6568–6584 Nucleic Acids Research, 2011, Vol. 39, No. 15 doi:10.1093/nar/gkr340 Published online 16 May 2011 Ultrafine anaphase bridges, broken DNA and illegitimate recombination induced by a replication fork barrier Sevil Sofueva, Fekret Osman, Alexander Lorenz, Roland Steinacher, Stefania Castagnetti, Jennifer Ledesma and Matthew C. Whitby* Department of Biochemistry, University of Oxford, South Parks Road, Oxford, OX1 3QU, UK Received November 25, 2010; Revised April 13, 2011; Accepted April 21, 2011 ABSTRACT Most DNA double-strand breaks (DSBs) in S- and G2-phase cells are repaired accurately by Rad51- dependent sister chromatid recombination. However, a minority give rise to gross chromosome rearrangements (GCRs), which can result in disease/ death. What determines whether a DSB is repaired accurately or inaccurately is currently unclear. We provide evidence that suggests that perturbing replication by a non-programmed protein–DNA replication fork barrier results in the persistence of replication intermediates (most likely regions of unreplicated DNA) into mitosis, which results in anaphase bridge formation and ultimately to DNA breakage. However, unlike previously chara- cterised replication-associated DSBs, these breaks are repaired mainly by Rad51-independent pro- cesses such as single-strand annealing, and are therefore prone to generate GCRs. These data high- light how a replication-associated DSB can be predisposed to give rise to genome rearrangements in eukaryotes. INTRODUCTION The path to complete genome duplication is littered with obstacles that can impede the progression of the replica- tion fork (1). These replication fork barriers (RFBs) include natural programmed RFBs, which are designed to ensure unidirectional replication of a given locus, as well as numerous accidental RFBs, including various DNA lesions, secondary structures in the template DNA, and protein–DNA complexes (2). Cells appear to have a variety of mechanisms for coping with RFBs, and it is thought that the nature of the barrier determines which is used in any given situation (1). In many cases it appears that the response is to protect the fork and its protein components (the replisome), until such time as the block has been removed and DNA syn- thesis can resume, or replication is completed by the arrival of the opposing fork (3). However, when fork pro- tection fails or the barrier proves persistent homologous recombination (HR) can be used to enable template switching to bypass the RFB or to promote replication restart if the fork has collapsed (= dissociation of the replisome from DNA) or broken (4,5). This recombin- ation typically occurs between equivalent DNA sequences on the sister chromatids, however occasionally HR acts between allelic or ectopic sites and consequently can result in changes to the genome such as loss of heterozy- gosity (LOH) and gross chromosome rearrangements (GCRs) including deletions, duplications, inversions and translocations that in humans can promote diseases such as cancer (5–8). Whether this misdirected recombination is solely stochastic or can be driven by the nature of the fork impediment is unknown. A key requirement in HR is the presence of a stretch of single-stranded (ss) DNA that is bound by recombinases, such as Rad51 and Rad52, which then promote either strand invasion of a homologous duplex DNA or anneal- ing to a complementary DNA strand (9). Replication forks naturally contain a region of ssDNA on the lagging template strand, and it is thought that this can become accessible to recombinases following fork stalling. Alternatively, the stalled fork can be remodelled through a process of fork reversal, which causes the nascent DNA strands to anneal to each other forming a duplex DNA end that can be resected to produce a *To whom correspondence should be addressed. Tel: 44 1865 613239; Fax: 44 1865 613343; Email: matthew.whitby@bioch.ox.ac.uk Present address: Sevil Sofueva, UCL Cancer Institute, 72 Huntley Street, London, WC1E 6BT, UK. The authors wish it to be known that, in their opinion, the ﬁrst three authors should be regarded as joint First Authors. The Author(s) 2011. Published by Oxford University Press. This is an Open Access article distributed under the terms of the Creative Commons Attribution Non-Commercial License (http://creativecommons.org/licenses/ by-nc/3.0), which permits unrestricted non-commercial use, distribution, and reproduction in any medium, provided the original work is properly cited.