SIMIAN-VIRUS-40 LARGE T-ANTIGEN AFFECTS THE SACCHAROMYCES-CEREVISIAE CELL-CYCLE AND INTERACTS WITH P34(CDC28)

Simian virus 40 tumor (T) antigen, an established viral oncoprotein, c auses alterations in cell growth control through interacting with, and altering the function of, cellular proteins. To examine the effects o f T antigen on cell growth control, and to identify the cellular prote ins with which it may functionally interact, T antigen was expressed i n the budding yeast Saccharomyces cerevisiae. The yeast cells expressi ng T antigen showed morphological alterations as well as growth inhibi tion attributable, at least in part, to a lag in progression from G(1) to S. This point in the cell cycle is also known to be affected by T antigen in mammalian cells. Both p34(CDC28) and p34(CDC2Hs) were shown to bind to a chimeric T antigen-glutathione S-transferase fusion prot ein, indicating that T antigen interacts directly with cell cycle prot eins which control the G(1) to S transition. This interaction was conf irmed by in vivo cross-linking experiments, in which T antigen and p34 (CDC28) were coimmunoprecipitated from extracts of T-antigen-expressin g yeast cells. These immunoprecipitated complexes could phosphorylate histone H1, indicating that kinase activity was retained. In addition, in autophosphorylation reactions, the complexes phosphorylated a nove l 60-kDa protein which appeared to be underphosphorylated (or underrep resented) in p34(CDC28)-containing complexes from cells which did not express T antigen. These results suggest that T antigen interacts with p34(CDC28) and alters the kinase function of p34(CDC28)-containing co mplexes. These events correlate with alterations in the yeast cell cyc le at the G(1) to S transition.