Transposition into Replicating DNA Occurs through Interaction with the Processivity Factor

Abstract

SUMMARY The bacterial transposon Tn7 directs transposition into actively replicating DNA by a mechanism involving the transposon-encoded protein TnsE. Here we show that TnsE physically and functionally interacts with the processivity factor of the DNA replication machinery in vivo and in vitro. Our work establishes an in vitro TnsABC+E transposition reaction reconstituted from purified proteins and target DNA structures. Using the in vitro reaction we confirm that the processivity factor specifically reorders TnsE-mediated transposition events on target DNAs in a way that matches the bias with active DNA replication in vivo. The TnsE interaction with an essential and conserved component of the replication machinery and a DNA structure reveals a new mechanism by which Tn7, and probably other elements, select target-sites associated with DNA replication.

Recruitment of CRISPR-Cas systems by Tn7-like transposons

“…However, the matches that could be detected were informative because they were to plasmids and bacteriophages associated with the same bacterial genera in which the respective elements are found (Table S2). We identified two cases (in Photobacterium kishitanii and Photobacterium leiognathi) of special interest, in which spacers matched the region adjacent to the tnsA-gene-proximal side of the element (Table S2), i.e., the specific region where complexes involved in targeting transposition events interact with the target DNA (44,56,58). An additional spacer match was found inside the transposon boundaries in several V. parahaemolyticus strains (Table S2).…”

“…Transposition events promoted by TnsABC+D are directed into a position 23 bp downstream of the region bound by TnsD. Tn7 transposition is orientation specific in all transposition pathways; the transposon end proximal to the tnsA gene (the "right" end of the element) is adjacent to the DNA sequence or a specific protein complex recognized in each pathway (44,(56)(57)(58).…”

“…Transposition into the attTn7 site shows no negative impact on the host, providing a "safe haven" for these elements that appear to be universally maintained in bacteria. Transposition mediated by TnsABC + TnsE is preferentially directed into plasmids and bacteriophages owing to the ability of TnsE to recognize complexes formed during specific types of DNA replication (42)(43)(44)(45). The TnsE-mediated transposition that preferentially directs insertion into other MGE is likely responsible for the wide distribution of Tn7 elements among bacteria.…”

Drosophila P elements preferentially transpose to replication origins

“…One of Tn7's distinct transposition pathways detects an altered DNA structure as well as factors associated with lagging-strand DNA synthesis, which leads to a strong orientational bias in insertion (20). Recently, the prokaryotic analog of Proliferating Cell Nuclear Antigen (PCNA), the β-clamp, was shown to be such a factor and to interact with Tn7 at lagging strands (31). Interactions with replication, including integration at origins, may be a feature shared by many transposons.…”

Unlocking Tn3-family transposase activity in vitro unveils an asymetric pathway for transposome assembly

“…In contrast, TnsC is addressed to specific sites by the Tn7-encoded target selector proteins TnsD or TnsE. TnsD promotes assembly of the target complex at a specific chromosomal site, attTn7, whereas TnsE directs transposition to DNA regions undergoing active lagging-strand DNA synthesis (30,46). Although Tn4430 inserts in a broad variety of DNA sequences in vivo, the data suggest that integration depends on specific host factors or cellular processes that locally affect DNA, directing transposition to specific DNA regions (6,18).…”