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Aberrant chromosome morphology in human cells defective for Holliday junction resolution


In somatic cells, Holliday junctions can be formed between sister chromatids during the recombinational repair of DNA breaks or after replication fork demise. A variety of processes act upon Holliday junctions to remove them from DNA, in events that are critical for proper chromosome segregation. In human cells, the BLM protein, inactivated in individuals with Bloom’s syndrome, acts in combination with topoisomerase IIIα, RMI1 and RMI2 (BTR complex) to promote the dissolution of double Holliday junctions1,2. Cells defective for BLM exhibit elevated levels of sister chromatid exchanges (SCEs) and patients with Bloom’s syndrome develop a broad spectrum of early-onset cancers caused by chromosome instability3. MUS81–EME1 (refs 4–7), SLX1–SLX4 (refs 8–11) and GEN1 (refs 12, 13) also process Holliday junctions but, in contrast to the BTR complex, do so by endonucleolytic cleavage. Here we deplete these nucleases from Bloom’s syndrome cells to analyse human cells compromised for the known Holliday junction dissolution/resolution pathways. We show that depletion of MUS81 and GEN1, or SLX4 and GEN1, from Bloom’s syndrome cells results in severe chromosome abnormalities, such that sister chromatids remain interlinked in a side-by-side arrangement and the chromosomes are elongated and segmented. Our results indicate that normally replicating human cells require Holliday junction processing activities to prevent sister chromatid entanglements and thereby ensure accurate chromosome condensation. This phenotype was not apparent when both MUS81 and SLX4 were depleted from Bloom’s syndrome cells, suggesting that GEN1 can compensate for their absence. Additionally, we show that depletion of MUS81 or SLX4 reduces the high frequency of SCEs in Bloom’s syndrome cells, indicating that MUS81 and SLX4 promote SCE formation, in events that may ultimately drive the chromosome instabilities that underpin early-onset cancers associated with Bloom’s syndrome.

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Figure 1: Contribution of GEN1, MUS81 and SLX4 to SCE frequency in Bloom’s syndrome cells.
Figure 2: Chromosome abnormalities in Bloom’s syndrome (BS) cells after siRNA-mediated depletion of GEN1 and MUS81.
Figure 3: Quantification of the chromosome segmentation phenotype observed in Bloom’s syndrome cells after GEN1 and MUS81 depletion.
Figure 4: Synthetic interactions between GEN1, MUS81 and SLX4.


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We thank I. Hickson for providing the Bloom’s syndrome cell lines and advice, P. Edwards for help and providing facilities for chromosome painting, S. Horswell for the statistical analysis, S. Ip for the GEN1 antibody, M.G. Blanco for assistance with SCE scoring and our laboratory colleagues for their encouragement and suggestions. We further thank K. Cimprich, the Cimprich laboratory members, and C. Wang, W. Johnson and A. Straight. This work was supported by Cancer Research UK, the Louis-Jeantet Foundation, the European Research Council, the Swiss Bridge Foundation and the Breast Cancer Campaign. S.N. was supported by a studentship from the UK Medical Research Council.

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T.W. and S.C.W. designed the project that was undertaken entirely by T.W. Expertise for the chromosome paints was provided by S.N. The manuscript was written by S.C.W. with help from T.W.

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Correspondence to Stephen C. West.

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The authors declare no competing financial interests.

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Wechsler, T., Newman, S. & West, S. Aberrant chromosome morphology in human cells defective for Holliday junction resolution. Nature 471, 642–646 (2011).

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