Participation of the lytic replicon in bacteriophage P1 plasmid maintenance

Abstract

P1 bacteriophage carries at least two replicons: a plasmid replicon and a viral lytic replicon. Since the isolated plasmid replicon can maintain itself stably at the low copy number characteristic of intact P1 prophage, it has been assumed that this replicon is responsible for driving prophage replication. We provide evidence that when replication from the plasmid replicon is prevented, prophage replication continues, albeit at a reduced rate. The residual plasmid replication is due to incomplete repression of the lytic replicon by the ci immunity repressor. Incomplete repression was particularly evident in lysogens of the thermoinducible P1 cl.100 prophage, whose replication at 32°C remained almost unaffected when use of the plasmid replicon was prevented. Moreover, the average plasmid copy number of P1 in a P1 cl.100 lysogen was elevated with respect to the copy number of P1 c1+. The capacity of the lytic replicon to act as an auxiliary in plasmid maintenance may contribute to the extraordinary stability of P1 plasmid prophage.P1 prophage replicates in Escherichia coli as a stable, low-copy-number, 90-kilobase (kb) plasmid. (See reference 47 for all but selected references to primary sources concerning P1.) A 4-kb segment of P1 DNA, the plasmid replicon, suffices to support stable plasmid maintenance at the low copy number. This segment consists of a 1.5-kb module, responsible for controlled replication from an origin known as oriR, and an adjacent 2.5-kb module, responsible for partitioning of the daughter plasmids at cell division.Replication from oriR requires the oriR-specific initiator, the product of the adjacent repA gene. A regulatory element, incA, is situated immediately downstream of r-epA and consists of a set of sites that, like oriR itself, bind initiator and interfere with oriR function. Also required for replication from oriR is the bacterial dnaA gene product, for which presumptive binding sites in oriR are present. P1 lytic replication initiates at an origin known as oriL (14,22,38). The lytic replicon appears not to require dnaA function (23). It does depend on the integrity of an open reading frame (repL) within which oriL is probably embedded (22,38). Control of the lytic replicon is exerted by the ci immunity repressor acting at an operator (designated Op53) to repress an operon that includes repL and an additional gene, not essential for replication, upstream of it (22,38). Repression by ci is antagonized by ant, normally controlled in P1 lysogens by the secondary immunity gene c4. The products of ci and c4 do not inhibit replication from the plasmid replicon (5), nor does the control element of the plasmid replicon (incA) inhibit the lytic replicon (unpublished results).We have examined the growth of P1 lysogens when either or both replicons are inhibited. Replication from oriR was inhibited in one of three ways: (i) mutational inactivation of

Genome analysis of the Clostridium difficile phage ΦCD6356, a temperate phage of the Siphoviridae family

“…However, in most Siphoviridae phages, the ori site is located close to or within the coding region of the replication protein and direct repeats were also evident within the ΦCD6356 putative replication protein. This may suggest that ΦCD6356 uses separate origins of replications for its lytic and lysogenic states, as is the case for the plasmid prophage, P1 (Yarmolinsky et al, 1989).…”

Characterization of the primary immunity region of the Escherichia coli linear plasmid prophage N15

“…Modulation of replication gene expression by antisense RNA is a strategy of several other plasmids (6,13,36,41) and of bacteriophage P1 (10). In the case of P1, the antisense RNA modulates expression of a C1-controlled replication gene that is normally specific for lytic development but which is also capable of sustaining plasmid replication when partially derepressed (46).…”

Isolation of high affinity human antibodies directly from large synthetic repertoires.

“…Phage P1 lysates were made by thermal induction (Rosner, 1972). E.coli C600 Su- (Appleyard, 1954) harbouring phage PlCm cl.100 r-m (Yarmolinsky et al, 1989) were grown in a 2 1 baffled flask containing 11 of 2XTY, 25 jig/ml chloramphenicol, 10 mM MgSO4 with vigorous shaking at 30°C to OD600 of 0.6. The temperature was then raised quickly to 42°C by shaking in a 70°C water bath and then shaking continued for a further 35 min in a 40°C water bath.…”