The coupling between the replication and parD (kis, kid) maintenance modules of R1 has been revisited here by the isolation of a significant collection of conditional replication mutants in the pKN1562 mini-R1 plasmid, and in its derivative, pJLV01, specifically affected in the RNase activity of the Kid toxin. This new analysis aims to identify key factors in this coupling. For this purpose we have quantified and characterized the restriction introduced by parD to isolate conditional replication mutants of this plasmid, a signature of the modular coupling. This restriction depends on the RNase activity of the Kid toxin and it is relieved by either over-expression of the Kis antitoxin or by preventing its degradation by Lon and ClpAP proteases. Based on these data and on the correlation between copy numbers and parD transcriptional levels obtained in the different mutants, it is proposed that a reduction of Kis antitoxin levels in response to inefficient plasmid replication is the key factor for coupling plasmid replication and parD modules.
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