Replication of simian virus 40 origin-containing DNA during infection with a recombinant Autographa californica multiple nuclear polyhedrosis virus expressing large T antigen

Abstract

Autographica californica multiple nuclear polyhedrosis virus (AcMNPV) has been shown to encode many of the enzymes involved in the replication of its own DNA. Although the AcMNPV genome contains multiple sets of reiterated sequences that are thought to function as origins of DNA replication, no initiator protein has yet been identified in the set of viral replication enzymes. In this study, the ability of a heterologous origin initiator system to promote DNA replication in AcMNPV-infected cells was examined. A recombinant AcMNPV that expressed the simian virus 40 (SV40) large T antigen was surprisingly found to induce the efficient replication of a transfected plasmid containing an SV40 origin. This replication was subsequently found to involve three essential components: (i) T antigen, since replication of SV40 origin-containing plasmids was not induced by wild-type AcMNPV which did not express this protein; (ii) an intact SV40 core origin, since deletion of specific functional motifs within the origin resulted in a loss of replicative abilities; and (iii) one or more AcMNPVencoded proteins, since viral superinfection was required for plasmid amplification. Characterization of the replicated DNA revealed that it existed as a high-molecular-weight concatemer and underwent significant levels of homologous recombination between inverted repeat sequences. These properties were consistent with an AcMNPV-directed mode of DNA synthesis rather than that of SV40 and suggested that T antigen-SV40 origin complexes may be capable of initiating DNA replication reactions that can be completed by AcMNPVencoded enzymes.

Replication, Integration, and Packaging of Plasmid DNA following Cotransfection with Baculovirus Viral DNA

“…These data are in full agreement with our previous results which demonstrated that nonspecific plasmid replication took place even in the presence of virus infection, if the infection occurred within about 4 h of plasmid transfection (42). In addition, our data are consistent with previous observations that the baculovirus DNA replication machinery supported the replication of plasmids carrying the simian virus 40 (SV40) origin in infected insect cells expressing the large T antigen (25). Apparently, when SV40 T antigen was synthesized, it recognized the SV40 origin on plasmid DNA, and replication, coupled with recombination, occurred in the presence of the baculovirus replication machinery.…”

Nucleocapsid Assembly of Baculoviruses

“…Recombination-dependent replication plays an essential role in the replication cycle of many dsDNA viruses, especially DNA viruses with large genomes. Baculoviruses produce high molecular weight DNA with an apparently branched structure [15]. A sequence alignment revealed that all baculoviruses encode a gene (alkaline nuclease) homologous to the lambda red α exonuclease in bacteriophages, which is a typical exonuclease in the Red recombination system [16].…”