SEQUENCES FLANKING THE PENTANUCLEOTIDE T-ANTIGEN BINDING-SITES IN THEPOLYOMAVIRUS CORE ORIGIN HELP DETERMINE SELECTIVITY OF DNA-REPLICATION

Replication of the genomes of the polyomaviruses requires two virus-sp ecified elements, the cis-acting origin of DNA replication, with its a uxiliary DNA elements, and the trans-acting viral large tumor antigen (T antigen). Appropriate interactions between them initiate the assemb ly of a replication complex which, together with cellular proteins, is responsible for primer synthesis and DNA chain elongation. The organi zation of cis-acting elements within the origins of the polyomaviruses which replicate in mammalian cells is conserved; however, these origi ns are sufficiently distinct that the T antigen of one virus may funct ion inefficiently or not at ail to initiate replication at the origin of another virus. We have studied the basis for such replication selec tivity between the murine polyomavirus T antigen and the primate lymph otropic polyomavirus origin. The murine polyomavirus T antigen is capa ble of carrying out the early steps of the assembly of an initiation c omplex at the lymphotropic papovavirus origin, including binding to an d deformation of origin sequences in vitro. However, the T antigen ine fficiently unwinds the origin, and unwinding is influenced by sequence s flanking the T antigen pentanucleotide binding sites on the late sid e of the viral core origin. These same sequences contribute to the rep lication selectivity observed in vivo and in vitro, suggesting that th e inefficient unwinding is the cause of the replication defect. These observations suggest a mechanism by which origins of DNA replication c an evolve replication selectivity and by which the function of diverse cellular origins might be temporally activated during the S phase of the eukaryotic cell cycle.