The three transforming regions of SV40 T antigen are required for immortalization of primary mouse embryo fibroblasts

• Published in: Oncogene Vol. 11; no. 11; p. 2295
• Main Authors: Conzen, S D, Cole, C N
• Format: Journal Article
• Language: English
• Published: England 07.12.1995
• Subjects: Animals; Antigens, Polyomavirus Transforming - genetics; Antigens, Polyomavirus Transforming - metabolism; Cell Division - genetics; Cell Transformation, Neoplastic - genetics; Cellular Senescence - genetics; Embryo, Mammalian - cytology; Embryo, Mammalian - metabolism; Fibroblasts - metabolism; Mice; Mutation; Protein Binding; Transfection; Tumor Suppressor Protein p53 - metabolism
• ISSN: 0950-9232

Simian virus 40 (SV40) is a small DNA tumor virus whose early region gene product, large T antigen, is sufficient to immortalize primary rodent cells and transform established rodent cell lines. Three functional domains of large T antigen are required for transformation of the rat embryo fibroblast REF 52 cell line: the extreme amino-terminal region, a domain which binds p105Rb family members, and the bipartite p53-binding region. Many studies have attempted to define the activities and regions of SV40 large T antigen required for immortalization of mouse embryo fibroblasts (MEFs). In most of these studies, investigators have used survival of T antigen-expressing primary MEF colonies at the time when controls MEFs undergo senescence as a measurement of 'immortalization' and concluded that immortalization of MEFs is correlated with large T antigen's ability to sequester the human tumor suppressor gene product p53 and separable from its p105Rb-binding or N terminal functions. In order to more rigorously define the regions of SV40 large T antigen required for escape from senescence, individual T antigen-expressing primary MEF colonies were systematically subcultured for > 60 population doublings beyond the time of control MEF senescence under conditions known to limit the number of spontaneously immortalized cells. We found that although interaction of T antigen with p53 was sufficient to substantially extend the lifespan of MEFs, all three SV40 large T antigen domains required for REF 52 transformation were necessary to immortalize primary MEFs. These results indicate that p53 inactivation alone is insufficient to immortalize primary MEFs; rather, immortalization requires multiple activities of T antigen which are also required for efficient transformation.