Cell type-specific replication of simian virus 40 conferred by hormone response elements in the late promoter

The late genes of SV40 are not expressed at significant levels until after the onset of viral DNA replication. We previously identified two hormone response elements (HREs) in the late promoter that contribute to this delay. Mutants defective in these HREs overexpress late RNA at early, but not late, times after transfection of CV-1PD cells. Overexpression of nuclear receptors (NRs) that recognize these HREs leads to repression of the late promoter in a sequence-specific and titratable manner, resulting in a delay in late gene expression. These observations led to a model in which the late promoter is repressed at early times after infection by NRs, with this repression being relieved by titration of these repressors through simian virus 40 (SV40) genome replication to high copy number. Here, we tested this model in the context of the viral life cycle. SV40 genomes containing mutations in either or both HREs that significantly reduce NR binding without altering the coding of any proteins were constructed. Competition for replication between mutant and wild-type viruses in low-multiplicity coinfections indicated that the +1 HRE offered a significant selective advantage to the virus within a few cycles of infection in African green monkey kidney cell lines CV-1, CV-1P, TC-7, MA-134, and Vero but not in CV-1PD' cells. Interestingly, the +55 HRE offered a selective disadvantage in MA-134 cells but had no effect in CV-1, CV-1P, TC-7, Vero, and CV-1PD' cells. Thus, we conclude that these HREs are biologically important to the virus, but in a cell type-specific manner.     

Programmed factor binding to simian virus 40 GC-box replication and transcription control sequences

Nuclear footprinting revealed a temporal program involving factor binding to the repetitive GC-box DNA elements present in the simian virus 40 regulatory region. This program specified ordered and directional binding to these tandem regulatory sequences in vivo during the late phase of infection. The program was interrupted by the DNA replication inhibitor aphidicolin or by inactivation of the viral replication factor simian virus 40 T antigen, suggesting a link between viral DNA replication and new factor binding. Measurements of DNA accumulation in viruses lacking either the distal or proximal halves of the GC-box region suggested that the region has a dual role in replication control. Overall, the data point to important relationships between DNA replication and factor binding to the GC-box DNA, a multifunctional regulatory region.     

Interference with simian virus 40 DNA replication by adenovirus type 2 during mixed infection of monkey cells.

Infection of monkey cells with human adenovirus (Ad) is abortive, but the infection can be enhanced by coinfecting with simian virus 40 (SV40). However, in the coinfected monkey cells, Ad interferes strongly with SV40 DNA biosynthesis. This interference was found to be a reproducible, delicately controlled phenomenon that was proportional to the multiplicity of infection of Ad and dependent on the active expression of the Ad genome. Newly synthesized SV40 DNA was not broken down in cells after delayed superinfection with Ad, and several early events of SV40 infection such as adsorption, penetration, uncoating, induction of cellular DNA synthesis, and enhancement of Ad infection were not markedly influenced by Ad-mediated interference. It is unlikely that interference is simply due to competition between SV40 and Ad for metabolites, enzymes, or replication sites. The interference effect could be partially neutralized by an increase in the multiplicity of coinfecting SV40 or by an increase in the time interval between SV40 infection and Ad coinfection. Interference was shown to be due to the activity of an Ad early gene product. However, the detailed mechanism of this Ad interference is still unclear.     

Protein-DNA interactions at the simian virus 40 origin of replication

Simian Virus 40 (SV40)-encoded large T antigen has an intrinsic ATP-dependent DNA-unwinding activity which is necessary for an early step in the activation of the viral origin of replication. Isolated T antigen unwinds any double-stranded DNA, regardless of whether it is linear or circularly closed. However, initiation of DNA replication depends on an intact origin of replication, and even minor deviations from the wild-type origin sequence abolish the template activity of an origin-bearing plasmid. This discrepancy suggests that T antigen may not be sufficient for origin activation and that other, probably cellular, functions are involved. We have isolated a cellular protein, the LOB protein, which specifically interacts with the AT-rich region of the SV40 origin and which induces a pronounced bending of the bound DNA.