An adenovirus type 5 gene function required for initiation of viral DNA replication

Abstract

Adenovirus type 5 (Ad5) DNA replication was studied after infection of human or monkey cells with two DNA-negative temperature-sensitive mutants belonging to different complementation groups (H5ts125 and H5ts36). When infection was carried out at the permissive temperature (32°) followed by a shift to the nonpermissive temperature (39.5°) viral DNA synthesis in H5ts125-infected cells was reduced 90% within 1 hr after shift-up, while a decline in DNA synthesis in H5ts36-infected cells is only observed after 6 hr. Analysis of the various forms of DNA synthesized under conditions of inhibition showed a constant ratio of replicating to mature viral DNA for both mutants, while no accumulation of replicating molecules was observed.

When H5ts125-infected cells were pulse-labeled with [3H]thymidine at 32 or 39.5° followed by a chase of the label at 39.5°, replicating DNA was converted into mature DNA at the same rate as in wild-type-infected cells. This indicates that chain propagation and termination could occur normally under nonpermissive conditions. The results of density labeling experiments performed at 39.5° are in agreement with an initiation block in H5ts125-infected cells at the nonpermissive temperature. It is concluded that the H5ts125 gene product and possibly also the H5ts36 gene product are required for the initiation of new rounds of replication. The potential role in initiation of the adenovirus-specific DNA binding protein, which is coded for by the H5ts125 gene, is discussed.