DNA sequences outside the simian virus 40 early region cause downregulation of T-antigen production in permissive simian cells.

Using a series of modified wtSV40 and early region SV40 DNAs we assayed the effect of viral late region sequences on T-antigen production by the SV40 early region. We found that SV40 late region (L-SV40) DNA sequences reduced T-antigen (T-Ag) production by the SV40 early region (E-SV40) when both viral regions were linked as they are in wtSV40 DNA. This was demonstrated by Western analysis which showed that E-SV40 DNA produced 10 times more T-Ag than wtSV40 DNA L-SV40, with its own promoter but unlinked to E-SV40 DNA, also greatly inhibited T-Ag production when it was contrasfected with E-SV40. Therefore, L-SV40 DNA inhibited T-Ag production by E-SV40 DNA when present in cis or in trans. We have shown that expression of the SV40 late transcription unit dominated that of the early (T-Ag gene) transcription unit because late region RNA accumulated to much higher levels than early viral RNA. However, in contrasfected cells L-SV40 DNA did not replicate to higher levels than E-SV40 DNA. We offer a model for control of T-Ag expression in which a relatively small amount of T-Ag activates late transcription at the expense of T-Ag gene transcription and that this represents a switch from early to late viral gene expression. We suggest that when activation of the late transcription unit occurs at the late promoter, expression of the T-Ag gene is greatly reduced. The L-SV40 promoter may inhibit T-Ag gene transcription by sequestering cellular factors required for early transcription, factors which may be present in limited amounts. We suggest further that activation of late transcription allows for the necessary production of large amounts of capsomeres and virions and downregulation of early transcription prevents the early region from interfering with capsid synthesis. We tested the model using a construct with a wild-type T-Ag gene but with mutations in the SV40 major late promoter which prevent the promoter from being bound by cellular repressors of late transcription. We found that this construct, which overproduces late SV40 RNA, was defective for T-Ag production. This indicates that activation of the late promoter results in repression of T-Ag gene expression.