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Disruption of mouse Slx4, a regulator of structure-specific nucleases, phenocopies Fanconi anemia

Abstract

The evolutionarily conserved SLX4 protein, a key regulator of nucleases, is critical for DNA damage response. SLX4 nuclease complexes mediate repair during replication and can also resolve Holliday junctions formed during homologous recombination. Here we describe the phenotype of the Btbd12 knockout mouse, the mouse ortholog of SLX4, which recapitulates many key features of the human genetic illness Fanconi anemia. Btbd12-deficient animals are born at sub-Mendelian ratios, have greatly reduced fertility, are developmentally compromised and are prone to blood cytopenias. Btbd12−/− cells prematurely senesce, spontaneously accumulate damaged chromosomes and are particularly sensitive to DNA crosslinking agents. Genetic complementation reveals a crucial requirement for Btbd12 (also known as Slx4) to interact with the structure-specific endonuclease Xpf-Ercc1 to promote crosslink repair. The Btbd12 knockout mouse therefore establishes a disease model for Fanconi anemia and genetically links a regulator of nuclease incision complexes to the Fanconi anemia DNA crosslink repair pathway.

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Figure 1: Btbd12 deficiency results in growth retardation and compromised fertility.
Figure 2: Some Btbd12−/− mice died prematurely and displayed brain and eye developmental defects.
Figure 3: A proportion of Btbd12−/− mice developed blood cytopenias associated with genomic instability.
Figure 4: Btbd12-deficient cells undergo premature replicative senescence, exhibiting spontaneous and inducible chromosomal instability.
Figure 5: Btbd12-deficient MEFs are hypersensitive to DNA interstrand crosslinking agents.
Figure 6: The interaction between Slx4 and Xpf-Ercc1 is required for crosslink repair.

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Acknowledgements

We wish to thank L. Neidernhofer for the kind gift of Ercc1-deficient and wild-type congenic MEFs. Fanca- and Fancc-deficient MEFs were obtained from the Fanconi Anaemia Research Fund Cell Repository, Eugene, Oregon, USA. We thank A. Shortland for substantial contributions in the planning of animal experiments. The authors would like to thank M. Taylor for cloning of the mouse Slx4 cDNA. We would also like to thank T. Langford, J. Cruickshank, A. Mead, M. Brown, J. Wiles, K. Robinson, C. Shepherd, M. Reed, V. Smith and R. Berks for assistance in animal husbandry and handling. We would also like to thank G. Nemeth (Mediso Ltd.) for assistance with the computed tomography methodology. D.J.A. was supported by Cancer Research-UK and the Wellcome Trust. P.H.L.G. is supported by ATIP-Centre National de la Recherche Scientifique, Association pour la Recherche sur le Cancer and Fondation pour la Recherche Medicale grants.

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K.J.P. and G.P.C. designed the study, the experiments and wrote the paper. G.P.C. performed the majority of the experiments presented. D.J.A. helped in the design of the experiments. M.J.A. analyzed histological samples and provided useful discussion. L.v.d.W. managed the colony and weighed and performed necropsies on mice at Wellcome Trust Sanger Institute. I.V.R. analyzed Fanconi pathway activation. F.L. assisted in analysis of developmental abnormalities. P.-H.L.G. helped in the design of some experiments. R.E.M. performed the micronucleus assay. S.M.G.P. performed the mouse phenotype pipeline analysis. F.G., M.I.K., D.Y.L. and K.B. performed imaging studies.

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Correspondence to Ketan J Patel.

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Crossan, G., van der Weyden, L., Rosado, I. et al. Disruption of mouse Slx4, a regulator of structure-specific nucleases, phenocopies Fanconi anemia. Nat Genet 43, 147–152 (2011). https://doi.org/10.1038/ng.752

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