Biochemistry, Genetics and Molecular Biology » "The Mechanisms of DNA Replication", book edited by David Stuart, ISBN 978-953-51-0991-4, Published: February 20, 2013 under CC BY 3.0 license. © The Author(s).

Chapter 3

The MCM and RecQ Helicase Families: Ancient Roles in DNA Replication and Genomic Stability Lead to Distinct Roles in Human Disease

By Dianne C. Daniel*, Ayuna V. Dagdanova and Edward M. Johnson
DOI: 10.5772/52961

Article top

Overview

Comparison of conserved ATPase motifs in RecQ proteins WRN, BLM and RECQL4 to the Walker A and Walker B boxes of MCM8. MCM8 was chosen as the MCM for comparison because it has a canonical GKS Walker A [12] and the signature MCM IDEFDKM Walker B ATP-binding domains. In the Walker B motif, note the conserved structural features and the conserved DE motif (containing aspartate, D, and the catalytic glutamate, E).
Figure 1. Comparison of conserved ATPase motifs in RecQ proteins WRN, BLM and RECQL4 to the Walker A and Walker B boxes of MCM8. MCM8 was chosen as the MCM for comparison because it has a canonical GKS Walker A [12] and the signature MCM IDEFDKM Walker B ATP-binding domains. In the Walker B motif, note the conserved structural features and the conserved DE motif (containing aspartate, D, and the catalytic glutamate, E).
Comparison of placement of selected structural motifs in human RecQ and MCM8 helicases. MCM8 has a splice variant that results in a 16 aa deletion in choriocarcinoma [12]. The Walker A and B boxes and the MCM Zn finger (ZnF) domains are indicated by filled rectangles. The MCM8 variant 2 deletion (Var2 del) is indicated by an open rectangle, as is its partially-homologous counterpart (del hom) in WRN and BLM. This conservatively structured domain consists of an N-terminal highly charged sequence followed by a conserved C and an aromatic-hydrophobic sequence. The open box is located in a different orientation relative to Walker A and B boxes in WRN vs. MCM8. The positions of black and white boxes are approximately to scale.
Figure 2. Comparison of placement of selected structural motifs in human RecQ and MCM8 helicases. MCM8 has a splice variant that results in a 16 aa deletion in choriocarcinoma [12]. The Walker A and B boxes and the MCM Zn finger (ZnF) domains are indicated by filled rectangles. The MCM8 variant 2 deletion (Var2 del) is indicated by an open rectangle, as is its partially-homologous counterpart (del hom) in WRN and BLM. This conservatively structured domain consists of an N-terminal highly charged sequence followed by a conserved C and an aromatic-hydrophobic sequence. The open box is located in a different orientation relative to Walker A and B boxes in WRN vs. MCM8. The positions of black and white boxes are approximately to scale.

The MCM and RecQ Helicase Families: Ancient Roles in DNA Replication and Genomic Stability Lead to Distinct Roles in Human Disease

Dianne C. Daniel*1, Ayuna V. Dagdanova1 and Edward M. Johnson1

1. Introduction

1.1. Rationale for comparison of MCM and RecQ helicase families

DNA helicases are currently organized into superfamilies based on their sequence structures and 3-D conformations. Within each superfamily, there are members that have further evolved for specialized functions [1]. There is conservation of RecQ proteins from bacteria to humans. Whereas bacteria have one RecQ helicase, humans have evolved at least five different proteins [2]. The RecQ members belong to the helicase Superfamily II, and as such have the characteristic Rec fold [1]. In this chapter, we will focus on RecQ family members WRN, BLM and RECQL4 (RecQ protein-like 4), which is also referred to in the literature as RECQ4. Eukaryotic and archaeal MCMs belong to the helicase Superfamily VI, and have the AAA+ (ATPases associated with diverse cellular activities) fold [1, 3, 4, 5]. Both Rec and AAA+ folds are based on the ancestral ASCE (additional strand conserved E) fold or an alpha-beta-alpha domain necessary for nucleoside triphosphate binding and catalysis [1, 6, 7]. A rationale for comparison of the RecQ and MCM family members relates to the importance of their activities for genomic integrity. The WRN and BLM proteins as well as other members of the RecQ family are characterized by this feature [8]. Both WRN and BLM are involved in DNA repair and a role for WRN in telomere homeostasis in humans is well established [2, 9]. MCM2-7 proteins, along with cofactors, are thought to function as the eukaryotic replicative helicase [10]. MCM8 [11, 12] and MCM9 [13, 14] are more recently discovered and their roles are less well defined. Although data point to a role for MCM8 in DNA replication, that role may be specialized in higher organisms. In human cells, MCM10 is recruited to chromosomal domains before they replicate and studies in yeast suggest a role in DNA replication, but not as a helicase [15-18]. Members of each family are essential for chromosome homeostasis. When replication forks stall, there may be involvement of members of each family. These proteins have interlocking functions since, for example, a stalled replication fork with attendant MCM proteins can lead to a DNA double-strand break (DSB), which requires RecQ proteins for repair [19].

2. RecQ and MCM family structures

Breaks in double-strand DNA can occur during DNA replication, at specific loci (e.g., at telomeres) and during meiosis. RecQ helicases are implicated in DNA repair based on their involvement in such processes as DNA end resection, branch migration, D-loop processing, Holliday Junction (HJ) and double Holliday junction (dHJ) resolution [2]. RecQ proteins function at multiple steps, both early and late, during repair of DSB [19]. RecQ helicases travel in a 3’ to 5’ direction on ssDNA [20]. The RecQ proteins have an ancient lineage based on an ancestral ASCE fold (αβα domain) of distant relation to P-loop NTPase folds. RecQ structural domains include a conserved core helicase domain for binding and hydrolysis of nucleoside triphosphate that is equivalent to the Walker A and Walker B boxes seen in MCM proteins [21]. They have a helicase and RNAase D C-terminal (HRDC) domain thought to mediate structure-specific nucleic acid binding, double HJ dissolution and protein-protein interactions. They also have a RecQ C-terminal (RQC) domain thought to mediate interactions with other proteins, structure-specific nucleic acid binding and metal cofactor binding [22]. Acidic regions present in many RecQ proteins aid in protein-protein interactions. There are also nuclear localization signals in some RecQ proteins (e.g., H. sapiens WRN and BLM). There are two RecQ members with an exonuclease domain, one of which is H. sapiens WRN. Two members have been functionally characterized as having an N-terminal strand exchange domain, one of which is H. sapiens BLM [2].

RECQL4 has been reported to have ssDNA binding and DNA strand-annealing activities. In this single study, RECQL4 did not display substrate unwinding or resolution of substrates resembling replication or recombination intermediates [23]. Recognizable HRDC and RQC domains that are important in BLM activity are missing in RECQL4 [22, 24]. As observed, the ssDNA annealing activity would allow RECQL4 to function during synthesis-dependent strand annealing (SDSA) along with another helicase. RECQL4 could help direct pathway choice during HR in DSB repair through aiding ssDNA annealing activity in non-homologous end joining (NHEJ) [19]. Thus, RECQL4 is similar to other RecQ helicases in its core helicase domain, but its function as a helicase is unclear [23, 25, 26].

AAA+ proteins, including MCMs, have a core molecular motor. Like the RecQ proteins, the AAA+ fold is also based on the ancestral ASCE fold. Acquisition of a catalytic glutamate (Fig. 1) to initiate efficient hydrolysis of ATP marked the emergence of the ASCE division from the ancestral P-loop fold [7]. For further discussion of the glutamate “switch” in AAA+ proteins, see reference [27]. Mechanisms of action are diverse, although members are typically oligomeric ring assemblies with inter-subunit communication. The central AAA+ motor has been adapted in evolution through structural changes to the core module and through domains added either N- or C-terminal to the AAA+ core. Activities are facilitated by recognition of protein partners functioning in these diverse events. Thus, AAA+ proteins display a variety of macromolecular remodelling events that are energy-driven by nucleotide hydrolysis thought to be occurring throughout what is typically a hexameric complex assembly [28, 29]. The conserved Walker A and Walker B motifs within the central module mediate ATP-binding and hydrolysis [7, 30-32]. MCM proteins have two active site motifs, the P-loop domain and the lid. Motifs in the P-loop include Walker A, Walker B and Sensor 1. The lid domain contains the arginine finger and Sensor 2. A catalytic site is created by a dimer interface that employs a cis P-loop from one subunit and a trans lid from the adjacent subunit [4, 33, 34]. A similar catalytic site created at the interfaces between adjacent monomers is also characteristic of the RecQ ATPase core [21].

media/image1.jpeg

Figure 1.

Comparison of conserved ATPase motifs in RecQ proteins WRN, BLM and RECQL4 to the Walker A and Walker B boxes of MCM8. MCM8 was chosen as the MCM for comparison because it has a canonical GKS Walker A [12] and the signature MCM IDEFDKM Walker B ATP-binding domains. In the Walker B motif, note the conserved structural features and the conserved DE motif (containing aspartate, D, and the catalytic glutamate, E).

3. RecQ and MCM helicases: association with disease and aging

3.1. BLM and Bloom syndrome, WRN and Werner syndrome, RECQL4 and Rothmund-Thomson syndrome

As a group, mutations in the RecQ helicases lead to adult segmental progeria with abnormalities in development, predisposition to cancer and acceleration of aging processes. Three of the RecQ family members are associated with rare autosomal recessive diseases [19]. These disorders Werner syndrome (WS) [35], Bloom syndrome (BS) [36] and Rothmund-Thomson syndrome (RTS) [37, 38] are caused by mutations in the genes coding for WRN, BLM and RECQL4, respectively. RTS is a heterogeneous disorder with about 60% of cases resulting from mutations in the RECQL4 gene [37]. Mutations in RECQL4 can also lead to two other disease phenotypes [39], but only RTS will be discussed here. The RecQ deficiency diseases are associated with cancer predisposition and several characteristics of aging [8, 20, 26, 40]. In BS cells, there is a 10-fold elevation in frequency of homologous recombination (HR), and reciprocal exchanges occur between homologous chromosomes and sister chromatids [41, 42]. WS cells, on the other hand, display large chromosome deletions and an increase in illegitimate recombination [43]. A higher frequency of chromosomal aberrations is reported for cells from RTS [44]. These deficiencies thus provide hints as to the cellular activities of these three helicases. Clinical features of these diseases, as referenced above, are as follows.

BS manifests in pleiotropic phenotypes such as growth retardation leading to proportional dwarfism, erythema with light sensitivity, skin lesions with hypo- and hyperpigmentation, immunodeficiency, susceptibility to type II diabetes, male infertility, female sub-fertility, reports of mental retardation, cancer predisposition (all types but at an earlier age of diagnosis than in the normal population).

WS leads to short stature and early onset age-related diseases, including greying hair, alopecia, bilateral cataracts, osteoporosis, arteriosclerosis, atherosclerosis, skin atrophy, hypogonadism, type II diabetes mellitus and susceptibility to tumors, especially those of mesenchymal origin (sarcomas).

RTS manifests as early growth deficiency, congenital bone defects, poikiloderma, cataracts, greying hair, alopecia, hypogonadism, and some increased susceptibility to cancer, especially osteogenic sarcomas.

3.2. MCMs and genomic stability

The MCM proteins 2-7 are necessary for DNA replication in yeast [45, 46], and this basic life function extends in evolution to a single MCM protein in archaea [47-49]. MCM2-7 are also essential for replication in Xenopus [50, 51] and have been proposed as a licensing factor for initiation of eukaryotic replication [52, 53]. The MCM proteins 2-8 have an identical Walker B-box motif of IDEFDKM. The MCM2-7 complex is enigmatic in that MCMs 4, 6 and 7 function alone as a heterohexameric helicase [54, 55]. For a discussion of individual MCM subunit arrangements and activities, see the references [33, 34]. MCM2-7 have now been shown to have helicase activity in vitro [56], and to be components of a holo-helicase Cdc45/MCM2-7/GINS (CMG) complex [57, 58]. The MCMs require a clamp-loading factor to assemble as a multimeric ring on DNA, and this function is fulfilled in known cases by the protein Cdc6 [59-64] although the regulation of this step in the formation of the CMG complex proceeds through multiple pathways [57, 58]. Various papers have dealt with the function of MCM proteins in DNA replication [10, 46, 65-68], and regarding their processive mechanism of DNA unwinding [27, 56, 58, 69, 70] and only certain lingering, disease-related questions will be considered here. A summary statement can be made regarding known relationships between MCM and RecQ helicases. Members of the MCM protein family are essential for the life-creating process of DNA replication, whereas members of the RecQ family are essential for the life-prolonging maintenance of the genome.

