Dissociation of Rb-binding and anchorage-independent growth from immortalization and tumorigenicity using SV40 mutants producing N-terminally truncated large T antigens

Abstract

The large T antigen of SV40 is both necessary and sufficient for conversion of primary mouse cells to cells with fully transformed phenotype. In this investigation, the influence of the N-terminal portion of T antigen on individual transformed cell characteristics was probed by using mutants bearing deletions in the 5'T antigen coding sequence. Specifically, DNA constructs expected to produce T antigens missing the first 109, 127, 150, or 176 amino acids or internal amino acid segments between 117 and 250 were tested for the ability to immortalize C57Bl/6 mouse embryo fibroblasts. The transformed cell properties displayed by clonally derived cell lines were then examined. The results indicated that neither the first 127 amino acids nor amino acids 127-250 of T antigen were necessary for efficient immortalization of primary cells or for their tumorigenicity. Functions mapped within these regions, including binding of the retinoblastoma susceptibility gene product (Rb) and transactivation of heterologous promoters, therefore, were not required to confer either of these growth properties. In addition the results showed that anchorage-independent growth was separable genetically from tumorigenicity and that removal of amino acids within the first 250 residues of T antigen compromised other transformed cell growth properties.