806 Research Article Introduction Faithful chromosome segregation requires that sister chromatids are held together from S phase, the time of DNA replication, to the moment in mitosis when all kinetochores have undergone bipolar attachment to spindle microtubules. Topological linkages between sister chromatids are established through both DNA catenation and cohesin protein complexes (Murray and Szostak, 1985; Nasmyth and Haering, 2005; Yanagida, 2005). Each cohesin complex is proposed to embrace both sister chromatids by forming a ring composed of two structural maintenance of chromosome (Smc) proteins (in human cells Smc1 and Smc3), linked by a kleisin subunit (Scc1; also known as hRad21 and hHR21) and an accessory protein (Scc3/SA) (Haering et al., 2008; Losada, 2007). During early mitosis, most cohesin is dissociated from chromosomes through the action of Wapl and two mitotic kinases, Plk1 and Aurora B (Dai et al., 2006; Gandhi et al., 2006; Gimenez-Abian et al., 2004; Kueng et al., 2006; Losada et al., 2002; Sumara et al., 2002). However, cohesin at the centromere is protected from this so-called ‘prophase pathway’ by a centromere-associated protein, shugoshin 1 (Sgo1), that acts in concert with protein phosphatase 2A (PP2A), most likely to counteract Plk1-dependent cohesin phosphorylation (Kitajima et al., 2006; McGuinness et al., 2005; Salic et al., 2004; Watanabe, 2005). Centromere cohesion is abolished only after silencing of the spindle assembly checkpoint (SAC). This surveillance mechanism ensures the fidelity of chromosome segregation by inhibiting mitotic progression until bipolar attachment of all chromosomes to spindle microtubules is established (Musacchio and Salmon, 2007). In mammalian cells, silencing of the SAC appears to depend primarily on the distance between inner centromeres and outer kinetochores or intra-kinetochore tension (Liu et al., 2009; Maresca and Salmon, 2009; O’Connell et al., 2008; Uchida et al., 2009), although inter-kinetochore tension may be monitored in yeast (Pinsky and Biggins, 2005). The SAC is thought to operate primarily via the sequestration of Cdc20, a cofactor of the anaphase- promoting complex/cyclosome (APC/C), which controls the onset of anaphase by targeting cyclin B and securin for ubiquitin- mediated proteasomal degradation. The degradation of cyclin B and securin results in the inactivation of the cyclin-dependent kinase 1 (Cdk1) and the activation of separase, which in turns cleaves the centromeric cohesin and triggers the onset of anaphase (Huang et al., 2005; Stemmann et al., 2001; Uhlmann et al., 1999). Catenation of sister chromatid DNA is the direct consequence of DNA replication (Sundin and Varshavsky, 1980). In human cells, the decatenation of DNA double strands is brought about primarily by Topoisomerase-II (Topo-II; also known as DNA Topoisomerase-II) and most catenation along chromosome arms is resolved before metaphase (Porter and Farr, 2004). At centromeres, however, the resolution of catenated DNA appears to be completed only during anaphase, as inferred from recent studies demonstrating the persistence of PICH (Plk1-interacting checkpoint helicase; also known as DNA excision repair protein ERCC-6 like)- positive DNA threads upon inhibition of Topo-II in human anaphase cells (Baumann et al., 2007; Wang et al., 2008). Similarly, a requirement for Topo-II activity after anaphase onset has been shown in a number of different organisms (Downes et al., 1991; Gimenez-Abian et al., 2002; Shamu and Murray, 1992; Uemura et al., 1987). The precise structures of PICH-positive DNA threads in anaphase cells (also referred to as ultrafine DNA bridges; UFBs) remain to be clarified. In particular, it is intriguing that Bloom syndrome RecQ helicase (as well as its complex partners RMI1 and Topoisomerase III) associates with some UFBs (Bachrati and Hickson, 2008; Chan et al., 2007). Moreover, a subpopulation of UFBs was recently shown to connect fragile site loci associated with Fanconi anaemia proteins FANCD2 and FANCI, thus, probably Centromere DNA decatenation depends on cohesin removal and is required for mammalian cell division Lily Hui-Ching Wang 1,2, *, Bernd Mayer 3,‡ , Olaf Stemmann 3,‡ and Erich A. Nigg 1,2 1 Department of Cell Biology, Max-Planck Institute of Biochemistry, D-82152 Martinsried, Germany 2 Biozentrum, University of Basel, Klingelbergstrasse 50/70, CH-4056 Basel, Switzerland 3 Department of Molecular Cell Biology, Max-Planck Institute of Biochemistry, D-82152 Martinsried, Germany *Author for correspondence (lily.wang@unibas.ch) ‡ Present address: Department of Genetics, University of Bayreuth, D-95440 Bayreuth, Germany Accepted 19 December 2009 Journal of Cell Science 123, 806-813 © 2010. Published by The Company of Biologists Ltd doi:10.1242/jcs.058255 Summary Sister chromatid cohesion is mediated by DNA catenation and proteinaceous cohesin complexes. The recent visualization of PICH (Plk1-interacting checkpoint helicase)-coated DNA threads in anaphase cells raises new questions as to the role of DNA catenation and its regulation in time and space. In the present study we show that persistent DNA catenation induced by inhibition of Topoisomerase- II can contribute to sister chromatid cohesion in the absence of cohesin complexes and that resolution of catenation is essential for abscission. Furthermore, we use an in vitro chromatid separation assay to investigate the temporal and functional relationship between cohesin removal and Topoisomerase-II-mediated decatenation. Our data suggest that centromere decatenation can occur only after separase activation and cohesin removal, providing a plausible explanation for the persistence of centromere threads after anaphase onset. Key words: PICH, Topoisomerase-II, Cohesin, Spindle-assembly checkpoint, NoCut Journal of Cell Science