Due to their essential roles, it is not surprising that there are few diseases directly ascribed to defects in MCMs 2-7. This does, however, bring up an unresolved MCM enigma: there are more MCM proteins than are required to form initiation complexes at cellular origins active within a given round of DNA replication [71]. In addition, MCM proteins in human cells remain at peak levels in G2 phase of the cell cycle, after DNA replication is complete [59, 72, 73] This high copy number could be an aspect of securing sufficient protein quantities for basic function. A role for MCM proteins in transcription has also been proposed, and this remains to be defined [74]; for further discussion see reference [46]. Redundancy of functions among MCM members is also a factor to be considered. Intriguingly, knockdown of MCM8 [75], not even present in yeast [12], retards S-phase approximately 25% in human cells in culture [75], suggesting a specialized function in higher eukaryotes critical for basic replication. It has been suggested that the excess MCMs license dormant origins, which are used under conditions of stress upon normally functional origins [71]. An overview provides a discussion of recent work connecting dormant origins of replication and tumor suppression based on the role of dormant origins during fork restart after repair of DNA damage [76].

Mutations in MCM family members have been studied in yeast and mouse models, and they confirm an essential role for MCM2-7 in DNA replication [34, 45, 77, 78]. A human MCM4 mutation (destabilizing the MCM2-7 complex) was recently reported concurrently by two groups who studied consanguineous families, and the resulting phenotype was found to be associated with immune deficiency (NK cells), adrenal insufficiency and short stature [79, 80]. Patient fibroblasts showed chromosome fragility [79]. The susceptibility of these patients to cancer is not currently known [80]. An MCM8 disruption and alternative splice form have been noted in hepatic carcinoma [11] and choriocarcinoma [12], respectively. Although not necessarily a cause of disease, the MCM proteins may be useful tools in diagnoses [81-85]. Elevated levels of MCM proteins 2-7 have been observed in several cancers [81, 84, 85]. In contrast, reduced levels of MCM8 mRNA have been reported in colon carcinoma [12]. Nuclear MCM7 is a good marker for proliferating cells [86], and MCMs 2, 5 or 7 may be an alternative to the Ki-67 marker to distinguish certain hyperproliferative disorders [83].

3.2.1. A structural domain deleted from MCM8 and present in WRN and BLM

A brief discussion of MCM8 is included in this section because it contains a motif that may be structurally similar to one found in BLM and WRN and is linked to neoplasia. Human MCM8 has a splice variant that results in a 16 amino acid (aa) deletion in a location between the Zn finger of MCM8 and its Walker A box [12], Fig. 2. Thus far this deletion has been detected by various different groups only in cases of choriocarcinoma. MCM8 with this same sequence deleted (Fig. 2) has, however, been detected in several higher eukaryotes other than humans, suggesting that the variant does have, or perhaps lacks, a function. That function is as yet unknown, but clues to it may be gleaned from a comparison of the MCM8 deletion with sequences from WRN and BLM. These RecQ proteins have a counterpart 16 aa domain with partial sequence homology and notable structural homology, as denoted in Fig. 2. This sequence in the RecQ proteins is located in a different orientation to that of MCM8 with regard to the helicase Walker A and B boxes (Fig. 2). In each case the first 8 aa of this sequence are highly charged and, in WRN, BLM and MCM8, contain a polar S. The 9th aa in this deletion is a conserved C. The remaining 7 aa contain a preponderance of aromatic and hydrophobic residues. This configuration of charged and aromatic residues is characteristic of known single-stranded nucleic acid binding proteins [87]. Among MCMs, this 16 aa domain is identifiable in the single MCM of Sulfolobus solfataricus, in which it has been implicated as a single-stranded DNA-binding “finger” [88]. Mutations of the positively charged amino acids strongly reduce single-stranded DNA binding of this MCM. In contrast, in BLM the polar S residue is thought to be involved in ATP binding [89]. Because of the aromatic nature of a portion of this domain, binding to one or more DNA nucleotide bases may be involved as a common link between functions of these 16 aa in RECQ and MCM helicases.

media/image2.jpeg

Figure 2.

Comparison of placement of selected structural motifs in human RecQ and MCM8 helicases. MCM8 has a splice variant that results in a 16 aa deletion in choriocarcinoma [12]. The Walker A and B boxes and the MCM Zn finger (ZnF) domains are indicated by filled rectangles. The MCM8 variant 2 deletion (Var2 del) is indicated by an open rectangle, as is its partially-homologous counterpart (del hom) in WRN and BLM. This conservatively structured domain consists of an N-terminal highly charged sequence followed by a conserved C and an aromatic-hydrophobic sequence. The open box is located in a different orientation relative to Walker A and B boxes in WRN vs. MCM8. The positions of black and white boxes are approximately to scale.

4. Supportive roles for WRN, BLM helicases and RECQL4 during replication elongation

During normal metabolism, such as in mitochondrial respiration, endogenous reactive oxygen species (ROS) are produced that lead to oxidative DNA modifications. In addition to endogenous mutagens, there are also environmental mutagens that damage the DNA [90]. Furthermore, during replication and transcription, duplex DNA is transiently opened, and there is an opportunity for non-B DNA structures to form in the genomic DNA [40]. RecQ helicases act on recombination intermediates, on preferred substrates including those resembling G-quadruplex DNA [91, 92], D-loops [93], HJ [94] and double HJ [95]. RecQ helicases may play a regulatory role in both pro- and anti-HR events. They function at the interface of HR with the stressed replication fork and may affect repair pathway choice. There is little evidence for a specific clear cut role, but for a discussion of proposed mechanistic models of RecQ protein function, see the references [19, 20].

4.1. BLM and RECQL4

In BS cells, there are S-phase defects in DNA replication involving abnormal replication intermediates, and replication elongation is slower [96-98]. These cells don’t recover well from induced fork stalling and accumulate DSB [99, 100]. In human cells, RECQL4 interacts with the MCM2-7 complex, Cdc45 and GINS. The interaction is facilitated by MCM10 [25]. BLM and RECQL4 are at their highest levels during S phase. At stalled replication forks, BLM physically associates with Rad51 and p53, and BLM and p53 function synergistically in HR [101]. BLM colocalizes in foci with PCNA and with the BASC (BRCA1-associated genome surveillance complex) [102]. BLM is phosphorylated by ATR [103]. When replication forks are stalled by use of hydroxyurea (HU), BLM colocalizes with Chk1 and p53BP1 foci. Chk1 is required for BLM and 53BP1 foci formation. Thus Chk1 may recruit BLM to stalled forks [104]. This implicates RecQ proteins as DNA damage checkpoint mediators in response to stalled forks. In vitro studies with use of substrates similar to Okazaki fragments showed BLM stimulation of flap endonuclease [105] (a protein that functions in lagging strand synthesis [106]). BLM functions to prevent the association of homologous sequences in the displaced flap DNA of the Okazaki fragment and the sister chromatid [107-110]. D-loops are formed when a ssDNA tail invades a homologous duplex, and BLM has a preference for dissociating D-loops with a 5’ invaded end suggesting a selection of recombination intermediates that are not extended by polymerase [93, 111, 112]. BLM is able to disrupt the initial Rad51 filament formation step to destabilize recombinase-nucleoprotein filaments. D-loops are susceptible to BLM activity when Rad51 is in an inactive form (ADP-bound) [112, 113]. BLM physically associates with CAF-1 (chromatin assembly factor I) largest subunit, and the colocalization of these two proteins occurs at sites of DNA synthesis. BLM inhibits CAF-1 function in chromatin assembly during DNA repair in vitro, and inhibits its mobilization after damage induction in vivo [114]. Mammalian WRN and BLM interact [115], and they both interact with RPA [116-118] and p53 [119-121]. Based on coimmunoprecipitation there is limited BLM and WRN interaction, but they may function in the same pathway during HR [20]. BLM helicase activity is stimulated by its binding to the RPA70 kDa subunit [116]. In mouse spermatocytes during meiotic prophase, BLM and RPA are nuclear colocalized [122]. This suggests a potential role for these proteins in resolution of recombination intermediates during meiosis [8]. BLM has a preference for unwinding G-quadruplex structures versus HJ [92]. Both mammalian WRN and BLM bind to G-quadruplex structures, which are roadblocks to polymerases [123, 124].

Aberrant replication intermediates arise in cells lacking WRN and BLM [97, 125]. Such unresolved replication or recombination structures lead to incomplete chromosome segregation. BLM, topoisomerase 3 alpha (Topo3α) and BLAP75/RMI1 (for BLM-associated polypeptide/RecQ-mediated genome instability) or a BLM-Topo3α-BLAP75/RMI1 complex localizes to resulting anaphase bridges [126]. A helicase known as PICH arrives first, followed by the resolution activity of BLM [127]. As helicases unwind duplex DNA, torsional stress produced in the DNA may require relief through topoisomerase activity, and such activity finally decatenates interlocked DNA molecules [128, 129]. In vitro studies show that BLM can partner with Topo3α to resolve dHJ and prevent sequence exchange through resolution of this recombination intermediate [95]. The double-junction dissolution reaction requires the HRDC domain of BLM [24]. An additional protein, BLAP75/DMI1 mediates formation of the “dissolvasome” (BLM, Topo3α and BLAP75/RMI1) [130-132]. Mammalian BLAP18/RMI2 has also been found to be part of this complex [133].

4.2. WRN

Over 50 distinct WRN mutations have been reported, most of which lead to premature termination of translation [19]. Recent missense mutations in the exonuclease domain in one patient compromised protein stability [134]. Most mutations in WS patients occur in the WRN C-terminal domain, which could disrupt the WRN/p53 interaction [20, 134]. Such premature termination could also disrupt the Del hom sequence shown in Fig. 2. No WS mutations have been reported that eliminate only helicase or only exonuclease activity. Both activities are compromised in the development of WS [134]. WS fibroblasts undergo replicative senescence prematurely [135-139]. Telomere defects in WS cells suggest WRN activity in human telomere homeostasis [19]. Telomeres are needed to avoid loss of genetic material. They are important for chromosome end replication and for protection of the ends from enzymatic attack [140]. Human telomeres contain 5 to 20 kb of the repetitive sequence TTAGGG [141]. At the terminal there are 100-200 bp of 3’ ssDNA overhang. This overhang can anneal with telomere DNA to form a stable D-loop leading to a structure referred to as a ‘t-loop’ [142, 143]. Alternatively, this free unannealed end may form G-quadruplex DNA [144]. Human WRN functions in lagging-strand synthesis, and in the replication of telomeric G-rich DNA ends [145]. C. elegans WRN-1 can disrupt D-loops [146] and human WRN can prevent aberrant recombination [147]. WRN 3’ to 5’ exonuclease is stimulated by the interacting Ku70/Ku86 complex supporting a role for WRN in DNA repair [148]. Evidence suggests that in the absence of telomerase, WRN and BLM have a role in the ALT (Alternate Lengthening of Telomeres) pathway for telomere maintenance [149, 150]. In biochemical experiments, WRN releases a 3‘ invading tail from a telomeric type D-loop by coordinated WRN helicase and exonuclease activities [149].

WRN and BLM catalytic activities are comparable except for 3’ to 5’ exonuclease activity of WRN [151-153], which BLM lacks. On dsDNA and on RNA-DNA hybrids, the WRN exonuclease activity degrades a 3’ recessed end. This activity can remove only one mismatched NT at the end of the recessed 3’ DNA and can initiate exonuclease activity from a gap or nick [154, 155]. The exonuclease activity of WRN can degrade abnormal DNA structures suggesting that WRN helicase and exonuclease activities are involved in resolution of aberrant DNA structures at stalled forks [156]. Human WRN interacts with proteins involved in DNA replication. WRN coimmunoprecipitates with PCNA and topoisomerase 1 [157]. WRN functionally and physically interacts with RPA [117], and it functionally interacts with DNA polymerase delta [152]. WS cells accumulate recombination intermediates that impede cell growth [158]. In cells treated with HU, WRN colocalizes with RPA foci and is thought to dissociate recombination intermediates at the stalled forks [94, 147]. WRN stimulates polymerase delta activity in the absence of its processivity factor PCNA. This suggests a role for WRN in recruiting polymerase delta for replication restart at blocked or collapsed forks [152]. RPA can stimulate the processivity of WRN. The stimulation of WRN by RPA is due to protein-protein interactions as opposed to enhanced ATPase activity [117, 118].

