Mouse SLX4 is a tumor suppressor that stimulates the activity of the nuclease XPF-ERCC1 in DNA crosslink repair.
Bottom Line: Slx4-deficient mice develop epithelial cancers and have a contracted hematopoietic stem cell pool.The N-terminal domain of SLX4 (mini-SLX4) that only binds to XPF-ERCC1 is sufficient to confer resistance to DNA crosslinking agents.Recombinant mini-SLX4 enhances XPF-ERCC1 nuclease activity up to 100-fold, directing specificity toward DNA forks.
Affiliation: MRC Laboratory of Molecular Biology, Francis Crick Avenue, Cambridge, CB2 0QH, UK.Show MeSH
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Mentions: As already mentioned, transformed MEF cells lines obtained from Slx4f3/f3 embryos were hypersensitive to ICL agents, such as Mitomycin C (MMC). The introduction of a full-length Slx4 transgene into these cells can complement this key phenotypic feature. This simple, cell-intrinsic DNA repair defect provides a system for functional dissection of the SLX4 polypeptide. SLX4 is a large 1565 amino acid polypeptide that serves as a binding platform for three nucleases (Figure 2A). An N-terminal MLR domain mediates the interaction with XPF-ERCC1, whereas MUS81-EME1 and SLX1 bind through regions mapping near the C terminus of SLX4 (Fekairi et al., 2009; Kim et al., 2013; Svendsen et al., 2009). Additionally, SLX4 possess two N-terminal UBZ domains and a central BTB/POZ protein dimerization/interaction domain. We created a truncation of mouse SLX4 (SLX4 1-758: mini-SLX4) that includes the XPF-ERCC1 binding region (MLR) and ectopically expressed this in Slx4f3/f3-deficient MEFs. Mini-SLX4 binds to endogenous XPF-ERCC1 as efficiently as the full-length SLX4 polypeptide (Figure 2B). This mini-SLX4 also significantly complements resistance to MMC (Figure 2C) (LD50 values: Slx4f3/f3 4 ng/ml, Mini-SLX4 23 ng/ml, and full-length SLX4 80 ng/ml).
Affiliation: MRC Laboratory of Molecular Biology, Francis Crick Avenue, Cambridge, CB2 0QH, UK.