Yeast cells that are deleted for both the Mus81-Eme1 endonuclease and Rqh1 DNA helicase are unviable. Our work established that the unviability of a Mus81 Rqh1 double mutant is due to a failure to process four-way DNA junctions that are formed during the repair of stalled and broken replication forks. Replication fork repair is a fundamental process in biology and mutations in the mammalian homologues of Mus81-Eme1 and Rqh1 result in genome instability and cancer predisposition.

Our work made use of a bacterial enzyme called RusA that specifically cleaves a DNA structure known as a Holliday junction (HJ). HJs are key intermediates of genetic recombination, and can also form at blocked replication forks. We had previously made a recombinant form of RusA that can function in eukaryotic cells. This has proved a useful and novel tool for identifying proteins that process HJs and HJ precursors in eukaryotic cells.

The faithful transmission of genetic information during cell division depends on exact replication and segregation of the chromosomes. The replication machinery frequently encounters lesions and proteins in the DNA template, which stall and block its progress. In order to recover DNA synthesis, cells can use their recombination machinery to overcome the blockage and restart replication. A key intermediate during this process is a four-way DNA junction. Our work helped to identify the roles of two key proteins involved in processing these junctions. In mammalian cells, loss of these proteins results in genetic instability and predisposition to cancer.

I became interested in enzymes that process HJs when working as a post-doc in Bob Lloyd’s laboratory at the University of Nottingham. We were involved in the identification and characterization of enzymes that branch migrate and resolve HJs in the bacterium Escherichia coli. When I left Nottingham to set up my own lab in Oxford, I chose to continue this area of research but I switched my focus to HJ processing in the eukaryotic model system Schizosaccharomyces pombe (fission yeast).