DNA transposition and site specific recombination

Part of the research in the laboratory is on the molecular mechanisms that mediate specialised DNA rearrangements in bacteria through transposition or site-specific recombination.
Transposition is the process used by a variety of mobile genetic elements (transposons, retrotransposons, viruses and retroviruses) to move from one site to another within or between genomes, whereas site-specific recombination is a usually efficient and tightly controlled DNA breakage and joining reaction occurring at the level of determined DNA sequences. This reaction is used to affect different biological functions related to bacteriophages life cycle, chromosomes and plasmids segregation, lateral transfers and virulence development.

The replicative transposition cycle of Tn4430 transposon

Tn4430 is a Tn3-family transposon from Bacillus thuringiensis. Transposition of this element is a multi-step process involving two transposon-encoded proteins:

  • The transposase TnpA
  • The tyrosine recombinase TnpI

At the initial steps of transposition, the TnpA protein co-operate with the host replication machinery to generate a DNA structure termed "cointegrate", in which the donor and target DNA molecules are joined by head-to-tail copies of the transposon. TnpA binding to the transposon ends also confers transposition immunity, a phenomenon that prevents insertion of multiple copies of the transposon in the same locus. The molecular basis of this "molecular repulsion" mechanism remains largely unexplored.

The TnpI recombinase is required to resolve the cointegrate transposition intermediate by catalysing a DNA site-specific recombination reaction between the duplicated copies of the transposon. Biochemical studies have shown that this reaction is controlled by a unique mechanism in which the TnpI protein acts both as a catalytic and regulatory component of the recombination complex.


Complementary genetical and biochemical approaches are used to investigate the molecular interactions that are involved in the assembly of the transposition and site-specific recombination complexes and the control of the DNA breakage and rejoining reactions catalysed by the TnpA and TnpI proteins.

Mutations produced in the proteins and their DNA substrates are studied in vivo, as well as in vitro using biochemical assays reproducing specific steps of the transposition/recombination reactions.
The aim is to provide a detailed picture describing how the transposition/recombination machines assemble and function and how they are controlled at the molecular level within the cell.

These machines provide interesting model systems for the understanding of DNA transactions occurring between separate regions of the genome. Similar mechanisms are involved in other important biological processes, such as DNA replication and repair, or the regulation of gene expression. Likewise transposition immunity conferred by the TnpA protein may represent a more general mechanism for "negative" regulation in bacterial systems.

From transposition and site-specific recombination mechanisms to the molecular biology toolbox

Also central to our project is the development of new technologies based on Tn4430 transposition and recombination mechanisms.

  • Pentapeptide scanning mutagenesis is a Tn4430-based method allowing to insert a variable 5-amino acid cassette at random positions of a protein. This method has already been used for the study several proteins.
  • The tyrosine recombinase TnpI was found to function on a variety of DNA substrates in vitro, as well as in different cell types in vivo, including mammal cells. This enzyme thus represents a valuable candidate for the development of new tools in molecular biology and genetic engineering.

Future work aims at implementing the TnpA- and TnpI-based technologies to further enlarge their field of application, including in functional geneomics studies.