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SV40 chromatin obtained from infected monkey cells was used to study the effect of the viral chromatin proteins on endogenous RNA polymerase II. Ammonium sulfate activated the rate of transcription by endogenous RNA polymerase in two ways: 1) by direct action on the enzyme; and 2) by causing a reversible conformational change in the viral chromatin. Under optimal reaction conditions, the viral chromatin proteins did not limit the rate of RNA chain elongation, and high molecular weight RNA (1.6 X 10(6) d) was synthesized by the SV40 chromatin.

The template activities for the processes of replication and transcription were compared for recently replicated ("new") and uniformly labeled ("old") simian virus 40 (SV40) DNA in infected monkey cells (line TC7). New SV40DNA (pulselabeled for 1 h) served as a template for a second round of replication withl a relatively high probability (8% of the DNA replicated per h) for a period of 5 h, after which time its template activity rapidly decreased by severalfold. Old SV40 DNA (labeled for 24 h) functioned as a template for replication with a constant probability (1.8% of the DNA replicated per 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 SV40 transcription complex. The proportion of RNA polymerase associated with nonreplicated SV40 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 SV40 DNA. Furthermore, after 9 h in bromodeoxyuridine, the recently replicated viral DNA had nearly three times more RNA polymerase per unit of DNA-than did the nonreplicated DNA. We thus conclude that recently replicated SV40-DNA is utilized preferentially as a template for transcription and for replication. Although it has been recognized for many ently than prereplicative templates. This specific years that changes occur in the transcriptional hypothesis was negated for SV40 by the recent program of bacterial and animal viruses after report of Birkenmeier et al. (2), which strongly

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ABSTRACT.A small fraction of the SV40 chromatin isolated from infected monkey cell cultures by the Triton method contains active RNA polymerase which had initiated transcription in vivo. This viral transcription complex (VTC) was utilized to answer the question of whether proteins dissociate from chromatin during transcription in vitro. 3H-RNA was synthesized by the VTC under conditions such that over half the label was in transcripts which were longer than half the length of the SV40 genome. Virtually all of the 3H-RNA remained associated with the SV40 chromatin, causing an increase in sedimentation rate from 55S to 78S. The density of the VTC-3H-RNA complex indicated that less than 5% of the original protein dissociated from the SV40 DNA which served as a template for transcription. We conclude that SV40 chromatin can be transcribed while the proteins remain associated with the DNA.INTRODUCTION.