XerD-mediated FtsK-independent integration of TLCϕ into the Vibrio cholerae genome

Abstract

As in most bacteria, topological problems arising from the circularity of the two Vibrio cholerae chromosomes, chrI and chrII, are resolved by the addition of a crossover at a specific site of each chromosome, dif, by two tyrosine recombinases, XerC and XerD. The reaction is under the control of a cell division protein, FtsK, which activates the formation of a Holliday Junction (HJ) intermediate by XerD catalysis that is resolved into product by XerC catalysis. Many plasmids and phages exploit Xer recombination for dimer resolution and for integration, respectively. In all cases so far described, they rely on an alternative recombination pathway in which XerC catalyzes the formation of a HJ independently of FtsK. This is notably the case for CTXϕ, the cholera toxin phage. Here, we show that in contrast, integration of TLCϕ, a toxin-linked cryptic satellite phage that is almost always found integrated at the chrI dif site before CTXϕ, depends on the formation of a HJ by XerD catalysis, which is then resolved by XerC catalysis. The reaction nevertheless escapes the normal cellular control exerted by FtsK on XerD. In addition, we show that the same reaction promotes the excision of TLCϕ, along with any CTXϕ copy present between dif and its left attachment site, providing a plausible mechanism for how chrI CTXϕ copies can be eliminated, as occurred in the second wave of the current cholera pandemic. cholera | site-specific DNA recombination | lysogenic conversion | chromosome segregation | lateral gene transfer

FtsK translocation permits discrimination between an endogenous and an imported Xer/ dif recombination complex

“…In all of the reported cases, integration of IMEXs recreates a bona fide dif site, thereby not interfering with chromosome dimer resolution, which would lead to their counter-selection. The best-described examples are Vibrio cholerae IMEXs, which carry crucial virulence determinants (5)(6)(7)17). These IMEXs have developed different strategies to integrate and to remain stably integrated, although the mechanisms ensuring their stable maintenance are not fully understood.…”

Effect of LexA on Chromosomal Integration of CTXϕ in Vibrio cholerae

“…4). Like CTX, both the phages use the same host-encoded XerC and XerD recombinases, but their integration in the chromosome of the cholera pathogen relies on the double-stranded attP substrates (25,26). We observed that integration of VGJ and TLC is not increased in lexA (Ind Ϫ ) V. cholerae cells (Table 5).…”

How Xer-exploiting mobile elements overcome cellular control

“…The attachment site of a V. cholerae IMEX, the toxin-linked cryptic phage (TLCϕ) harbors four of these bases (Fig. 1D, blue bases of dif GGI3 and attP TLC ) (25). We previously demonstrated that XerD poorly bound to attP TLC , which is sufficient to prevent XerD-mediated FtsK-dependent recombination (25).…”

“…1D, blue bases of dif GGI3 and attP TLC ) (25). We previously demonstrated that XerD poorly bound to attP TLC , which is sufficient to prevent XerD-mediated FtsK-dependent recombination (25). Thus, it is tempting to propose that GGI are IMEX and dif GGI sites were selected not only to escape but also to overcome the normal cellular control imposed on Xer recombination by FtsK.…”

“…Thus, it is tempting to propose that GGI are IMEX and dif GGI sites were selected not only to escape but also to overcome the normal cellular control imposed on Xer recombination by FtsK. GGI harboring dif GGI3 probably belong to the TLCϕ class of IMEX, which integrate into and excise from the genome of their host via a XerD-first FtsK-independent recombination pathway (25). GGI harboring dif GGI1 and dif GGI2 probably define a new class of IMEX.…”