Replication and Active Partition of Integrative and Conjugative Elements (ICEs) of the SXT/R391 Family: The Line between ICEs and Conjugative Plasmids Is Getting Thinner

摘要

Integrative and Conjugative Elements (ICEs) of the SXT/R391 family disseminate multidrug resistance among pathogenic Gammaproteobacteria such as Vibrio cholerae . SXT/R391 ICEs are mobile genetic elements that reside in the chromosome of their host and eventually self-transfer to other bacteria by conjugation. Conjugative transfer of SXT/R391 ICEs involves a transient extrachromosomal circular plasmid-like form that is thought to be the substrate for single-stranded DNA translocation to the recipient cell through the mating pore. This plasmid-like form is thought to be non-replicative and is consequently expected to be highly unstable. We report here that the ICE R391 of Providencia rettgeri is impervious to loss upon cell division. We have investigated the genetic determinants contributing to R391 stability. First, we found that a hipAB -like toxin/antitoxin system improves R391 stability as its deletion resulted in a tenfold increase of R391 loss. Because hipAB is not a conserved feature of SXT/R391 ICEs, we sought for alternative and conserved stabilization mechanisms. We found that conjugation itself does not stabilize R391 as deletion of traG , which abolishes conjugative transfer, did not influence the frequency of loss. However, deletion of either the relaxase-encoding gene traI or the origin of transfer ( oriT ) led to a dramatic increase of R391 loss correlated with a copy number decrease of its plasmid-like form. This observation suggests that replication initiated at oriT by TraI is essential not only for conjugative transfer but also for stabilization of SXT/R391 ICEs. Finally, we uncovered srpMRC , a conserved locus coding for two proteins distantly related to the type II (actin-type ATPase) parMRC partitioning system of plasmid R1. R391 and plasmid stabilization assays demonstrate that srpMRC is active and contributes to reducing R391 loss. While partitioning systems usually stabilizes low-copy plasmids, srpMRC is the first to be reported that stabilizes a family of ICEs. Author Summary Integrative and conjugative elements (ICEs) constitute a class of mobile genetic elements defined by their ability to integrate into the chromosome of their host cell and to transfer by conjugation. Some of the most studied ICEs belong to the SXT/R391 family, which are major drivers of multidrug resistance dissemination among various pathogenic Gammaproteobacteria . Transfer of SXT/R391 ICEs to a new host first requires its excision from the chromosome as a circular molecule, which may be lost if the cell divides. In silico analyses revealed several putative stabilization systems carried by R391, a prototypical member of the SXT/R391 ICEs family originally isolated from Providencia rettgeri . We discovered that, besides stabilization by integration into the chromosome, stability of SXT/R391 ICEs also depends on toxin/antitoxin systems and plasmid-like features including intracellular replication and active partition. Thus, although it has been known for a long time that ICEs and conjugative plasmids use similar strategies to transfer between bacterial populations, our work reveals additional unforeseen similarities in their mechanisms of maintenance in the host cell.