Identification of Two Replicons in Phage-Plasmid P4

Abstract

DNA replication of phage-plasmid P4 proceeds bidirectionally from theori1site (previously namedori), but requires a secondcis-acting region,crr.Replication depends on the product of the P4 α gene, a protein with primase and helicase activity, that binds bothori1andcrr.A negative regulator of P4 DNA replication, the Cnr protein, is required for copy number control of plasmid P4. Using a plasmid complementation test for replication, we found that two replicons, both dependent on the α gene product, coexist in P4. The first replicon is made by thecnrand α genes and theori1andcrrsites. The second is limited to the α andcrrregion. Thus, in the absence of theori1region, replication can initiate at a different site. By deletion mapping, acis-acting region,ori2,essential for replication of the α-crrreplicon was mapped within a 270-bp fragment in the first half of the α gene. Theori2site was found to be dispensable in a replicon that containsori1.A construct that besidescrrand α carries also thecnrgene was unable to replicate, suggesting that Cnr not only controls replication fromori1,but also silencesori2.