indicating that EAD inhibitors may have therapeutic potential.
The primary structure of the EAD is highly repetitive, containing 31 copies of a dispersed Degenerate Hexapeptide Repeats (DHR, consensus SYGQQS), in which tyrosine is the only absolutely conserved residue. The previous data in our lab indicated that DHRs may play an important role in the function of EAD. To date, the DHRs have not been systematically studied due to lack of a sensitive assay for the small region of the EAD with limited number of DHRs.
In my work, an assay has been set up to solve this problem. Based on this, systematic mutational analysis was carried out. The results showed that: 1. DHRs play a major role in EAD-mediated trans-activation and they function cooperatively to activate the transcription. 2. The tyrosine in DHRs is critical for trans-activation. 3. EAD is a novel tyrosine dependent transcriptional activation domain. These findings, for the first time, provide the evidences for the role of the DHRs in trans-activation and the mechanism of EAD action.
The above results identify a small peptide motif, DHR, which might represent an EAD inhibitor or might be an effective target for rational drug design. I have attempted to establish an assay that will allow testing of the inhibitory potential of DHR related peptides (or other small molecules). By exploring several methods, my results indicate that it is feasible to effectively deliver peptides and monitor EAD-mediated trans-activation in a well defined, homogenous, cell system.
Overall, my study has provided useful reagents (a series of mutants) and a firm platform for the future molecular study of the EAD.
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