Recombinant adenoviruses that express high levels of the simian virus 40 (SV40) small-t (ST) antigen have been used to study the requirement for ST to drive cell cycle proliferation of confluent human diploid fibroblasts. This occurs when either large-T (LT) antigen or serum is added to provide a second signal. While cells readily completed S phase in these experiments, they were found to accumulate with 4N DNA content. Cellular and nuclear morphology, as well as the biochemical status of cyclin B complexes, showed that these cells entered mitosis but were blocked prior to mitotic metaphase. The defect appears to reflect an inability of cells overexpressing ST to form organized centrosomes that duplicate and separate normally during the cell cycle and, therefore, the absence of a mitotic spindle. The ability of ST to bind protein phosphatase 2A was required for this pattern, suggesting that altered phosphorylation of key centrosomal components may occur when ST is overexpressed. Although the possible significance of ST effects on the centrosome cycle is not fully understood, these findings suggest that ST could influence chromosomal instability patterns that are a hallmark of SV40-transformed cells and LT expression.The early region of simian virus 40 (SV40) encodes three proteins that can be detected in nonpermissive, transforming infections (44, 50). The best understood of these is the large-T (LT) antigen, which binds tumor suppressor proteins p53 and pRb and has DNA binding, helicase, and transactivation capabilities (reviewed in references 12, 22, and 32). An aminoterminal dnaJ domain is also required for viral DNA replication and transformation (40,49). In LT, the dnaJ domain modulates the protein stability of p130 and p107, members of the pRb family. (41). A key function of the small-t (ST) antigen is its binding to protein phosphatase 2A (PP2A). ST mimics cellular regulatory B subunits (28,29,48) of this trimeric enzyme and, presumably, modifies the substrate specificity and intracellular localization of PP2A (36, 39). ST expression in primary cells results in activation of key cellular kinases and growth regulators, such as mitogen-activated protein kinase, its kinase MEK, and the ion transporter, the Na-H antiporter (19,38). These enzymes are all more highly phosphorylated in the presence of ST, consistent with an inhibition of phosphatase activity against these target molecules.ST enhances the efficiency of virus transformation and tumor formation in animal model systems. A role for ST in hamsters (2) or transgenic mice is particularly apparent in nondividing tissues (5), consistent with the general concept that ST enhances cell cycle progression. Considerable evidence to support this concept came from early tissue culture studies (18, 23), one of which showed that a few rounds of cell division could bypass the ST requirement in a hamster cell system (23).ST is not required for the transformation of all cell types in cultures. However, whenever ST is required, its ability to interact with PP2A has pro...
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