The SV40 large tumor (T) antigen is both necessary and sufficient to transform growing primary cells. Stable expression of this single protein restructures the molecular program of normal cells. T antigen induces a panoply of altered growth characteristics that culminate in tumorigenic cell lines in culture or emerging tumors in experimental animals. We recently showed that discrete regions of T antigen confer or are required for expression of the individual growth patterns that collectively characterize the transformed cell phenotype. This genetic dissection of the transforming regions of T antigen holds the promise of identifying, by virtue of colinearity, functions involved in each cell growth alteration. By using immortalization of C57B1/6 mouse embryo fibroblasts (B6/MEF) as a selective assay, and characterizing individual cell lines for other growth characteristics, we identified a broad segment of T antigen (amino acids 251-625) that is sufficient for immortalization and tumorigenicity in nude mice. We also showed that amino acids 1-126 appear to be sufficient to confer loss of contact inhibition, and that anchorage independent growth and growth in medium with reduced concentrations of growth factors require integrity of the T antigen segment containing amino acids 1-250. Functions within each region are implicated in individual cell growth patterns. We propose to define more precisely the regions and functions of T antigen that are sufficient to confer each of the transformed cell growth characteristics. For these analyses, three types of probes will be used: (1) a series of in-frame deletion mutations that remove sequential fifty codons between 250 and 650; (2) overlapping in-frame deletion mutations generated from libraries of deletion-linker insertion mutations; and (3) fusion proteins containing specific T antigen regions. Coordinately, we will define and genetically isolate the portions of T antigen that are sufficient to stimulate host DNA synthesis, transactive heterologous promoters, reactivate silent ribosomal RNA genes, and bind the anti- oncogene product 53. Experiments should ultimately determine the individual and combined roles of these activities, which are located within the immortalization and tumor-forming region, as well as the significance binding the retinoblastoma susceptibility gene product (Rb), in transformed cell growth, generation of tumors from transformed cell lines, and induction of preneoplastic and neoplastic disease in transgenic mice.
| Tatum, Angela M; Mylin, Lawrence M; Bender, Susan J et al. (2008) CD8+ T cells targeting a single immunodominant epitope are sufficient for elimination of established SV40 T antigen-induced brain tumors. J Immunol 181:4406-17 |
