v-rel is the transforming gene of the avian reticuloendotheliosis virus, strain T (REV-T), which causes fatal lymphomas in young birds and also transforms avian lymphoid cells in vitro. The v-rel oncogene codes for a protein of 503 amino acids. The v-rel protein exhibits extensive homology within its N-terminal 300 amino acids with several mammalian $\kappa$B enhancer binding proteins including 50-kDa and 65-kDa subunits of NF-$\kappa$B transcription factor. In this study, an array of biochemical functions of mutant v-rel proteins was investigated in order to understand the mechanism of transformation by v-rel oncogene. Since the v-rel protein shows striking homology with NF-$\kappa$B subunits, wild-type v-rel or deletion and linker insertion mutants of v-rel were expressed in insect cells to determine the $\kappa$B-specific DNA binding activity. C-terminal deletions of up to 100 amino acids, or deletion of 11 N-terminal amino acids had no effect on DNA binding activity of v-rel protein, whereas N-terminal deletion of 99 amino acids abolished DNA binding activity completely. All of the mutant v-rel proteins with transforming activity also retained the DNA binding activity like wild-type v-rel protein, whereas all of the mutant v-rel proteins which were nontransforming lost DNA binding activity. Therefore, it is clear that the $\kappa$B-specific DNA binding activity is necessary for transformation by v-rel. All of the non-DNA binding, non-transforming mutants of v-rel exhibited a transdominant negative effect. These results suggest that the formation of DNA binding complexes by v-rel protein with NF-$\kappa$B subunits is necessary for transformation. In order to determine the possible role of pp40 association in transformation, pp40 association with v-rel protein was examined. Association of pp40 with the DNA binding mutant v-rel proteins including wild-type v-rel protein resulted in inhibition of DNA binding activity. These results imply that the physical association with pp40 itself might not be sufficient for transformation by v-rel. Since the v-rel protein directly binds to the $\kappa$B site, the $\kappa$B-dependent transcriptional activity of v-rel protein was investigated. The non-DNA binding, non-transforming mutants function as transdominant inhibitors of the rel family of transcription factors. These results support a model in which activation of gene expression directly by v-rel protein is required for its ability to induce oncogenic transformation. (Abstract shortened by UMI.)
| Identifer | oai:union.ndltd.org:uottawa.ca/oai:ruor.uottawa.ca:10393/10381 |
| Date | January 1996 |
| Creators | Park, Mahnhoon. |
| Contributors | Kang, C. Yong, |
| Publisher | University of Ottawa (Canada) |
| Source Sets | Université d’Ottawa |
| Detected Language | English |
| Type | Thesis |
| Format | 229 p. |
Page generated in 0.0342 seconds