5. Unification of BLM, WRN, RECQL4 and MCM2-7 activities in DNA replication and recombination/repair

5.1. BLM: Role in DNA damage response with a complex role in inhibiting or promoting HR

BLM is found mostly in fine granules throughout the nucleoplasm at highest levels during S and G2 phases of the cell cycle. Its focal localization is in PML nuclear bodies (PML-NB). The name PML derives from the promyelocytic leukemia protein, PML [159-162]. This protein forms the structural groundwork of the PML bodies, which store various nuclear proteins [163]. These PML-NB store repair proteins (e.g., Topo3α, MRN and p53) and may be involved in sensing DNA damage [163]. By regulating the availability of repair proteins, response can be directed to DNA damage sites. Trafficking of proteins to the PML-NB is regulated by sumoylation [164]. The sumoylation pathway involves E1, E2 and E3 enzymes, which regulate respectively, SUMO activation, SUMO conjugation and targeting of specific substrates for sumoylation through ligation [165]. BLM contains a motif for SUMO binding that would facilitate its integration into this repair protein storage network [166]. In addition, BLM is SUMO-1 and SUMO-2 modified [167]. When mutants are prepared in which the SUMO-binding sequences are deleted, BLM cannot localize to PML-NB [168]. When mutants are prepared that do not allow BLM localization to PML-NB, there is about a two-fold increase in sister chromatid exchange. These findings indicate that there is a need for BLM-SUMO interaction in order for BLM to localize to PML-NB, and that BLM activity, such as its accumulation at stalled replication forks, may be regulated by this specific localization [168].

At sites of DSB, repair foci form. A central player, H2AX, is phosphorylated when DSBs are induced, and this phosphorylation involves ATM, ATR and DNA-PK. Over one million bp are then marked by phosphorylated H2AX (γH2AX) on each side of the break [112, 168-171]. γH2AX recruits additional repair proteins to the damage site [172]. In studies where normal S-phase cells are treated with DNA damaging agents (HU, UV and cross-linking agents), BLM responds by leaving the PML-NBs to relocate to repair foci and colocalize with the marker γH2AX [101, 173]. BLM interacts physically and functionally with γH2AX as well as with ATM and ATR [103, 173-175]. In damage that is S-phase specific, BLM associates with the complex ATR/CHK1/53BP1, which gathers at repair foci as an early response to the damage. Based on kinetic studies, BLM may facilitate BRCA1 and MRN complex localization at repair foci in S phase [173], which may involve BLM regulation of p53 in these foci [101]. This early function of BLM at repair foci may allow for BML to influence the choice of repair pathways and facilitate a BLM function in anti-recombination at stalled forks that perhaps involves SUMO regulation [19].

As discussed in the previous section, BLM interacts with the recombinase, Rad51. Rad51 catalyzes the pairing of a ssDNA tail and a homologous stretch of dsDNA to promote strand exchange early in the HR pathway [111]. Following ionizing radiation, Rad51 foci contained BLM [159]. Rad51 functions in HR, and localizes to ssDNA when DSB are induced [176]. BML can displace the recombinase from the ssDNA filament [112], which can be viewed as an anti-recombinagenic function [19]. In Rad51-associated D-loops, BLM can interact with Rad51 and unwind DNA in front of the polymerase [177]. This could favor SDSA leading to pro-recombinagenic function [19, 177]. BLM, however, also functions in resolution of G-quadruplex DNA structure and has higher binding affinity for it compared to HJ. BLM helicase activity is required for resolution of this structure [92]. Whereas the BLM/Rad51 interaction would represent an early event in a DNA damage/repair process, the formation of HJ, on the other hand, is a late event in HR. As discussed in section 5.1, the BLM-Topo3α-BLAP75/RMI1 complex functions to “dissolve” dHJ by convergent fork migration to generate non-cross-over products [178]. This is facilitated by Topo3α relief of superhelicity and by its ability to cleave and rejoin one strand of a DNA duplex. In the absence of BLM, Topo3α activity would involve break and rejoining activities instead of dissolution, which could lead to crossover events and an increase in sister chromatid exchange in BS cells [19]. BLM that is mutated to be unable to interact with Topo3α can only partially rescue the frequency of sister chromatid exchange [168]. Thus, together in a complex, BLM and Topo3α achieve dissolution of a recombinogenic intermediate. BLM has been proposed to regulate ploidy based on a role along with other members of this complex in resolution of anaphase bridges [126]. BLM has been found in complexes with mismatch repair protein MLH1 [179, 180]. It has been found to be present in large complexes containing not only BLM-Topo3a-BLAF75, but also additional factors. These additional factors could include several BLAF factors, proteins from the FA (Fanconi anemia) pathway, RPA, and mismatch repair protein MLH1. [181, 182]. The interactive role of these pathway components remains to be determined.

5.2. WRN: Helicase and exonuclease activities in concert

When WRN biochemical activities were compared using nonhydrolyzable ATPγS to inhibit only WRN helicase activity or aa substitutions to eliminate only the WRN exonuclease activity, the exonuclease activity was shown to function in degradation of the leading strand on replication fork-type substrates and in degradation of the annealed telomere overhangs on substrates resembling D-loop structures. WRN binding proteins were inhibitory [149, 183]. In addition, WRN was found to degrade ssDNA substrates longer than 40 nt with dependence upon the helicase activity [183]. WRN activities were also explored using WS fibroblasts. In WS cells, WRN and the enzyme telomerase are able to reverse the phenotype of excess chromosome fusion. In these cells, the anaphase bridges were missing telomere DNA. Dominant negative telomerase was not able to rescue the phenotype indicating that a stable telomere length was needed for rescue [184]. Telomeres are stabilized by a complex of proteins that bind DNA, known as the shelterin complex [185]. This complex consists of TRF1, TRF2 (double-strand DNA binding proteins), and POT1 (single-strand DNA binding protein) as well as adaptor proteins. The coordinate action of these proteins in the presence of telomerase is needed to regulate telomere length. Without telomerase, telomeres shorten in length each cell division [185]. In S phase, WRN colocalizes with TRF2 [145, 186]. TRF1 and TRF2 limit WRN exonuclease activity on synthetic telomere D-loops [149]. WRN helicase is stimulated by single-stranded DNA binding proteins, RPA or POT1. These proteins modulate WRN exonuclease degradation of the 3’ overhang [149, 187]. The hypothesis is that WRN could unwind D-loops to facilitate leading strand synthesis through telomeric DNA, and there is also the possibility that WRN prevents interchromosomal interactions between telomeres [19].

5.2.1. WRN activities in response to G-quadruplex structures in the lagging strand

At the telomere, the G-rich strand is duplicated by lagging-strand synthesis. It may assume a G-quadruplex structure, which would interrupt the replication fork. Experimentally, when inhibiting WRN by use of overexpression of a dominant-negative WRN, the helicase deficiency leads to loss almost entirely affecting the sister telomere on the lagging strand [145]. WRN interaction with FEN-1 (the flap endonuclease involved in the processing of Okazaki fragments, [188]) could assist in the maturation of these fragments [105, 189]. Data suggest that WRN may function in concert with lagging strand synthesis perhaps in unwinding complex structure at the telomere. WRN interacts physically with DNA polymerase delta to stimulate its activity [190]. WRN can prevent stalling of polymerases delta at telomere sequence in vitro [191]. WRN stimulation of polymerase delta happens only in the absence of PCNA, which suggests that WRN has a role at stalled forks rather than in the regulation of processive DNA synthesis [19]. WRN, as opposed to BLM, is specific for G-quadruplex structure in the trinucleotide repeat of Fragile X syndrome [192].

5.2.2. WRN and Ku in suppression of aberrant recombination at telomeres

Proteins involved in the NHEJ path for repair of DSBs that are found at telomeres include Ku heterodimer, DNA-PK, MRN complex and ATM [193]. Through interaction with NBS1, WRN colocalizes with two of these components, Ku and the MRN complex [194]. Under unstable conditions, Ku can suppress sister chromatid exchange at telomeres [195]. Mouse knockout studies show that WRN normally suppresses aberrant recombination at telomeres [196].

5.2.3. WRN activities in base excision repair and interstrand cross link repair by HR

WS cells show increased sensitivity to alkylating agents and to agents that increase ROS. Human fibroblasts in which WRN has been knocked down respond to oxidative stress with an increase in DNA damage [197]. These observations support a role for WRN in repair of such damage [198]. DNA damage resulting from oxidation, alkylation, methylation and deamination is repaired by the base excision repair (BER) pathway. WRN interacts with proteins in this pathway. In addition, WRN helicase activity stimulates and is stimulated by polymerase beta activity [199]. There is evidence that WRN activity in BER is regulated by PARP-1, but the reader is referred to references [200, 201]. BRCA1 and BRCA2 play an important role in DSB repair in the HR subpathway [202, 203]. HR is a pathway for repairing interstrand crosslinks (ICL). Cells deficient in either WRN or BRCA1 are hypersensitive to induction of ICLs [204, 205]. A physical interaction of WRN with BRCA1 enhances the activities of WRN [206]. In addition, data from WRN and/or BRCA1 knockdown studies indicate that BRCA1 may act cooperatively with WRN in HR during ICL repair [206].

5.3. RECQL4

Data from Recql4-/- mice indicate a role for mouse RECQL4 in genomic stability and in the promoting cohesion between sister chromatids [207]. Depleting the X. laevis homologue of RTS, xRTS, in Xenopus egg extracts led to reduced DNA synthesis and an inhibition of RPA stabilization of ssDNA prior to polymerase loading at unwound origins. The addition of purified human RECQL4 could reverse this effect [208]. A nonhelical region in the N terminus of xRTS could be important in initiation of DNA replication based on its interaction with the Cut5 protein and the importance of xRTS in loading DNA polymerase alpha onto chromatin [209]. The N terminus of xRTS is not homologous to that of RECQL4 in mammals, however, and this role may not be conserved [19]. RECQL4, along with Ctf4 and MCM10, has been shown to be required for stable association of the CMG complex in human cells. In this study, Cut5/TopBP1 was not required for CMG stabilization [210]. In the Xenopus replication model, RECQL4 binds to chromatin that has been processed to resemble DSBs with a dependence on RPA and ATM activity [211]. Chromatin immunoprecipitation experiments show that RECQL4 functions on DNA in close association with Ku [212]. In HeLa cells human RECQL4 forms a complex with Rad51 and colocalizes with Rad51 foci formed after treatment with etoposide [212]. PML-NB contain a portion of RECQL4 [212]. It is also found in the nucleolus [213]. When using a T7 phage display screen, RECQL4 was found to interact with PARP-1, and the association influenced the nuclear localization of RECQL4. PARP-1 has a role in RECQL4 movement to the nucleolus from the nucleoplasm. When oxidative stress is induced, as opposed to other types of damage induction, RECQL4 increasingly localizes in the nucleolus. This trafficking is inhibited by inhibition of PARP [213]. RTS cells decrease proliferation and synthesis of DNA when exposed to hydrogen peroxide [214]. Lack of proper response to ROS could lead to premature aging as see in RTS. PARP is also involved in an end-joining pathway of DNA repair [215, 216]. The role of RECQL4 in its interaction with PARP-1 is not known (for further discussion see reference [19]).

