New insights into the formation and resolution of ultra-fine anaphase bridges

Abstract

Recent data indicate an unexpected requirement for proteins that were hitherto considered to be dedicated to DNA repair to facilitate the faithful disjunction of sister chromatids in anaphase. These include the Bloom's syndrome gene product, BLM and its partners, as well as a number of proteins that are important for preventing Fanconi anemia (FA) in man. As part of an analysis of the roles of these proteins in mitosis, we identified a novel class of anaphase bridge structure, called an ultra-fine anaphase bridge (UFB). These UFBs are also defined by the presence of a SNF2 family protein called PICH. In this review, we will discuss the possible sources of UFBs, and how the BLM, PICH and FA proteins might serve to process these structures in order to maintain genome stability.

Introduction

A human body contains more than 10 trillion cells. Remarkably, almost all of the cells inherit an identical set of chromosomes even after many rounds of cell division. In order to achieve such high fidelity in genomic maintenance, cells have developed elegant and carefully controlled DNA metabolic pathways to replicate, repair and segregate the genetic material. These pathways involve a large number of proteins that typically manipulate DNA structure in one way or another such as DNA polymerases, nucleases, topoisomerases and helicases. In this review, we will focus on a discussion of a particular family of DNA metabolising enzymes called the RecQ helicases. This is because defects of RecQ helicases cause severe genetic disorders in humans that are associated with an increase in genomic instability, and a predisposition to cancer and premature aging.

In humans, there are 5 members of RecQ helicase family: BLM, WRN, RecQ1, RecQ4 and RecQ5. Defects in BLM, WRN and RecQ4 are associated with genetic disorders; namely, Bloom's syndrome (BS), Werner's syndrome (WS) and Rothmund–Thomson syndrome (RTS), respectively. All of these disorders show signs of growth retardation, skin abnormalities and cancer predisposition, amongst other things. Patients with WS and RTS display premature aging, which is particularly striking in the case of WS, and are prone to the development of certain types of cancer, such as sarcomas and skin cancers. In contrast, BS patients are predisposed to all types of cancers that are found in general population. BS cells display an approximately 10-fold elevation in the frequency of sister-chromatids exchange (SCE), a marker for increased homologous recombination [1], [2], [3]. Therefore, BLM is of particular interest for studying the relationship between genomic instability and tumorigenesis.

RecQ helicases are highly conserved from bacteria to humans, primarily in the seven ‘signature’ helicase motifs (I, Ia, II–VI) [4]. In most cases, this domain is adjacent to a so-called RQC domain that interacts with DNA and contains a characteristic Zn²⁺ binding motif. RecQ helicases have Mg²⁺- and DNA-dependent ATPase activity, as well as 3′–5′ DNA helicase activity [5]. BLM and WRN, as well as their bacterial and budding yeast orthologues, RecQ and Sgs1, contain an additional conserved domain; the Helicase and RNaseD C-terminal (HRDC) domain, which has been shown also to be involved in DNA binding. In contrast, regions outside these domains are poorly conserved, and are thought to be involved in other functions such as mediating protein–protein interactions. A particularly conserved BLM binding partner is topoisomerase III (TopoIIIα in humans) [6], [7]. Recently, two additional partners, RMI1 and RMI2, were identified in humans, and studies showed that these partners have regulatory functions to facilitate the activities of BLM-TopoIIIα [8], [9], [10], [11], [12], [13]. Purified BLM, on its own, can unwind a variety of DNA structures that mimic replication and recombination intermediates. These include a partial duplex DNA with a 3′ tail, a forked structure, a D-loop structure and a 4-way/X-junction. It can also unwind non-B-form G-quadruplex (G4) DNA, a structure that is proposed to form in guanine-rich sequences such as telomeric and ribosomal DNA regions (Fig. 1A) [14]. Moreover, BLM can catalyze branch migration of a central intermediate in homologous recombination, the Holliday junction (HJ) (Fig. 1B) [2]. Very interestingly, acting in concert with TopoIII/RMI1, BLM and Sgs1 are capable of catalyzing a unique reaction, termed ‘double-HJ dissolution’ (Fig. 1C) which serves to complete homologous recombination reactions [2], [15], [16]. This reaction requires both HJ branch migration and ssDNA decatenation. Recently, two studies demonstrated that an additional function of Sgs1/TopoIII/Rmi1 is to facilitate 5′–3′ end resection of double stranded DNA catalyzed by the Dna2 nuclease [17], [18]. This reaction is proposed to initiate homologous recombination [19]. Overall, the findings indicate that the RecQ helicases play multiple roles in processing early and late DNA intermediates for replication and recombination. In this review, we will focus on another newly identified role of the BLM complex, which is to ensure faithful chromosome segregation. A detailed discussion of Bloom's syndrome and of BLM's biochemical properties can be found in other recent reviews [1], [2], [3].