We continue our studies of the replication and partition mechanisms that account for the stable inheritance of unit-copy bacterial plasmids for which P1 plasmid prophage serves as exemplar. In the period of this report we have (1) provided more compelling evidence that had been previously obtained for a DNA looping mediated by the P1-specified initiator protein, RepA, (2) shown, by site-directed mutagenesis and deletion analysis, that RepA protein appears not to be the cause of the replication inhibition observed when the repA gene is overexpressed, but that mutations within repA appear able to relieve this inhibition. These mutations are being exploited in the construction of a versatile miniP1-based cloning and expression vector, (3) devised promising selection procedures for the isolation of host mutants affecting plasmid maintenance, (4) determined the contribution of the recently characterized lytic Pl replicon to plasmid replication as a function of the expression of phage immunity genes, (5) demonstrated that P1 is an addictive genetic element; once acquired, the plasmid becomes necessary for continued bacterial survival, (6) shown that one of the two P1- determined proteins that are essential for partitioning of the plasmid to daughter cells binds to the plasmid centromere with an integration host factor (IHF). IHF is the first host component of a plasmid partitioning system to have been identified.