5.4. MCM2-7

Replication fork stalling, can lead to DSB and chromosomal rearrangements. An S phase checkpoint is triggered by these events and there is a block to elongation. When this occurs, the proteins Mrc1, Tof1, and Csm3 (M/T/C complex) interact with the MCM2-7 complex to stabilize the replication fork. When the M/T/C complex is missing, the replisome continues, but synthesis stops. This may be partially due to loss of DNA polymerase epsilon from the fork [217-219]. Studies in yeast provide insight into these activities. The M/T/C complex associates with MCM2-7 [218, 220-222] and also with polymerase epsilon [221, 223]. These physical interactions permit communication between polymerase epsilon and the MCM complex [223]. The M/T/C complex may be part of the normal replication fork protein entourage [218, 220, 221, 224]. Mrc1 and Cdc45 coimmunoprecipitate, indicating an interaction of Mrc1 with the The Replication Progression Complex core, which includes Cdc45, the MCMs and GINS [223]. In each cell cycle Mrc loads onto replication origins along with the polymerases. This occurs after the Replication Progression Complex forms. Mrc migrates with the replication forks. [218, 219, 224-226]. Tof1, like Mrc, also coimmunoprecipitates with Cdc45 [227]. The exact mechanism of action of the M/T/C complex is not known. In a yeast study, the Tof1 homologue could switch regulation between pro- and anti-recombination activities in a site-specific manner [228]. Data indicate that a Mrc1 and MCM6-C terminal interaction senses alkylated DNA damage [221]. The other two subunits Tof1 and Csm3, may function to sense other types of damage. Although the helicase domains of MCM2-7 are conserved, the N and C terminals are divergent. Other negative regulators could differentially bind to these regions to regulate powering of elongation by the MCM2-7 helicase during times of stress [10]. A future question relates to the extent to which leading or lagging strand polymerase arrest is associated with formation of ssDNA, fork regression and formation of abnormal DNA structures [66]. These data indicate a functional connection between the MCM proteins, which act at stalled forks, and the RecQ proteins, which facilitate repair of the resulting damage.

6. Summary

BLM, WRN and RECQL4 act during events that stress the advancing replication fork providing relief through DNA damage repair and through resolution of aberrant replication/ recombination intermediates, including those present at the telomere. At checkpoint, the replication proteins at a stalled fork are held stable through communication that occurs due to proteins that bind and signal to both the MCM complex and polymerase(s). This would allow repair proteins such as WRN and BLM helicases and RECQL4 to resolve the stress and thus aid in fork restart. Advancing our knowledge of the RecQ and MCM family activities and the mechanisms and signalling behind these activities will increase our understanding of cancer and aging and perhaps enlighten us regarding how to accommodate these challenges to human health.

Acknowledgments

This. work was supported by NIH funding to EMJ and resources derived from grant funding from Virginia’s Commonwealth Health Research Board to DCD.

