Drosophila MUS312 and the Vertebrate Ortholog BTBD12 Interact with DNA Structure-Specific Endonucleases in DNA Repair and Recombination

Abstract

Summary DNA recombination and repair pathways require structure-specific endonucleases to process DNA structures that include forks, flaps, and Holliday junctions. Previously, we determined that the Drosophila MEI-9-ERCC1 endonuclease interacts with the novel MUS312 protein to produce meiotic crossovers, and that MUS312 has a MEI-9-independent role in interstrand crosslink (ICL) repair. The importance of MUS312 to pathways crucial for maintaining genomic stability in Drosophila prompted us to search for orthologs in other organisms. Based on sequence, expression pattern, conserved protein-protein interactions, and ICL repair function, we determined that the mammalian ortholog of MUS312 is BTBD12. Orthology between these proteins and S. cerevisiae Slx4 helped identify a conserved interaction with a second structure-specific endonuclease, SLX1. Genetic and biochemical evidence described here and in related papers suggest that MUS312 and BTBD12 direct Holliday junction resolution by at least two distinct endonucleases in different recombination and repair contexts.

Recruitment and positioning determine the specific role of the XPF ‐ ERCC 1 endonuclease in interstrand crosslink repair

“…We then examined the XE ΔNSGW mutant and found that it was recruited normally to the ICL (Fig 5D and Appendix Fig S2C). This is striking, because this region was shown to be important for the interaction between XPF and SLX4 (Andersen et al , 2009) and we have previously shown that SLX4 is important for the recruitment of XPF to the ICL. Lastly, we examined the recruitment of the nuclease domain mutants XE R670S and XE S767F .…”

“…These mutated residues correspond to residues L219 and C225 in Xenopus laevis XPF that is 75% identical to human XPF (Fig 1A). Another mutation in XPF's helicase‐like domain, hs G325E ( xl G314E), was reported to disrupt the interaction of XPF with the BTB domain of SLX4 (Andersen et al , 2009). This interaction is likely specific to ICL repair and not NER because SLX4‐deficient cells are not UV sensitive (Crossan et al , 2011).…”

“…The hs G325E mutation in XPF abrogates the interaction between XPF and the BTB domain of SLX4 (Andersen et al , 2009). We found that the XE G314E and XE ΔNSGW mutants were both able to interact normally with full‐length SLX4 (Figs 6B and EV5D).…”

“…A specific residue in the helicase‐like domain of XPF has been implicated in the interaction with SLX4 (Yildiz et al , 2002; Andersen et al , 2009). A glycine to glutamic acid mutation at residue 325 of human XPF abrogated the interaction with SLX4 in a yeast two‐hybrid assay (Andersen et al , 2009). This yeast two‐hybrid assay was performed with C‐terminal deletion mutants of SLX4 and the interaction with XPF was pinpointed to the BTB domain.…”

A new class of ultrafine anaphase bridges generated by homologous recombination

“…single HJs and D-loop structures), are processed by a second mechanism which involves structure-selective endonuclease (SSE)-mediated resolution [31,32,39,40]. There are two genetically distinct resolution pathways: one is mediated by the SLX1–SLX4, MUS81-EME1 and XPF-ERCC1 (SMX) tri-nuclease complex [27,31,41–46], and the other is mediated by GEN1 endonuclease [47–50]. …”