Role of DNA polymerase .gamma. in adenovirus DNA replication. Mechanism of inhibition by 2',3'-dideoxynucleoside 5'-triphosphates

Abstract

In contrast to cellular or SV40 DNA replication, adenovirus type 5 (Ad5) or type 2 (Ad2) DNA synthesis in isolated nuclei is strongly inhibited by low concentrations of 2',3'-dideoxythymidine 5'-triphosphate (ddTTP). On the basis of differential sensitivity of cellular DNA polymerases, a role of DNA polymerase gamma in adenovirus DNA replication has been proposed. We have investigated the mechanism of inhibition of adenovirus DNA synthesis, using [alpha-32P]ddTTP and other dNTP analogues. Both ddATP and ddGTP were as inhibitory as ddTTP, while ddCTP had an even stronger effect on adenovirus DNA replication. DNA polymerase alpha was resistant to all four ddNTP's, while DNA polymerase gamma was very sensitive. The inhibition by ddTTP in isolated infected nuclei was slowly reversible. [alpha-32P]ddTTP was incorporated into Ad5 DNA as a chain-terminating nucleotide, and the analogue could be used as a substrate by DNA polymerase gamma. Under similar conditions, incorporation in cellular DNA or using DNA polymerase alpha was not observed. The nucleoside analogues ddA and ddC suppressed adenovirus. DNA replication in intact cells and reduced plaque formation. These results provide further evidence for a function of DNA polymerase gamma in adenovirus DNA synthesis.

Antiadenovirus Activities of Several Classes of Nucleoside and Nucleotide Analogues

“…An intriguing finding is the inhibition of adenovirus DNA synthesis by 2Ј,3Ј-dideoxynucleoside analogues, as reported for zalcitabine (ddC) more than 20 years ago (27), and further documented by Mentel et al (19). At this time, alovudine (FddT) is in phase II trials for human immunodeficiency virus (HIV) therapy.…”

Anti-adenoviral effect of anti-HIV agents in vitro in serotypes inducing keratoconjunctivitis

“…Several nucleoside analogues have been discovered to have antiviral activity against AdV by inhibiting viral DNA polymerase-mediated DNA replication in AdV-infected cells [24,36]. Nucleoside analogues require activation by cellular enzymes to exert their antiviral activity [29].…”

Inhibitory effect of 2',3'-didehydro-2',3'-dideoxynucleosides on infectivity, cytopathic effects, and replication of human immunodeficiency virus

“…Our experiments showed that there seems to be no direct effect on the virus particles by these nucleoside analogs (data not shown). It is known that the 5'-triphosphates of dideoxynucleosides, such as 2' ,3'-dideoxyadenosine, -dideoxyguanosine, -dideoxycytidine, and -dideoxythymidine, can rather easily inhibit cellular DNA polymerases 1B and -y, as well as viral reverse transcriptase, but no mammalian DNA polymerase a (2,15,(18)(19)(20). Both DHT and DHC are also 2',3'-dideoxynucleosides and have double bonds between the 2' and 3' positions.…”