References

1 - Berger JM. SnapShot: nucleic acid helicases and translocases. Cell. 2008 Sep 5;134(5):888- e1. 18775318
2 - Bernstein KA, Gangloff S, Rothstein R. The RecQ DNA helicases in DNA repair. Annu Rev Genet. 2010;44:393-417. 21047263
3 - Kelman Z, Hurwitz J. Structural lessons in DNA replication from the third domain of life. Nat Struct Biol. 2003 Mar;10(3):148-50. 12605215
4 - Erzberger JP, Berger JM. Evolutionary relationships and structural mechanisms of AAA+ proteins. Annu Rev Biophys Biomol Struct. 2006;35:93-114. 16689629
5 - Koonin EV. A common set of conserved motifs in a vast variety of putative nucleic acid-dependent ATPases including MCM proteins involved in the initiation of eukaryotic DNA replication. Nucleic Acids Res. 1993 Jun 11;21(11):2541-7. 8332451
6 - Iyer LM, Leipe DD, Koonin EV, Aravind L. Evolutionary history and higher order classification of AAA+ ATPases. J Struct Biol. 2004 Apr-May;146(1-2):11-31. 15037234
7 - Leipe DD, Koonin EV, Aravind L. Evolution and classification of P-loop kinases and related proteins. J Mol Biol. 2003 Oct 31;333(4):781-815. 14568537
8 - Mohaghegh P, Hickson ID. DNA helicase deficiencies associated with cancer predisposition and premature ageing disorders. Hum Mol Genet. 2001 Apr;10(7):741-6. 11257107
9 - Brosh RM, Jr., Bohr VA. Human premature aging, DNA repair and RecQ helicases. Nucleic Acids Res. 2007;35(22):7527-44. 18006573
10 - Bochman ML, Schwacha A. The Mcm complex: unwinding the mechanism of a replicative helicase. Microbiol Mol Biol Rev. 2009 Dec;73(4):652-83. 19946136
11 - Gozuacik D, Chami M, Lagorce D, Faivre J, Murakami Y, Poch O, Biermann E, Knippers R, Brechot C, Paterlini-Brechot P. Identification and functional characterization of a new member of the human Mcm protein family: hMcm8. Nucleic Acids Res. 2003 Jan 15;31(2):570-9. 12527764
12 - Johnson EM, Kinoshita Y, Daniel DC. A new member of the MCM protein family encoded by the human MCM8 gene, located contrapodal to GCD10 at chromosome band 20p12.3-13. Nucleic Acids Res. 2003 Jun 1;31(11):2915-25. 12771218
13 - Lutzmann M, Maiorano D, Mechali M. Identification of full genes and proteins of MCM9, a novel, vertebrate-specific member of the MCM2-8 protein family. Gene. 2005 Dec 5;362:51-6. 16226853
14 - Yoshida K. Identification of a novel cell-cycle-induced MCM family protein MCM9. Biochem Biophys Res Commun. 2005 Jun 3;331(2):669-74. 15850810
15 - Fien K, Cho YS, Lee JK, Raychaudhuri S, Tappin I, Hurwitz J. Primer utilization by DNA polymerase alpha-primase is influenced by its interaction with Mcm10p. J Biol Chem. 2004 Apr 16;279(16):16144-53. 14766746
16 - Ricke RM, Bielinsky AK. Mcm10 regulates the stability and chromatin association of DNA polymerase-alpha. Mol Cell. 2004 Oct 22;16(2):173-85. 15494305
17 - Sawyer SL, Cheng IH, Chai W, Tye BK. Mcm10 and Cdc45 cooperate in origin activation in Saccharomyces cerevisiae. J Mol Biol. 2004 Jul 2;340(2):195-202. 15201046
18 - Yang X, Gregan J, Lindner K, Young H, Kearsey SE. Nuclear distribution and chromatin association of DNA polymerase alpha-primase is affected by TEV protease cleavage of Cdc23 (Mcm10) in fission yeast. BMC Mol Biol. 2005;6:13. 15941470
19 - Ouyang KJ, Woo LL, Ellis NA. Homologous recombination and maintenance of genome integrity: cancer and aging through the prism of human RecQ helicases. Mech Ageing Dev. 2008 Jul-Aug;129(7-8):425-40. 18430459
20 - Bachrati CZ, Hickson ID. RecQ helicases: suppressors of tumorigenesis and premature aging. Biochem J. 2003 Sep 15;374(Pt 3):577-606. 12803543
21 - Singleton MR, Dillingham MS, Wigley DB. Structure and mechanism of helicases and nucleic acid translocases. Annu Rev Biochem. 2007;76:23-50. 17506634
22 - Killoran MP, Keck JL. Sit down, relax and unwind: structural insights into RecQ helicase mechanisms. Nucleic Acids Res. 2006;34(15):4098-105. 16935877
23 - Macris MA, Krejci L, Bussen W, Shimamoto A, Sung P. Biochemical characterization of the RECQ4 protein, mutated in Rothmund-Thomson syndrome. DNA Repair (Amst). 2006 Feb 3;5(2):172-80. 16214424
24 - Wu L, Chan KL, Ralf C, Bernstein DA, Garcia PL, Bohr VA, Vindigni A, Janscak P, Keck JL, Hickson ID. The HRDC domain of BLM is required for the dissolution of double Holliday junctions. Embo J. 2005 Jul 20;24(14):2679-87. 15990871
25 - Xu X, Rochette PJ, Feyissa EA, Su TV, Liu Y. MCM10 mediates RECQ4 association with MCM2-7 helicase complex during DNA replication. Embo J. 2009 Oct 7;28(19):3005-14. 19696745
26 - Chu WK, Hickson ID. RecQ helicases: multifunctional genome caretakers. Nat Rev Cancer. 2009 Sep;9(9):644-54. 19657341
27 - Mogni ME, Costa A, Ioannou C, Bell SD. The glutamate switch is present in all seven clades of AAA+ protein. Biochemistry. 2009 Sep 22;48(37):8774-5. 19702328
28 - Snider J, Thibault G, Houry WA. The AAA+ superfamily of functionally diverse proteins. Genome Biol. 2008;9(4):216. 18466635
29 - Davey MJ, Jeruzalmi D, Kuriyan J, O'Donnell M. Motors and switches: AAA+ machines within the replisome. Nat Rev Mol Cell Biol. 2002 Nov;3(11):826-35. 12415300
30 - Ogura T, Wilkinson AJ. AAA+ superfamily ATPases: common structure--diverse function. Genes Cells. 2001 Jul;6(7):575-97. 11473577
31 - Saraste M, Sibbald PR, Wittinghofer A. The P-loop--a common motif in ATP- and GTP-binding proteins. Trends Biochem Sci. 1990 Nov;15(11):430-4. 2126155
32 - Walker JE, Saraste M, Runswick MJ, Gay NJ. Distantly related sequences in the alpha- and beta-subunits of ATP synthase, myosin, kinases and other ATP-requiring enzymes and a common nucleotide binding fold. Embo J. 1982;1(8):945-51. 6329717
33 - Davey MJ, Indiani C, O'Donnell M. Reconstitution of the Mcm2-7p heterohexamer, subunit arrangement, and ATP site architecture. J Biol Chem. 2003 Feb 14;278(7):4491-9. 12480933
34 - Schwacha A, Bell SP. Interactions between two catalytically distinct MCM subgroups are essential for coordinated ATP hydrolysis and DNA replication. Mol Cell. 2001 Nov;8(5):1093-104. 11741544
35 - Yu CE, Oshima J, Fu YH, Wijsman EM, Hisama F, Alisch R, Matthews S, Nakura J, Miki T, Ouais S, Martin GM, Mulligan J, Schellenberg GD. Positional cloning of the Werner's syndrome gene. Science. 1996 Apr 12;272(5259):258-62. 8602509
36 - Bloom D. Congenital telangiectatic erythema resembling lupus erythematosus in dwarfs; probably a syndrome entity. AMA Am J Dis Child. 1954 Dec;88(6):754-8. 13206391
37 - Kitao S, Lindor NM, Shiratori M, Furuichi Y, Shimamoto A. Rothmund-thomson syndrome responsible gene, RECQL4: genomic structure and products. Genomics. 1999 Nov 1;61(3):268-76. 10552928
38 - Siitonen HA, Sotkasiira J, Biervliet M, Benmansour A, Capri Y, Cormier-Daire V, Crandall B, Hannula-Jouppi K, Hennekam R, Herzog D, Keymolen K, Lipsanen-Nyman M, Miny P, Plon SE, Riedl S, Sarkar A, Vargas FR, Verloes A, Wang LL, Kaariainen H, Kestila M. The mutation spectrum in RECQL4 diseases. Eur J Hum Genet. 2009 Feb;17(2):151-8. 18716613
39 - Hanada K, Hickson ID. Molecular genetics of RecQ helicase disorders. Cell Mol Life Sci. 2007 Sep;64(17):2306-22. 17571213
40 - Karow JK, Wu L, Hickson ID. RecQ family helicases: roles in cancer and aging. Curr Opin Genet Dev. 2000 Feb;10(1):32-8. 10679384
41 - German J. Bloom syndrome: a mendelian prototype of somatic mutational disease. Medicine (Baltimore). 1993 Nov;72(6):393-406. 8231788
42 - German J. Bloom's syndrome. Dermatol Clin. 1995 Jan;13(1):7-18. 7712653
43 - Shen JC, Loeb LA. The Werner syndrome gene: the molecular basis of RecQ helicase-deficiency diseases. Trends Genet. 2000 May;16(5):213-20. 10782115
44 - Vasseur F, Delaporte E, Zabot MT, Sturque MN, Barrut D, Savary JB, Thomas L, Thomas P. Excision repair defect in Rothmund Thomson syndrome. Acta Derm Venereol. 1999 Mar;79(2):150-2. 10228638
45 - Tye BK. MCM proteins in DNA replication. Annu Rev Biochem. 1999;68:649-86. 10872463
46 - Forsburg SL. Eukaryotic MCM proteins: beyond replication initiation. Microbiol Mol Biol Rev. 2004 Mar;68(1):109-31. 15007098
47 - Chong JP, Hayashi MK, Simon MN, Xu RM, Stillman B. A double-hexamer archaeal minichromosome maintenance protein is an ATP-dependent DNA helicase. Proc Natl Acad Sci U S A. 2000 Feb 15;97(4):1530-5. 10677495
48 - Kelman Z, Lee JK, Hurwitz J. The single minichromosome maintenance protein of Methanobacterium thermoautotrophicum DeltaH contains DNA helicase activity. Proc Natl Acad Sci U S A. 1999 Dec 21;96(26):14783-8. 10611290
49 - Tye BK. Insights into DNA replication from the third domain of life. Proc Natl Acad Sci U S A. 2000 Mar 14;97(6):2399-401. 10716976
50 - Madine MA, Swietlik M, Pelizon C, Romanowski P, Mills AD, Laskey RA. The roles of the MCM, ORC, and Cdc6 proteins in determining the replication competence of chromatin in quiescent cells. J Struct Biol. 2000 Apr;129(2-3):198-210. 10806069
51 - Romanowski P, Madine MA, Rowles A, Blow JJ, Laskey RA. The Xenopus origin recognition complex is essential for DNA replication and MCM binding to chromatin. Curr Biol. 1996 Nov 1;6(11):1416-25. 8939603
52 - Blow JJ, Hodgson B. Replication licensing--defining the proliferative state? Trends Cell Biol. 2002 Feb;12(2):72-8. 11849970
53 - Madine MA, Khoo CY, Mills AD, Musahl C, Laskey RA. The nuclear envelope prevents reinitiation of replication by regulating the binding of MCM3 to chromatin in Xenopus egg extracts. Curr Biol. 1995 Nov 1;5(11):1270-9. 8574584
54 - Ishimi Y. A DNA helicase activity is associated with an MCM4, -6, and -7 protein complex. J Biol Chem. 1997 Sep 26;272(39):24508-13. 9305914
55 - Lee JK, Hurwitz J. Processive DNA helicase activity of the minichromosome maintenance proteins 4, 6, and 7 complex requires forked DNA structures. Proc Natl Acad Sci U S A. 2001 Jan 2;98(1):54-9. 11136247
56 - Bochman ML, Schwacha A. The Mcm2-7 complex has in vitro helicase activity. Mol Cell. 2008 Jul 25;31(2):287-93. 18657510
57 - Boos D, Frigola J, Diffley JF. Activation of the replicative DNA helicase: breaking up is hard to do. Curr Opin Cell Biol. 2012 Jun;24(3):423-30. 22424671
58 - Kang YH, Galal WC, Farina A, Tappin I, Hurwitz J. Properties of the human Cdc45/Mcm2-7/GINS helicase complex and its action with DNA polymerase epsilon in rolling circle DNA synthesis. Proc Natl Acad Sci U S A. 2012 Apr 17;109(16):6042-7. 22474384
59 - Kinoshita Y, Johnson EM. Site-specific loading of an MCM protein complex in a DNA replication initiation zone upstream of the c-MYC gene in the HeLa cell cycle. J Biol Chem. 2004 Aug 20;279(34):35879-89. 15190069
60 - Schepers A, Diffley JF. Mutational analysis of conserved sequence motifs in the budding yeast Cdc6 protein. J Mol Biol. 2001 May 11;308(4):597-608. 11350163
61 - Nevis KR, Cordeiro-Stone M, Cook JG. Origin licensing and p53 status regulate Cdk2 activity during G(1). Cell Cycle. 2009 Jun 15;8(12):1952-63. 19440053
62 - Coverley D, Pelizon C, Trewick S, Laskey RA. Chromatin-bound Cdc6 persists in S and G2 phases in human cells, while soluble Cdc6 is destroyed in a cyclin A-cdk2 dependent process. J Cell Sci. 2000 Jun;113 ( Pt 11):1929-38. 10806104
63 - Alexandrow MG, Hamlin JL. Cdc6 chromatin affinity is unaffected by serine-54 phosphorylation, S-phase progression, and overexpression of cyclin A. Mol Cell Biol. 2004 Feb;24(4):1614-27. 14749377
64 - Shin JH, Grabowski B, Kasiviswanathan R, Bell SD, Kelman Z. Regulation of minichromosome maintenance helicase activity by Cdc6. J Biol Chem. 2003 Sep 26;278(39):38059-67. 12837750
65 - Daniel DC, Johnson EM. Addressing the enigma of MCM8 in DNA replication. In: Kušić-Tišma J, ed. Fundamental Aspects of DNA Replication: INTECH 2011:37-52. DOI: 10.5772/21177. http://www.intechopen.com/books/fundamental-aspects-of-dna-replication/addressing-the-enigma-of-mcm8-in-dna-replication
