Abstract
Toxin-antitoxin (TA) loci were initially identified on conjugative plasmids, and one function of plasmid-encoded TA systems is to stabilize plasmids or increase plasmid competition via postsegregational killing. Here, we discovered that the type II TA system, Pseudoalteromonas rubra plasmid toxin-antitoxin PrpT/PrpA, on a low-copy-number conjugative plasmid, directly controls plasmid replication. Toxin PrpT resembles ParE of plasmid RK2 while antitoxin PrpA (PF03693) shares no similarity with previously characterized antitoxins. Surprisingly, deleting this prpA-prpT operon from the plasmid does not result in plasmid segregational loss, but greatly increases plasmid copy number. Mechanistically, the antitoxin PrpA functions as a negative regulator of plasmid replication, by binding to the iterons in the plasmid origin that inhibits the binding of the replication initiator to the iterons. We also demonstrated that PrpA is produced at a higher level than PrpT to prevent the plasmid from overreplicating, while partial or complete degradation of labile PrpA derepresses plasmid replication. Importantly, the PrpT/PrpA TA system is conserved and is widespread on many conjugative plasmids. Altogether, we discovered a function of a plasmid-encoded TA system that provides new insights into the physiological significance of TA systems.
Highlights
| | | | toxin–antitoxin plasmid replication plasmid copy number ParE origin of replication hypothesis proposes that TA systems have been selected on plasmids through horizontal plasmid propagation rather than due to vertical propagation [11]
Since conjugative plasmids are usually large and may carry genes encoding functions that are detrimental to the bacterial host, minimizing plasmid copy number is critical for reducing the host burden
By further searching the available sequenced plasmids, we found that ParE/PF03693 pairs are often found in conjugative plasmids of pathogenic and environmental bacteria, including Salmonella enterica, Enterobacter kobei, and P. rubra (Fig. 1D and SI Appendix, Table S1)
Summary
| | | | toxin–antitoxin plasmid replication plasmid copy number ParE origin of replication hypothesis proposes that TA systems have been selected on plasmids through horizontal plasmid propagation rather than due to vertical propagation [11]. The competition hypothesis is strongly supported by the fact that a plasmid encoding the ParE/ ParD TA system excluded an isogenic plasmid devoid of this TA module under conditions of horizontal gene transfer [11], and competition between plasmids led to a higher accumulation of TA systems on plasmids relative to chromosomes when transposonencoded TA systems were added [13]. Conjugative plasmids are extrachromosomal genetic elements that carry genetic determinants for adaptive traits enabling host bacteria to colonize diverse environments They are maintained in bacterial communities through both vertical inheritance and horizontal transfer [1, 2].
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