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. 1979 May;30(2):497–507. doi: 10.1128/jvi.30.2.497-507.1979

Protein synthesized early after infection is linked to the termini of adenovirus type 2 DNA synthesized in vivo and in vitro.

T Yamashita, M Arens, M Green
PMCID: PMC353353  PMID: 224213

Abstract

The human adenovirus DNA genome contains a protein (CBP, or covalently bound protein) linked to each 5' terminus. To assess whether CBP is synthesized early, infected cells were incubated with hydroxyurea from 1 to 18 h postinfection, the hydroxyurea was removed, cycloheximide was added, and viral DNA was labeled with [3H]thymidine from 18 to 23 h postinfection. Removal of hydroxyurea at 18 h postinfection permits the synthesis of viral DNA, whereas cycloheximide maintains the block in late viral protein synthesis. Three lines of evidence are presented to show that viral 3H-labeled DNA prepared by this procedure was linked to CBP: (I) the DNA sedimented more rapidly than protein-free DNA (i.e., protinase treated) in neutral sucrose gradients containing guanidine hydrochloride; (ii) the DNA banded at a lower density than protein-free DNA in CsCl gradients containing guanidine hydrochloride; and (iii) neither the 3H-labeled DNA nor the end fragments produced by EcoRI digestion entered a 1.4% agarose gel during electrophoresis. These experiments are strong evidence that CBP is not a product of a late viral gene and is therefore the product of either an early viral gene or a cell gene. Experiments were performed to test whether CBP is attached to viral DNA synthesized in vitro by a soluble complex that synthesizes exclusively viral DNA as completed viral genomes in vitro. In vitro-labeled DNA was analyzed by velocity sedimentation, equilibrium sedimentation, and agarose gel electrophoresis as described above. Our results indicate that the majority of in vitro-synthesized DNA molecules were attached to CBP. These results, which indicate that CBP is synthesized early after infection and is attached to viral DNA labeled in vitro by a soluble replication complex, are consistent with the idea that CBP may play a role in viral DNA replication.

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Selected References

These references are in PubMed. This may not be the complete list of references from this article.

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