Recently replicated simian virus 40 DNA is a preferential template for transcription and replication

Abstract

The template activities for the processes of replication and transcription were compared for recently replicated (new) and uniformly labeled (old) SV-40 DNA in infected [African green] monkey [kidney] cells (line TC7). New SV-40 DNA (pulse-labeled for 1 h) served as a template for a 2nd round of replication with a relatively high probability (8% of the DNA replicated/h) for a period of 5 h, after which time its template activity rapidly decreased by several-fold. Old SV-40 DNA (labeled for 24 h) functioned as a template for replication with a constant probability (1.8% of the DNA replicated/h) for at least 12 h. The proportion of RNA polymerase with nonreplicated and with recently replicated (bromodeoxyuridine-substituted) viral DNA was determined by an assay that used the Triton-soluble SV-40 transcription complex. The proportion of RNA polymerase associated with nonreplicated SV-40 DNA decreased very slowly (to 50% in 6 h), strongly suggesting that replicating viral genomes are not required as templates for the initiation of late transcription. This hypothesis was supported by the finding that the RNA synthesized in vitro was associated with covalently closed circular SV-40 DNA. After 9 h in bromodeoxyuridine, the recently replicated viral DNA had nearly 3 times more RNA polymerase/unit of DNA than did the nonreplicated DNA. Recently replicated SV-40 DNA is apparently utilized preferentially as a template for transcription and for replication.