The effect of the location of the proteic post-segregational stability system within the replicon of plasmid pTF-FC2 on the fine regulation of plasmid replication.

The broad host-range IncQ-2 family plasmid, pTF-FC2, is a mobilizable, medium copy number plasmid that lacks an active partitioning system. Plasmid stability is enhanced by a toxin-antitoxin (TA) system known as pas (plasmid addiction system) that is located within the replicon between the repB (primase) and the repA (helicase) and repC (DNA-binding) genes. The discovery of a closely related IncQ-2 plasmid, pRAS3, with a completely different TA system located between the repB and repAC genes raised the question of whether the location of pas within the replicon had an effect on the plasmid in addition to its ability to act as a TA system. In this work we demonstrate that the presence of the strongly expressed, autoregulated pas operon within the replicon resulted in an increase in the expression of the downstream repAC genes when autoregulation was relieved. While deletion of the pas module did not affect the average plasmid copy number, a pas-containing plasmid exhibited increased stability compared with a pas deletion plasmid even when the TA system was neutralized. It is proposed that the location of a strongly expressed, autoregulated operon within the replicon results in a rapid, but transient, expression of the repAC genes that enables the plasmid to rapidly restore its normal copy number should it fall below a threshold.