66 - Forsburg SL. The MCM helicase: linking checkpoints to the replication fork. Biochem Soc Trans. 2008 Feb;36(Pt 1):114-9. 18208397
67 - Masai H, You Z, Arai K. Control of DNA replication: regulation and activation of eukaryotic replicative helicase, MCM. IUBMB Life. 2005 Apr-May;57(4-5):323-35. 16036617
68 - Costa A, Onesti S. The MCM complex: (just) a replicative helicase? Biochem Soc Trans. 2008 Feb;36(Pt 1):136-40. 18208401
69 - Brewster AS, Wang G, Yu X, Greenleaf WB, Carazo JM, Tjajadi M, Klein MG, Chen XS. Crystal structure of a near-full-length archaeal MCM: functional insights for an AAA+ hexameric helicase. Proc Natl Acad Sci U S A. 2008 Dec 23;105(51):20191-6. 19073923
70 - Makarova KS, Koonin EV, Kelman Z. The CMG (CDC45/RecJ, MCM, GINS) complex is a conserved component of the DNA replication system in all archaea and eukaryotes. Biol Direct. 2012;7:7. 22329974
71 - Woodward AM, Gohler T, Luciani MG, Oehlmann M, Ge X, Gartner A, Jackson DA, Blow JJ. Excess Mcm2-7 license dormant origins of replication that can be used under conditions of replicative stress. J Cell Biol. 2006 Jun 5;173(5):673-83. 16754955
72 - Claycomb JM, MacAlpine DM, Evans JG, Bell SP, Orr-Weaver TL. Visualization of replication initiation and elongation in Drosophila. J Cell Biol. 2002 Oct 28;159(2):225-36. 12403810
73 - Hirai K, Shirakata M. Replication licensing of the EBV oriP minichromosome. Curr Top Microbiol Immunol. 2001;258:13-33. 11443858
74 - Yankulov K, Todorov I, Romanowski P, Licatalosi D, Cilli K, McCracken S, Laskey R, Bentley DL. MCM proteins are associated with RNA polymerase II holoenzyme. Mol Cell Biol. 1999 Sep;19(9):6154-63. 10454562
75 - Volkening M, Hoffmann I. Involvement of human MCM8 in prereplication complex assembly by recruiting hcdc6 to chromatin. Mol Cell Biol. 2005 Feb;25(4):1560-8. 15684404
76 - Klotz-Noack K, Blow JJ. A role for dormant origins in tumor suppression. Mol Cell. 2011 Mar 4;41(5):495-6. 21362544
77 - Gomez EB, Catlett MG, Forsburg SL. Different phenotypes in vivo are associated with ATPase motif mutations in Schizosaccharomyces pombe minichromosome maintenance proteins. Genetics. 2002 Apr;160(4):1305-18. 11973289
78 - You Z, Komamura Y, Ishimi Y. Biochemical analysis of the intrinsic Mcm4-Mcm6-mcm7 DNA helicase activity. Mol Cell Biol. 1999 Dec;19(12):8003-15. 10567526
79 - Gineau L, Cognet C, Kara N, Lach FP, Dunne J, Veturi U, Picard C, Trouillet C, Eidenschenk C, Aoufouchi S, Alcais A, Smith O, Geissmann F, Feighery C, Abel L, Smogorzewska A, Stillman B, Vivier E, Casanova JL, Jouanguy E. Partial MCM4 deficiency in patients with growth retardation, adrenal insufficiency, and natural killer cell deficiency. J Clin Invest. 2012 Mar 1;122(3):821-32. 22354167
80 - Hughes CR, Guasti L, Meimaridou E, Chuang CH, Schimenti JC, King PJ, Costigan C, Clark AJ, Metherell LA. MCM4 mutation causes adrenal failure, short stature, and natural killer cell deficiency in humans. J Clin Invest. 2012 Mar 1;122(3):814-20. 22354170
81 - Chatrath P, Scott IS, Morris LS, Davies RJ, Rushbrook SM, Bird K, Vowler SL, Grant JW, Saeed IT, Howard D, Laskey RA, Coleman N. Aberrant expression of minichromosome maintenance protein-2 and Ki67 in laryngeal squamous epithelial lesions. Br J Cancer. 2003 Sep 15;89(6):1048-54. 12966424
82 - Davidson EJ, Morris LS, Scott IS, Rushbrook SM, Bird K, Laskey RA, Wilson GE, Kitchener HC, Coleman N, Stern PL. Minichromosome maintenance (Mcm) proteins, cyclin B1 and D1, phosphohistone H3 and in situ DNA replication for functional analysis of vulval intraepithelial neoplasia. Br J Cancer. 2003 Jan 27;88(2):257-62. 12610511
83 - Freeman A, Morris LS, Mills AD, Stoeber K, Laskey RA, Williams GH, Coleman N. Minichromosome maintenance proteins as biological markers of dysplasia and malignancy. Clin Cancer Res. 1999;5(8):2121-32.
84 - Gonzalez MA, Pinder SE, Callagy G, Vowler SL, Morris LS, Bird K, Bell JA, Laskey RA, Coleman N. Minichromosome maintenance protein 2 is a strong independent prognostic marker in breast cancer. J Clin Oncol. 2003 Dec 1;21(23):4306-13. 14645419
85 - Hunt DP, Freeman A, Morris LS, Burnet NG, Bird K, Davies TW, Laskey RA, Coleman N. Early recurrence of benign meningioma correlates with expression of mini-chromosome maintenance-2 protein. Br J Neurosurg. 2002 Feb;16(1):10-5. 11928726
86 - Khalili K, Del Valle L, Muralidharan V, Gault WJ, Darbinian N, Otte J, Meier E, Johnson EM, Daniel DC, Kinoshita Y, Amini S, Gordon J. Puralpha Is Essential for Postnatal Brain Development and Developmentally Coupled Cellular Proliferation As Revealed by Genetic Inactivation in the Mouse. Mol Cell Biol. 2003 Oct 1;23(19):6857-75. 12972605
87 - Bergemann AD, Johnson EM. The HeLa Pur factor binds single-stranded DNA at a specific element conserved in gene flanking regions and origins of DNA replication. Mol Cell Biol. 1992;12:1257-65.
88 - Liu W, Pucci B, Rossi M, Pisani FM, Ladenstein R. Structural analysis of the Sulfolobus solfataricus MCM protein N-terminal domain. Nucleic Acids Res. 2008 Jun;36(10):3235-43. 18417534
89 - Rong SB, Valiaho J, Vihinen M. Structural basis of Bloom syndrome (BS) causing mutations in the BLM helicase domain. Mol Med. 2000 Mar;6(3):155-64. 10965492
90 - Lindahl T. Instability and decay of the primary structure of DNA. Nature. 1993 Apr 22;362(6422):709-15. 8469282
91 - Sun H, Karow JK, Hickson ID, Maizels N. The Bloom's syndrome helicase unwinds G4 DNA. J Biol Chem. 1998 Oct 16;273(42):27587-92. 9765292
92 - Huber MD, Lee DC, Maizels N. G4 DNA unwinding by BLM and Sgs1p: substrate specificity and substrate-specific inhibition. Nucleic Acids Res. 2002 Sep 15;30(18):3954-61. 12235379
93 - van Brabant AJ, Ye T, Sanz M, German IJ, Ellis NA, Holloman WK. Binding and melting of D-loops by the Bloom syndrome helicase. Biochemistry. 2000 Nov 28;39(47):14617-25. 11087418
94 - Karow JK, Constantinou A, Li JL, West SC, Hickson ID. The Bloom's syndrome gene product promotes branch migration of holliday junctions. Proc Natl Acad Sci U S A. 2000 Jun 6;97(12):6504-8. 10823897
95 - Wu L, Hickson ID. The Bloom's syndrome helicase suppresses crossing over during homologous recombination. Nature. 2003 Dec 18;426(6968):870-4. 14685245
96 - Hand R, German J. A retarded rate of DNA chain growth in Bloom's syndrome. Proc Natl Acad Sci U S A. 1975 Feb;72(2):758-62. 1054854
97 - Lonn U, Lonn S, Nylen U, Winblad G, German J. An abnormal profile of DNA replication intermediates in Bloom's syndrome. Cancer Res. 1990 Jun 1;50(11):3141-5. 2110504
98 - Han H, Hurley LH. G-quadruplex DNA: a potential target for anti-cancer drug design. Trends Pharmacol Sci. 2000 Apr;21(4):136-42. 10740289
99 - Davies SL, North PS, Hickson ID. Role for BLM in replication-fork restart and suppression of origin firing after replicative stress. Nat Struct Mol Biol. 2007 Jul;14(7):677-9. 17603497
100 - Rao VA, Conti C, Guirouilh-Barbat J, Nakamura A, Miao ZH, Davies SL, Sacca B, Hickson ID, Bensimon A, Pommier Y. Endogenous gamma-H2AX-ATM-Chk2 checkpoint activation in Bloom's syndrome helicase deficient cells is related to DNA replication arrested forks. Mol Cancer Res. 2007 Jul;5(7):713-24. 17634426
101 - Sengupta S, Linke SP, Pedeux R, Yang Q, Farnsworth J, Garfield SH, Valerie K, Shay JW, Ellis NA, Wasylyk B, Harris CC. BLM helicase-dependent transport of p53 to sites of stalled DNA replication forks modulates homologous recombination. Embo J. 2003 Mar 3;22(5):1210-22. 12606585
102 - Wang Y, Cortez D, Yazdi P, Neff N, Elledge SJ, Qin J. BASC, a super complex of BRCA1-associated proteins involved in the recognition and repair of aberrant DNA structures. Genes Dev. 2000 Apr 15;14(8):927-39. 10783165
103 - Davies SL, North PS, Dart A, Lakin ND, Hickson ID. Phosphorylation of the Bloom's syndrome helicase and its role in recovery from S-phase arrest. Mol Cell Biol. 2004 Feb;24(3):1279-91. 14729972
104 - Sengupta S, Robles AI, Linke SP, Sinogeeva NI, Zhang R, Pedeux R, Ward IM, Celeste A, Nussenzweig A, Chen J, Halazonetis TD, Harris CC. Functional interaction between BLM helicase and 53BP1 in a Chk1-mediated pathway during S-phase arrest. J Cell Biol. 2004 Sep 13;166(6):801-13. 15364958
105 - Brosh RM, Jr., Driscoll HC, Dianov GL, Sommers JA. Biochemical characterization of the WRN-FEN-1 functional interaction. Biochemistry. 2002 Oct 8;41(40):12204-16. 12356323
106 - Kao HI, Veeraraghavan J, Polaczek P, Campbell JL, Bambara RA. On the roles of Saccharomyces cerevisiae Dna2p and Flap endonuclease 1 in Okazaki fragment processing. J Biol Chem. 2004 Apr 9;279(15):15014-24. 14747468
107 - Bartos JD, Wang W, Pike JE, Bambara RA. Mechanisms by which Bloom protein can disrupt recombination intermediates of Okazaki fragment maturation. J Biol Chem. 2006 Oct 27;281(43):32227-39. 16950766
108 - Sharma S, Otterlei M, Sommers JA, Driscoll HC, Dianov GL, Kao HI, Bambara RA, Brosh RM, Jr. WRN helicase and FEN-1 form a complex upon replication arrest and together process branchmigrating DNA structures associated with the replication fork. Mol Biol Cell. 2004 Feb;15(2):734-50. 14657243
109 - Sharma S, Sommers JA, Gary RK, Friedrich-Heineken E, Hubscher U, Brosh RM, Jr. The interaction site of Flap Endonuclease-1 with WRN helicase suggests a coordination of WRN and PCNA. Nucleic Acids Res. 2005;33(21):6769-81. 16326861
110 - Sharma S, Sommers JA, Wu L, Bohr VA, Hickson ID, Brosh RM, Jr. Stimulation of flap endonuclease-1 by the Bloom's syndrome protein. J Biol Chem. 2004 Mar 12;279(11):9847-56. 14688284
111 - Bachrati CZ, Borts RH, Hickson ID. Mobile D-loops are a preferred substrate for the Bloom's syndrome helicase. Nucleic Acids Res. 2006;34(8):2269-79. 16670433
112 - Bugreev DV, Yu X, Egelman EH, Mazin AV. Novel pro- and anti-recombination activities of the Bloom's syndrome helicase. Genes Dev. 2007 Dec 1;21(23):3085-94. 18003860
113 - Bugreev DV, Brosh RM, Jr., Mazin AV. RECQ1 possesses DNA branch migration activity. J Biol Chem. 2008 Jul 18;283(29):20231-42. 18495662
114 - Jiao R, Bachrati CZ, Pedrazzi G, Kuster P, Petkovic M, Li JL, Egli D, Hickson ID, Stagljar I. Physical and functional interaction between the Bloom's syndrome gene product and the largest subunit of chromatin assembly factor 1. Mol Cell Biol. 2004 Jun;24(11):4710-9. 15143166
115 - von Kobbe C, Karmakar P, Dawut L, Opresko P, Zeng X, Brosh RM, Jr., Hickson ID, Bohr VA. Colocalization, physical, and functional interaction between Werner and Bloom syndrome proteins. J Biol Chem. 2002 Jun 14;277(24):22035-44. 11919194
116 - Brosh RM, Jr., Li JL, Kenny MK, Karow JK, Cooper MP, Kureekattil RP, Hickson ID, Bohr VA. Replication protein A physically interacts with the Bloom's syndrome protein and stimulates its helicase activity. J Biol Chem. 2000 Aug 4;275(31):23500-8. 10825162
117 - Brosh RM, Jr., Orren DK, Nehlin JO, Ravn PH, Kenny MK, Machwe A, Bohr VA. Functional and physical interaction between WRN helicase and human replication protein A. J Biol Chem. 1999 Jun 25;274(26):18341-50. 10373438
118 - Shen JC, Gray MD, Oshima J, Loeb LA. Characterization of Werner syndrome protein DNA helicase activity: directionality, substrate dependence and stimulation by replication protein A. Nucleic Acids Res. 1998 Jun 15;26(12):2879-85. 9611231
119 - Blander G, Kipnis J, Leal JF, Yu CE, Schellenberg GD, Oren M. Physical and functional interaction between p53 and the Werner's syndrome protein. J Biol Chem. 1999 Oct 8;274(41):29463-9. 10506209
