Biology of SV40 transplantation antigen (TrAg): I. Demonstration of SV40 TrAg on glutaraldehyde-fixed SV40-infected African green monkey kidney cells

Abstract

The synthesis of papovavirus SV40 transplantation antigen (TrAg) in SV40-infected permissive African green monkey kidney cells (TC7) was demonstrated by using the in vivo transplantation-rejection test. Adult BALB c mice immunized with SV40-infected cells, which had been fixed with 0.25% glutaraldehyde for 5 min at room temperature, rejected a challenge of syngeneic virus-free SV40-transformed mouse cells (VLM). Fixed uninfected TC-7 cells failed to induce transplantation immunity against VLM cells in mice. The induction of SV40 tumor immunity in mice after immunization with glutaraldehyde-fixed infected TC-7 cells was mediated by the TrAg synthesized in permissive monkey cells as a result of virus infection, rather than by the residual infectious virus in the glutaraldehyde-fixed infected cells. The treatment of infectious SV40 with 0.25% glutaraldehyde drastically reduced both viral infectivity and tumor immunity-inducing ability of the virus. Glutaraldehyde-inactivated virus also failed to induce SV40 T antigen in monkey cells, thus ruling out any possibility that the inactivated cell-associated virus was responsible for the induction of tumor immunity in vivo. The amount of infectious SV40 required to induce detectable tumor immunity was found to be 2 × 10 5 PFU of virus administered to adult mice, and is far greater than the amount of virus (1 × 10 4 PFU) which remained associated with SV40-infected monkey cells after fixation of cells with glutaraldehyde. The synthesis of SV40 TrAg in SV40-infected TC-7 cells was inhibited by 5 μg/ml of actinomycin D, but not by 15 μg/ml of cytosine arabinoside, indicating that transcription in the infected cells, but not viral-DNA synthesis, is required for the expression of TrAg. By using 0.25% glutaraldehyde to fix infected cells, it should be possible to test available temperature-sensitive mutants of SV40 for their ability to synthesize TrAg at the nonpermissive temperature, as the glutaraldehyde fixation reduces infectivity of intracellular and extracellular virus, stops the infectious cycle at a particular stage, and preserves immunogenicity of TrAg.