The onset of DNA replication is an important step within the life cycle of the human neurotropic polyomavirus JC. In this report, evidence that both the human and the murine tumor suppressor protein p53 strongly inhibit JCV DNA replication in vivo is presented. This inhibition is dose-dependent and not a secondary effect of a decreased expression of JCV large T-antigen in response to p53. Using deletion mutants of murine p53 and tumor-derived point mutations of human p53, the basis of the suppression of JCV DNA replication by p53 was dissected. Deletion of either the amino- or the carboxy-terminal domain of murine p53 did not interfere with the repression of JCV DNA replication. However, deletion of the highly conserved central region of p53 abolished the inhibitory effect on replication. The tumor-derived human mutant p53(His273) inhibited JCV DNA replication significantly, whereas another tumorigenic mutant, p53(His175), had no inhibitory effect Concomitantly, a direct protein-protein interaction between p53 and JCV large T-antigen was lost in mutants which did not affect JCV DNA replication. These results strongly suggest that p53 inhibits JCV DNA replication by interacting with JCV large T-antigen.

Interleukin 2 (IL-2) and interleukin 4 (IL-4) secreted by activated but not by resting mature T cells are pleiotropic cytokines affecting growth and differentiation of diverse cell types, such as T cells, B cells, and mast cells. There is little information about the molecular basis for the constitutive repression of IL-2 and IL-4 gene expression in unstimulated T cells. We investigated the possibility that wild-type (wt) p53, a nuclear tumor suppressor protein, might serve to repress IL-2 and IL-4 gene expression in murine E14 T lymphoma and in human Jurkat cells. We transiently cotransfected these cells with constitutive simian virus 40 (SV 40) early promoter expression plasmids overproducing wt or mutant murine p53 and with appropriate luciferase (luc) reporter plasmids containing the promoter elements of murine IL-2 and IL-4 genes to evaluate the effect of various p53 species on these promoters. Murine wt p53 derived from pSG5p53cD strongly repressed the IL-2 and IL-4 promoters in both cell lines induced by the phorbol ester TPA and the Ca2+ ionophore ionomycin but not, however, in uninduced cells. In similar transient transfection experiments with lymphoma cells, overexpression of deletion mutant species of murine p53 revealed that the N-terminal and C-terminal domains are crucial for inhibition of both IL-2 and IL-4 gene expression. These parts of p53 comprise the transactivation domain at the amino terminal side, which has previously also been shown to interact with the TATA-box binding-protein TBP and the carboxy-terminal oligomerization domain. Additionally, it was shown that a previously described inhibitory protein, the high-mobility-group protein HMG-I/Y, does not functionally interact with p53. Cotransfection of expression plasmids for both p53 and HMG-I/Y did not alter the extent of inhibition by the individual proteins. These data suggest that p53 can downmodulate both IL-2 and IL-4 gene expression and that both the transactivation and oligomerization domains of the tumor suppressor protein are essential for this transcriptional repression.