Sequential induction of three recombination directionality factors directs assembly of tripartite integrative and conjugative elements

Abstract

Tripartite integrative and conjugative elements (ICE3) are a novel form of ICE that exist as three separate DNA regions integrated within the genomes of Mesorhizobium spp. Prior to conjugative transfer the three ICE3 regions of M. ciceri WSM1271 ICEMcSym1271 combine and excise to form a single circular element. This assembly requires three coordinated recombination events involving three site-specific recombinases IntS, IntG and IntM. Here, we demonstrate that three excisionases–or recombination directionality factors—RdfS, RdfG and RdfM are required for ICE3 excision. Transcriptome sequencing revealed that expression of ICE3 transfer and conjugation genes was induced by quorum sensing. Quorum sensing activated expression of rdfS, and in turn RdfS stimulated transcription of both rdfG and rdfM. Therefore, RdfS acts as a “master controller” of ICE3 assembly and excision. The dependence of all three excisive reactions on RdfS ensures that ICE3 excision occurs via a stepwise sequence of recombination events that avoids splitting the chromosome into a non-viable configuration. These discoveries expose a surprisingly simple control system guiding molecular assembly of these novel and complex mobile genetic elements and highlight the diverse and critical functions of excisionase proteins in control of horizontal gene transfer.

Horizontally Acquired Homologs of Xenogeneic Silencers: Modulators of Gene Expression Encoded by Plasmids, Phages and Genomic Islands

“…Under certain conditions, GIs can be excised from the chromosome through the site-specific recombination between the direct repeated sequences (DRSs) that flank the element, in a reaction catalyzed by the integrase protein encoded within or outside the island [80][81][82][83]. This reaction, which is usually promoted by a recombination directionality factor also encoded inside or outside the GI, results in the formation of a circular element [84,85]. The insertion site is reconstituted in the chromosome, and a copy remains as part of the excised GI.…”

Regulation Mediated by N-Acyl Homoserine Lactone Quorum Sensing Signals in the Rhizobium-Legume Symbiosis

“…Here, we will focus on the role of QS on relevant traits such as EPS production and motility and we will discuss the relevance of this regulatory system in the establishment of symbiosis. For information about QS-controlled transfer of plasmids and symbiosis islands, readers are referred to more detailed studies [ 69 , 77 , 83 , 87 , 92 , 93 , 94 ].…”

Delineation of the integrase-attachment and origin-of-transfer regions of the symbiosis island ICEMlSymR7A

“…We have previously demonstrated that a synthetic "mini-ICE" carrying only intS and attP integrates into the phe-tRNA gene in R7ANS; a derivative of R7A from which ICEMlSym R7A was cured (20). The ICEMlSym R7A RDF, RdfS, is a putative member of the MerR superfamily of winged helixturn-helix (wHTH) DNA-binding proteins (12,33,34 (20). Escherichia coli DC10B (41) cultures were grown in Lysogeny broth (LB) at 37⁰C.…”

“…1). rdfS is the first gene in a polycistronic operon encoding traF, msi107 and probably rlxS (33,58). The traF start site also overlaps with the rdfS stop codon suggesting they may be translationally coupled.…”