Adenovirus-2 DNA: In Vitro Replication and Analysis of Viral Early Proteins

Abstract

The synthesis of DNA is complex, involving numerous enzymes and other proteins that may function in multienzyme systems. Animal DNA viruses (adenoviruses, papovaviruses, herpesviruses, poxviruses, parvoviruses) provide simple, powerful systems for studying eukaryotic DNA replication. We have chosen adenoviruses (Ads) because cells that replicate Ad genomes are particularly useful models for studying DNA replication and its regulation for several reasons: (1) Host-cell DNA synthesis is blocked late after infection with human Ads, thus permitting the unambiguous analysis of viral DNA synthesis. (2) Isolated nuclei and subnuclear complexes have been prepared from Ad2-, Ad5-, and Ad12-infected cells that synthesize almost exclusively viral DNA sequences in vitro. (3) DNA-negative Ad2, Ad5, and Ad12 temperature-sensitive (ts) mutants are available that are very useful in understanding DNA replication. (4) With the exception of several virus-coded proteins, Ad DNA replication involves mainly cellular enzymes and other proteins, and thus the study of Ad DNA replication...