120 - Spillare EA, Robles AI, Wang XW, Shen JC, Yu CE, Schellenberg GD, Harris CC. p53-mediated apoptosis is attenuated in Werner syndrome cells. Genes Dev. 1999 Jun 1;13(11):1355-60. 10364153
121 - Wang XW, Tseng A, Ellis NA, Spillare EA, Linke SP, Robles AI, Seker H, Yang Q, Hu P, Beresten S, Bemmels NA, Garfield S, Harris CC. Functional interaction of p53 and BLM DNA helicase in apoptosis. J Biol Chem. 2001 Aug 31;276(35):32948-55. 11399766
122 - Walpita D, Plug AW, Neff NF, German J, Ashley T. Bloom's syndrome protein, BLM, colocalizes with replication protein A in meiotic prophase nuclei of mammalian spermatocytes. Proc Natl Acad Sci U S A. 1999 May 11;96(10):5622-7. 10318934
123 - Li JL, Harrison RJ, Reszka AP, Brosh RM, Jr., Bohr VA, Neidle S, Hickson ID. Inhibition of the Bloom's and Werner's syndrome helicases by G-quadruplex interacting ligands. Biochemistry. 2001 Dec 18;40(50):15194-202. 11735402
124 - Popuri V, Bachrati CZ, Muzzolini L, Mosedale G, Costantini S, Giacomini E, Hickson ID, Vindigni A. The Human RecQ helicases, BLM and RECQ1, display distinct DNA substrate specificities. J Biol Chem. 2008 Jun 27;283(26):17766-76. 18448429
125 - Poot M, Hoehn H, Runger TM, Martin GM. Impaired S-phase transit of Werner syndrome cells expressed in lymphoblastoid cell lines. Exp Cell Res. 1992 Oct;202(2):267-73. 1327851
126 - Chan KL, North PS, Hickson ID. BLM is required for faithful chromosome segregation and its localization defines a class of ultrafine anaphase bridges. Embo J. 2007 Jul 25;26(14):3397-409. 17599064
127 - Baumann C, Korner R, Hofmann K, Nigg EA. PICH, a centromere-associated SNF2 family ATPase, is regulated by Plk1 and required for the spindle checkpoint. Cell. 2007 Jan 12;128(1):101-14. 17218258
128 - Johnson FB, Lombard DB, Neff NF, Mastrangelo MA, Dewolf W, Ellis NA, Marciniak RA, Yin Y, Jaenisch R, Guarente L. Association of the Bloom syndrome protein with topoisomerase IIIalpha in somatic and meiotic cells. Cancer Res. 2000 Mar 1;60(5):1162-7. 10728666
129 - Wu L, Davies SL, North PS, Goulaouic H, Riou JF, Turley H, Gatter KC, Hickson ID. The Bloom's syndrome gene product interacts with topoisomerase III. J Biol Chem. 2000 Mar 31;275(13):9636-44. 10734115
130 - Bussen W, Raynard S, Busygina V, Singh AK, Sung P. Holliday junction processing activity of the BLM-Topo IIIalpha-BLAP75 complex. J Biol Chem. 2007 Oct 26;282(43):31484-92. 17728255
131 - Raynard S, Zhao W, Bussen W, Lu L, Ding YY, Busygina V, Meetei AR, Sung P. Functional role of BLAP75 in BLM-topoisomerase IIIalpha-dependent holliday junction processing. J Biol Chem. 2008 Jun 6;283(23):15701-8. 18390547
132 - Wu L, Bachrati CZ, Ou J, Xu C, Yin J, Chang M, Wang W, Li L, Brown GW, Hickson ID. BLAP75/RMI1 promotes the BLM-dependent dissolution of homologous recombination intermediates. Proc Natl Acad Sci U S A. 2006 Mar 14;103(11):4068-73. 16537486
133 - Singh TR, Ali AM, Busygina V, Raynard S, Fan Q, Du CH, Andreassen PR, Sung P, Meetei AR. BLAP18/RMI2, a novel OB-fold-containing protein, is an essential component of the Bloom helicase-double Holliday junction dissolvasome. Genes Dev. 2008 Oct 15;22(20):2856-68. 18923083
134 - Huang S, Lee L, Hanson NB, Lenaerts C, Hoehn H, Poot M, Rubin CD, Chen DF, Yang CC, Juch H, Dorn T, Spiegel R, Oral EA, Abid M, Battisti C, Lucci-Cordisco E, Neri G, Steed EH, Kidd A, Isley W, Showalter D, Vittone JL, Konstantinow A, Ring J, Meyer P, Wenger SL, von Herbay A, Wollina U, Schuelke M, Huizenga CR, Leistritz DF, Martin GM, Mian IS, Oshima J. The spectrum of WRN mutations in Werner syndrome patients. Hum Mutat. 2006 Jun;27(6):558-67. 16673358
135 - Davis T, Singhrao SK, Wyllie FS, Haughton MF, Smith PJ, Wiltshire M, Wynford-Thomas D, Jones CJ, Faragher RG, Kipling D. Telomere-based proliferative lifespan barriers in Werner-syndrome fibroblasts involve both p53-dependent and p53-independent mechanisms. J Cell Sci. 2003 Apr 1;116(Pt 7):1349-57. 12615976
136 - Epstein CJ, Martin GM, Schultz AL, Motulsky AG. Werner's syndrome a review of its symptomatology, natural history, pathologic features, genetics and relationship to the natural aging process. Medicine (Baltimore). 1966 May;45(3):177-221. 5327241
137 - Faragher RG, Kill IR, Hunter JA, Pope FM, Tannock C, Shall S. The gene responsible for Werner syndrome may be a cell division "counting" gene. Proc Natl Acad Sci U S A. 1993 Dec 15;90(24):12030-4. 8265666
138 - Schulz VP, Zakian VA, Ogburn CE, McKay J, Jarzebowicz AA, Edland SD, Martin GM. Accelerated loss of telomeric repeats may not explain accelerated replicative decline of Werner syndrome cells. Hum Genet. 1996 Jun;97(6):750-4. 8641691
139 - Tahara H, Tokutake Y, Maeda S, Kataoka H, Watanabe T, Satoh M, Matsumoto T, Sugawara M, Ide T, Goto M, Furuichi Y, Sugimoto M. Abnormal telomere dynamics of B-lymphoblastoid cell strains from Werner's syndrome patients transformed by Epstein-Barr virus. Oncogene. 1997 Oct 16;15(16):1911-20. 9365237
140 - Sandell LL, Zakian VA. Loss of a yeast telomere: arrest, recovery, and chromosome loss. Cell. 1993 Nov 19;75(4):729-39. 8242745
141 - Nurnberg P, Thiel G, Weber F, Epplen JT. Changes of telomere lengths in human intracranial tumours. Hum Genet. 1993 Mar;91(2):190-2. 8462979
142 - Griffith JD, Comeau L, Rosenfield S, Stansel RM, Bianchi A, Moss H, de Lange T. Mammalian telomeres end in a large duplex loop. Cell. 1999 May 14;97(4):503-14. 10338214
143 - Murti KG, Prescott DM. Telomeres of polytene chromosomes in a ciliated protozoan terminate in duplex DNA loops. Proc Natl Acad Sci U S A. 1999 Dec 7;96(25):14436-9. 10588723
144 - Williamson JR, Raghuraman MK, Cech TR. Monovalent cation-induced structure of telomeric DNA: the G-quartet model. Cell. 1989 Dec 1;59(5):871-80. 2590943
145 - Crabbe L, Verdun RE, Haggblom CI, Karlseder J. Defective telomere lagging strand synthesis in cells lacking WRN helicase activity. Science. 2004 Dec 10;306(5703):1951-3. 15591207
146 - Hyun M, Bohr VA, Ahn B. Biochemical characterization of the WRN-1 RecQ helicase of Caenorhabditis elegans. Biochemistry. 2008 Jul 15;47(28):7583-93. 18558712
147 - Constantinou A, Tarsounas M, Karow JK, Brosh RM, Bohr VA, Hickson ID, West SC. Werner's syndrome protein (WRN) migrates Holliday junctions and co-localizes with RPA upon replication arrest. EMBO Rep. 2000 Jul;1(1):80-4. 11256630
148 - Cooper MP, Machwe A, Orren DK, Brosh RM, Ramsden D, Bohr VA. Ku complex interacts with and stimulates the Werner protein. Genes Dev. 2000 Apr 15;14(8):907-12. 10783163
149 - Opresko PL, Otterlei M, Graakjaer J, Bruheim P, Dawut L, Kolvraa S, May A, Seidman MM, Bohr VA. The Werner syndrome helicase and exonuclease cooperate to resolve telomeric D loops in a manner regulated by TRF1 and TRF2. Mol Cell. 2004 Jun 18;14(6):763-74. 15200954
150 - Bhattacharyya S, Sandy A, Groden J. Unwinding protein complexes in ALTernative telomere maintenance. J Cell Biochem. 2010 2010;109(1):7-15. 19911388
151 - Huang S, Li B, Gray MD, Oshima J, Mian IS, Campisi J. The premature ageing syndrome protein, WRN, is a 3'-->5' exonuclease. Nat Genet. 1998 Oct;20(2):114-6. 9771700
152 - Kamath-Loeb AS, Johansson E, Burgers PM, Loeb LA. Functional interaction between the Werner Syndrome protein and DNA polymerase delta. Proc Natl Acad Sci U S A. 2000 Apr 25;97(9):4603-8. 10781066
153 - Shen JC, Gray MD, Oshima J, Kamath-Loeb AS, Fry M, Loeb LA. Werner syndrome protein. I. DNA helicase and dna exonuclease reside on the same polypeptide. J Biol Chem. 1998 Dec 18;273(51):34139-44. 9852073
154 - Huang S, Beresten S, Li B, Oshima J, Ellis NA, Campisi J. Characterization of the human and mouse WRN 3'-->5' exonuclease. Nucleic Acids Res. 2000 Jun 15;28(12):2396-405. 10871373
155 - Kamath-Loeb AS, Shen JC, Loeb LA, Fry M. Werner syndrome protein. II. Characterization of the integral 3' --> 5' DNA exonuclease. J Biol Chem. 1998 Dec 18;273(51):34145-50. 9852074
156 - Shen JC, Loeb LA. Werner syndrome exonuclease catalyzes structure-dependent degradation of DNA. Nucleic Acids Res. 2000 Sep 1;28(17):3260-8. 10954593
157 - Lebel M, Spillare EA, Harris CC, Leder P. The Werner syndrome gene product co-purifies with the DNA replication complex and interacts with PCNA and topoisomerase I. J Biol Chem. 1999 Dec 31;274(53):37795-9. 10608841
158 - Prince PR, Emond MJ, Monnat RJ, Jr. Loss of Werner syndrome protein function promotes aberrant mitotic recombination. Genes Dev. 2001 Apr 15;15(8):933-8. 11316787
159 - Bischof O, Kim SH, Irving J, Beresten S, Ellis NA, Campisi J. Regulation and localization of the Bloom syndrome protein in response to DNA damage. J Cell Biol. 2001 Apr 16;153(2):367-80. 11309417
160 - Dutertre S, Ababou M, Onclercq R, Delic J, Chatton B, Jaulin C, Amor-Gueret M. Cell cycle regulation of the endogenous wild type Bloom's syndrome DNA helicase. Oncogene. 2000 May 25;19(23):2731-8. 10851073
161 - Sanz MM, Proytcheva M, Ellis NA, Holloman WK, German J. BLM, the Bloom's syndrome protein, varies during the cell cycle in its amount, distribution, and co-localization with other nuclear proteins. Cytogenet Cell Genet. 2000;91(1-4):217-23. 11173860
162 - Zhong S, Hu P, Ye TZ, Stan R, Ellis NA, Pandolfi PP. A role for PML and the nuclear body in genomic stability. Oncogene. 1999 Dec 23;18(56):7941-7. 10637504
163 - Dellaire G, Ching RW, Ahmed K, Jalali F, Tse KC, Bristow RG, Bazett-Jones DP. Promyelocytic leukemia nuclear bodies behave as DNA damage sensors whose response to DNA double-strand breaks is regulated by NBS1 and the kinases ATM, Chk2, and ATR. J Cell Biol. 2006 Oct 9;175(1):55-66. 17030982
164 - Matunis MJ, Zhang XD, Ellis NA. SUMO: the glue that binds. Dev Cell. 2006 Nov;11(5):596-7. 17084352
165 - Johnson ES. Protein modification by SUMO. Annu Rev Biochem. 2004;73:355-82. 15189146
166 - Shen TH, Lin HK, Scaglioni PP, Yung TM, Pandolfi PP. The mechanisms of PML-nuclear body formation. Mol Cell. 2006 Nov 3;24(3):331-9. 17081985
167 - Eladad S, Ye TZ, Hu P, Leversha M, Beresten S, Matunis MJ, Ellis NA. Intra-nuclear trafficking of the BLM helicase to DNA damage-induced foci is regulated by SUMO modification. Hum Mol Genet. 2005 May 15;14(10):1351-65. 15829507
168 - Hu P, Beresten SF, van Brabant AJ, Ye TZ, Pandolfi PP, Johnson FB, Guarente L, Ellis NA. Evidence for BLM and Topoisomerase IIIalpha interaction in genomic stability. Hum Mol Genet. 2001 Jun 1;10(12):1287-98. 11406610
169 - Paull TT, Rogakou EP, Yamazaki V, Kirchgessner CU, Gellert M, Bonner WM. A critical role for histone H2AX in recruitment of repair factors to nuclear foci after DNA damage. Curr Biol. 2000 Jul 27-Aug 10;10(15):886-95. 10959836
170 - Pilch DR, Sedelnikova OA, Redon C, Celeste A, Nussenzweig A, Bonner WM. Characteristics of gamma-H2AX foci at DNA double-strand breaks sites. Biochem Cell Biol. 2003 Jun;81(3):123-9. 12897845
171 - Rogakou EP, Boon C, Redon C, Bonner WM. Megabase chromatin domains involved in DNA double-strand breaks in vivo. J Cell Biol. 1999 Sep 6;146(5):905-16. 10477747
172 - Bassing CH, Alt FW. H2AX may function as an anchor to hold broken chromosomal DNA ends in close proximity. Cell Cycle. 2004 Feb;3(2):149-53. 14712078
173 - Davalos AR, Campisi J. Bloom syndrome cells undergo p53-dependent apoptosis and delayed assembly of BRCA1 and NBS1 repair complexes at stalled replication forks. J Cell Biol. 2003 Sep 29;162(7):1197-209. 14517203
174 - Beamish H, Kedar P, Kaneko H, Chen P, Fukao T, Peng C, Beresten S, Gueven N, Purdie D, Lees-Miller S, Ellis N, Kondo N, Lavin MF. Functional link between BLM defective in Bloom's syndrome and the ataxia-telangiectasia-mutated protein, ATM. J Biol Chem. 2002 Aug 23;277(34):30515-23. 12034743
