Crosstalk between transposase subunits during cleavage of the mariner transposon.

Corentin Claeys Bouuaert,Ronald Chalmers,Danxu Liu,Neil Walker

doi:10.1093/nar/gku172

Corentin Claeys Bouuaert, Ronald Chalmers + Show 2 more

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https://doi.org/10.1093/nar/gku172

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Abstract

Mariner transposition is a complex reaction that involves three recombination sites and six strand breaking and joining reactions. This requires precise spatial and temporal coordination between the different components to ensure a productive outcome and minimize genomic instability. We have investigated how the cleavage events are orchestrated within the mariner transpososome. We find that cleavage of the non-transferred strand is completed at both transposon ends before the transferred strand is cleaved at either end. By introducing transposon-end mutations that interfere with cleavage, but leave transpososome assembly unaffected, we demonstrate that a structural transition preceding transferred strand cleavage is coordinated between the two halves of the transpososome. Since mariner lacks the DNA hairpin intermediate, this transition probably reflects a reorganization of the transpososome to allow the access of different monomers onto the second pair of strands, or the relocation of the DNA within the same active site between two successive hydrolysis events. Communication between transposase subunits also provides a failsafe mechanism that restricts the generation of potentially deleterious double-strand breaks at isolated sites. Finally, we identify transposase mutants that reveal that the conserved WVPHEL motif provides a structural determinant of the coordination mechanism.

Highlights

Transposons are genetic elements that mobilize and amplify within a host genome
For strand cleavage analyses of reactions using a supercoiled substrate, the products of transposition reactions were digested with the BsaHI restriction endonuclease and 3 -labeled with ␣32P-dCTP and the Klenow enzyme or dephosphorylated with Antarctic phosphatase and 5 -labeled with ␥ -32P-ATP and the polynucleotide kinase (PNK) enzyme
Several important features of transposon end cleavage have already been established for Mos1 and the correct interpretation of the current work depends on knowing whether Hsmar1 uses the same mechanism

Summary

Introduction

Transposons are genetic elements that mobilize and amplify within a host genome. they are generally detrimental, and often considered as molecular parasites, they are important evolutionary forces [1]. To excise the transposon from the donor site, the transposase generates a double-strand break (DSB) at both ends of the element. This is followed by integration of the transposon in a new target site (Figure 1A). Since the transposition reaction represents a danger to the host and the transposon itself, regulation is important to avoid deleterious events. These include the formation of DSBs at an isolated site, the failure of the transposon to reintegrate or other more complex genome rearrangements such as those that arise from single-end strand transfer These include the formation of DSBs at an isolated site, the failure of the transposon to reintegrate or other more complex genome rearrangements such as those that arise from single-end strand transfer (e.g. [2])

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