THE EFFECTS OF SV40 LARGE T-ANTIGEN AND P53 ON NUCLEAR TRANSPORT CAPACITY IN BALB C 3T3 CELLS/

In a previous study, it was found that the microinjection of purified SV40 large T antigen into the cytoplasm of BALB/c 3T3 cells significan tly increased both the relative rate of signal-mediated nuclear transp ort and the functional size of the transport channels that are located within the pores. In this investigation, a series of mutants were emp loyed to identify the region of large T responsible for the permeabili ty increase. Plasmids encoding wild-type or mutant forms of large T we re injected into the nucleoplasm of proliferating BALB/c 3T3 cells, an d the nuclear import of nucleoplasmin-coated gold particles was analyz ed approximately 18 h later. The large T mutants that were not effecti ve in inducing the increase in nuclear transport capacity were also un able to bind p53. Further evidence that transport activity and p53 bin ding localize to the same region of large T was obtained by simultaneo usly injecting plasmids that overexpress wild-type or mutant p53 and p lasmids that encode active forms of large T. It was found that wild-ty pe p53 prevented the large T-induced transport increase; however, muta nt p53, which is unable to bind to large T, had no effect. Decreasing the concentration of endogenous p53 in cells that do not contain large T, by injecting anti-p53 antibodies or plasmids that express mutant p 53, resulted in a significant increase in the nuclear import of nucleo plasmin-coated gold. The latter results suggest that p53 might normall y act as a transport suppressor. (C) 1994 Academic Press, Inc.