Bovine RP-A Functions in SV40 DNA Replication in vitro, but Bovine Polymerase α-Primase Inhibits Replication Competitively

Abstract

The replication of SV40 DNA in a cell-free assay (Li and Kelly, 1984) containing primate cell extract has facilitated the identification and purification of a set of cellular replication proteins that, in concert with a single viral regulatory protein T antigen, are sufficient to replicate SV40 origin-containing DNA (Tsurimoto et al., 1990; Weinberg et al., 1990). Origin binding and unwinding to form a pre-initiation complex at the origin requires T antigen and the cellular single-stranded DNA binding protein RP-A (RF-A) (reviewed by Borowiec et al., 1990). The initial primer synthesis is thought to be catalyzed by DNA polymerase α-primase, but how the establishment of the pre-initiation complex is coupled to synthesis of RNA primers and DNA remains speculative. Protein-protein interactions among the initiation proteins, as observed in procaryotic model systems (reviewed by Bramhill and Kornberg, 1988), could serve to couple origin unwinding and DNA synthesis. Indeed, an association between T antigen and DNA polymerase a-primase in crude extracts of both primate and rodent cells has been reported (Smale and Tjian, 1986; Gough et al., 1988; Gannon and Lane, 1987, 1990).