Characterization of XerC‐ and XerD‐dependent CTX phage integration in Vibrio cholerae

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Abstract
CTXφ is a filamentous bacteriophage that encodes cholera toxin and integrates site-specifically into the larger of the two Vibrio cholerae chromosomes. The CTXφ genome lacks an integrase; instead, its integration depends on the chromosome-encoded tyrosine recombinases XerC and XerD. During integration, recombination occurs between regions of homology in CTXφ and the V. cholerae chromosome. Here, we define the elements on the phage genome (attP ) and bacterial chromosome (attB) required for CTXφ integration. attB is a short sequence composed of one binding site for XerC and XerD spanning the site of recombination. Together, XerC and XerD bind to two sites within attP. While one XerC/D binding site in attP spans the core recombination region, the other site is ≈ 80 bp away. Although integration occurs at the core XerC/D binding site in attP, the second site is required for CTXφ integration, suggesting it performs an architectural role in the integration reaction. In vitro cleavage reactions showed that XerC and XerD are capable of cleaving attB and attP sequences; however, additional cellular processes such as DNA replication or Holliday junction resolution by a host resolvase may contribute to integration in vivo.