Site-specific mutagenesis in the analysis of a viral replicon.

Abstract

The small circular genome of Simian Virus 40 (SV40) is a relatively simple mammalian replicon, a DNA molecule that contains a signal for the origin of DNA replication and gene(s) for protein(s) involved in initiating DNA synthesis at the origin (Jacob and Brenner 1963). SV40 DNA replicates in the nucleus of infected monkey cells, replication commencing at a specific site in the DNA located in a regulatory segment of the viral genome between the start of early and late genes, and proceeding bidirectionally from that site (see Fig. 1) (Danna and Nathans 1972; Fareed et al. 1972). That SV40 DNA codes for a protein involved in the initiation of viral DNA replication was inferred from the observation that tsA mutants of SV40 are defective in initiation of DNA replication at the nonpermissive temperature (Tegtmeyer 1972). The mutational alterations in tsA mutants map in the gene for the SV40 T antigen (Lai and Nathans 1975). Direct interaction in vitro of T antigen with the SV40 replication origin region (Tjian 1978a, b) suggests that specific T antigen binding to the origin signal may be the initial event in SV40 DNA replication.