175 - Franchitto A, Pichierri P. Bloom's syndrome protein is required for correct relocalization of RAD50/MRE11/NBS1 complex after replication fork arrest. J Cell Biol. 2002 Apr 1;157(1):19-30. 11916980
176 - Raderschall E, Golub EI, Haaf T. Nuclear foci of mammalian recombination proteins are located at single-stranded DNA regions formed after DNA damage. Proc Natl Acad Sci U S A. 1999 Mar 2;96(5):1921-6. 10051570
177 - Adams MD, McVey M, Sekelsky JJ. Drosophila BLM in double-strand break repair by synthesis-dependent strand annealing. Science. 2003 Jan 10;299(5604):265-7. 12522255
178 - Plank JL, Wu J, Hsieh TS. Topoisomerase IIIalpha and Bloom's helicase can resolve a mobile double Holliday junction substrate through convergent branch migration. Proc Natl Acad Sci U S A. 2006 Jul 25;103(30):11118-23. 16849422
179 - Langland G, Kordich J, Creaney J, Goss KH, Lillard-Wetherell K, Bebenek K, Kunkel TA, Groden J. The Bloom's syndrome protein (BLM) interacts with MLH1 but is not required for DNA mismatch repair. J Biol Chem. 2001 Aug 10;276(32):30031-5. 11325959
180 - Pedrazzi G, Perrera C, Blaser H, Kuster P, Marra G, Davies SL, Ryu GH, Freire R, Hickson ID, Jiricny J, Stagljar I. Direct association of Bloom's syndrome gene product with the human mismatch repair protein MLH1. Nucleic Acids Res. 2001 Nov 1;29(21):4378-86. 11691925
181 - Meetei AR, Sechi S, Wallisch M, Yang D, Young MK, Joenje H, Hoatlin ME, Wang W. A multiprotein nuclear complex connects Fanconi anemia and Bloom syndrome. Mol Cell Biol. 2003 May;23(10):3417-26. 12724401
182 - Yin J, Sobeck A, Xu C, Meetei AR, Hoatlin M, Li L, Wang W. BLAP75, an essential component of Bloom's syndrome protein complexes that maintain genome integrity. Embo J. 2005 Apr 6;24(7):1465-76. 15775963
183 - Machwe A, Xiao L, Lloyd RG, Bolt E, Orren DK. Replication fork regression in vitro by the Werner syndrome protein (WRN): holliday junction formation, the effect of leading arm structure and a potential role for WRN exonuclease activity. Nucleic Acids Res. 2007;35(17):5729-47. 17717003
184 - Crabbe L, Jauch A, Naeger CM, Holtgreve-Grez H, Karlseder J. Telomere dysfunction as a cause of genomic instability in Werner syndrome. Proc Natl Acad Sci U S A. 2007 Feb 13;104(7):2205-10. 17284601
185 - de Lange T. Shelterin: the protein complex that shapes and safeguards human telomeres. Genes Dev. 2005 Sep 15;19(18):2100-10. 16166375
186 - Opresko PL, von Kobbe C, Laine JP, Harrigan J, Hickson ID, Bohr VA. Telomere-binding protein TRF2 binds to and stimulates the Werner and Bloom syndrome helicases. J Biol Chem. 2002 Oct 25;277(43):41110-9. 12181313
187 - Opresko PL, Mason PA, Podell ER, Lei M, Hickson ID, Cech TR, Bohr VA. POT1 stimulates RecQ helicases WRN and BLM to unwind telomeric DNA substrates. J Biol Chem. 2005 Sep 16;280(37):32069-80. 16030011
188 - Kao HI, Campbell JL, Bambara RA. Dna2p helicase/nuclease is a tracking protein, like FEN1, for flap cleavage during Okazaki fragment maturation. J Biol Chem. 2004 Dec 3;279(49):50840-9. 15448135
189 - Sharma S, Sommers JA, Brosh RM, Jr. In vivo function of the conserved non-catalytic domain of Werner syndrome helicase in DNA replication. Hum Mol Genet. 2004 Oct 1;13(19):2247-61. 15282207
190 - Szekely AM, Chen YH, Zhang C, Oshima J, Weissman SM. Werner protein recruits DNA polymerase delta to the nucleolus. Proc Natl Acad Sci U S A. 2000 Oct 10;97(21):11365-70. 11027336
191 - Kamath-Loeb AS, Loeb LA, Johansson E, Burgers PM, Fry M. Interactions between the Werner syndrome helicase and DNA polymerase delta specifically facilitate copying of tetraplex and hairpin structures of the d(CGG)n trinucleotide repeat sequence. J Biol Chem. 2001 May 11;276(19):16439-46. 11279038
192 - Fry M, Loeb LA. Human werner syndrome DNA helicase unwinds tetrahelical structures of the fragile X syndrome repeat sequence d(CGG)n. J Biol Chem. 1999 Apr 30;274(18):12797-802. 10212265
193 - Riha K, Heacock ML, Shippen DE. The role of the nonhomologous end-joining DNA double-strand break repair pathway in telomere biology. Annu Rev Genet. 2006;40:237-77. 16822175
194 - Cheng WH, von Kobbe C, Opresko PL, Arthur LM, Komatsu K, Seidman MM, Carney JP, Bohr VA. Linkage between Werner syndrome protein and the Mre11 complex via Nbs1. J Biol Chem. 2004 May 14;279(20):21169-76. 15026416
195 - Celli GB, Denchi EL, de Lange T. Ku70 stimulates fusion of dysfunctional telomeres yet protects chromosome ends from homologous recombination. Nat Cell Biol. 2006 Aug;8(8):885-90. 16845382
196 - Laud PR, Multani AS, Bailey SM, Wu L, Ma J, Kingsley C, Lebel M, Pathak S, DePinho RA, Chang S. Elevated telomere-telomere recombination in WRN-deficient, telomere dysfunctional cells promotes escape from senescence and engagement of the ALT pathway. Genes Dev. 2005 Nov 1;19(21):2560-70. 16264192
197 - Harrigan JA, Wilson DM, 3rd, Prasad R, Opresko PL, Beck G, May A, Wilson SH, Bohr VA. The Werner syndrome protein operates in base excision repair and cooperates with DNA polymerase beta. Nucleic Acids Res. 2006;34(2):745-54. 16449207
198 - Blank A, Bobola MS, Gold B, Varadarajan S, D DK, Meade EH, Rabinovitch PS, Loeb LA, Silber JR. The Werner syndrome protein confers resistance to the DNA lesions N3-methyladenine and O6-methylguanine: implications for WRN function. DNA Repair (Amst). 2004 Jun 3;3(6):629-38. 15135730
199 - Harrigan JA, Opresko PL, von Kobbe C, Kedar PS, Prasad R, Wilson SH, Bohr VA. The Werner syndrome protein stimulates DNA polymerase beta strand displacement synthesis via its helicase activity. J Biol Chem. 2003 Jun 20;278(25):22686-95. 12665521
200 - Li B, Navarro S, Kasahara N, Comai L. Identification and biochemical characterization of a Werner's syndrome protein complex with Ku70/80 and poly(ADP-ribose) polymerase-1. J Biol Chem. 2004 Apr 2;279(14):13659-67. 14734561
201 - von Kobbe C, Harrigan JA, Schreiber V, Stiegler P, Piotrowski J, Dawut L, Bohr VA. Poly(ADP-ribose) polymerase 1 regulates both the exonuclease and helicase activities of the Werner syndrome protein. Nucleic Acids Res. 2004;32(13):4003-14. 15292449
202 - Bryant HE, Schultz N, Thomas HD, Parker KM, Flower D, Lopez E, Kyle S, Meuth M, Curtin NJ, Helleday T. Specific killing of BRCA2-deficient tumours with inhibitors of poly(ADP-ribose) polymerase. Nature. 2005 Apr 14;434(7035):913-7. 15829966
203 - Farmer H, McCabe N, Lord CJ, Tutt AN, Johnson DA, Richardson TB, Santarosa M, Dillon KJ, Hickson I, Knights C, Martin NM, Jackson SP, Smith GC, Ashworth A. Targeting the DNA repair defect in BRCA mutant cells as a therapeutic strategy. Nature. 2005 Apr 14;434(7035):917-21. 15829967
204 - Bohr VA, Souza Pinto N, Nyaga SG, Dianov G, Kraemer K, Seidman MM, Brosh RM, Jr. DNA repair and mutagenesis in Werner syndrome. Environ Mol Mutagen. 2001;38(2-3):227-34. 11746759
205 - Yun J, Zhong Q, Kwak JY, Lee WH. Hypersensitivity of Brca1-deficient MEF to the DNA interstrand crosslinking agent mitomycin C is associated with defect in homologous recombination repair and aberrant S-phase arrest. Oncogene. 2005 Jun 9;24(25):4009-16. 15782115
206 - Cheng WH, Kusumoto R, Opresko PL, Sui X, Huang S, Nicolette ML, Paull TT, Campisi J, Seidman M, Bohr VA. Collaboration of Werner syndrome protein and BRCA1 in cellular responses to DNA interstrand cross-links. Nucleic Acids Res. 2006;34(9):2751-60. 16714450
207 - Mann MB, Hodges CA, Barnes E, Vogel H, Hassold TJ, Luo G. Defective sister-chromatid cohesion, aneuploidy and cancer predisposition in a mouse model of type II Rothmund-Thomson syndrome. Hum Mol Genet. 2005 Mar 15;14(6):813-25. 15703196
208 - Sangrithi MN, Bernal JA, Madine M, Philpott A, Lee J, Dunphy WG, Venkitaraman AR. Initiation of DNA replication requires the RECQL4 protein mutated in Rothmund-Thomson syndrome. Cell. 2005 Jun 17;121(6):887-98. 15960976
209 - Matsuno K, Kumano M, Kubota Y, Hashimoto Y, Takisawa H. The N-terminal noncatalytic region of Xenopus RecQ4 is required for chromatin binding of DNA polymerase alpha in the initiation of DNA replication. Mol Cell Biol. 2006 Jul;26(13):4843-52. 16782873
210 - Im JS, Ki SH, Farina A, Jung DS, Hurwitz J, Lee JK. Assembly of the Cdc45-Mcm2-7-GINS complex in human cells requires the Ctf4/And-1, RecQL4, and Mcm10 proteins. Proc Natl Acad Sci U S A. 2009 Sep 15;106(37):15628-32. 19805216
211 - Kumata Y, Tada S, Yamanada Y, Tsuyama T, Kobayashi T, Dong YP, Ikegami K, Murofushi H, Seki M, Enomoto T. Possible involvement of RecQL4 in the repair of double-strand DNA breaks in Xenopus egg extracts. Biochim Biophys Acta. 2007 Apr;1773(4):556-64. 17320201
212 - Petkovic M, Dietschy T, Freire R, Jiao R, Stagljar I. The human Rothmund-Thomson syndrome gene product, RECQL4, localizes to distinct nuclear foci that coincide with proteins involved in the maintenance of genome stability. J Cell Sci. 2005 Sep 15;118(Pt 18):4261-9. 16141230
213 - Woo LL, Futami K, Shimamoto A, Furuichi Y, Frank KM. The Rothmund-Thomson gene product RECQL4 localizes to the nucleolus in response to oxidative stress. Exp Cell Res. 2006 Oct 15;312(17):3443-57. 16949575
214 - Werner SR, Prahalad AK, Yang J, Hock JM. RECQL4-deficient cells are hypersensitive to oxidative stress/damage: Insights for osteosarcoma prevalence and heterogeneity in Rothmund-Thomson syndrome. Biochem Biophys Res Commun. 2006 Jun 23;345(1):403-9. 16678792
215 - Wang M, Wu W, Wu W, Rosidi B, Zhang L, Wang H, Iliakis G. PARP-1 and Ku compete for repair of DNA double strand breaks by distinct NHEJ pathways. Nucleic Acids Res. 2006;34(21):6170-82. 17088286
216 - Audebert M, Salles B, Calsou P. Involvement of poly(ADP-ribose) polymerase-1 and XRCC1/DNA ligase III in an alternative route for DNA double-strand breaks rejoining. J Biol Chem. 2004 Dec 31;279(53):55117-26. 15498778
217 - Bailis JM, Luche DD, Hunter T, Forsburg SL. Minichromosome maintenance proteins interact with checkpoint and recombination proteins to promote s-phase genome stability. Mol Cell Biol. 2008 Mar;28(5):1724-38. 18180284
218 - Gambus A, Jones RC, Sanchez-Diaz A, Kanemaki M, van Deursen F, Edmondson RD, Labib K. GINS maintains association of Cdc45 with MCM in replisome progression complexes at eukaryotic DNA replication forks. Nat Cell Biol. 2006 Apr;8(4):358-66. 16531994
219 - Katou Y, Kanoh Y, Bando M, Noguchi H, Tanaka H, Ashikari T, Sugimoto K, Shirahige K. S-phase checkpoint proteins Tof1 and Mrc1 form a stable replication-pausing complex. Nature. 2003 Aug 28;424(6952):1078-83. 12944972
220 - Calzada A, Hodgson B, Kanemaki M, Bueno A, Labib K. Molecular anatomy and regulation of a stable replisome at a paused eukaryotic DNA replication fork. Genes Dev. 2005 Aug 15;19(16):1905-19. 16103218
221 - Komata M, Bando M, Araki H, Shirahige K. The direct binding of Mrc1, a checkpoint mediator, to Mcm6, a replication helicase, is essential for the replication checkpoint against methyl methanesulfonate-induced stress. Mol Cell Biol. 2009 Sep;29(18):5008-19. 19620285
222 - Nedelcheva MN, Roguev A, Dolapchiev LB, Shevchenko A, Taskov HB, Shevchenko A, Stewart AF, Stoynov SS. Uncoupling of unwinding from DNA synthesis implies regulation of MCM helicase by Tof1/Mrc1/Csm3 checkpoint complex. J Mol Biol. 2005 Apr 1;347(3):509-21. 15755447
223 - Lou H, Komata M, Katou Y, Guan Z, Reis CC, Budd M, Shirahige K, Campbell JL. Mrc1 and DNA polymerase epsilon function together in linking DNA replication and the S phase checkpoint. Mol Cell. 2008 Oct 10;32(1):106-17. 18851837
224 - Szyjka SJ, Viggiani CJ, Aparicio OM. Mrc1 is required for normal progression of replication forks throughout chromatin in S. cerevisiae. Mol Cell. 2005 Sep 2;19(5):691-7. 16137624
225 - Alcasabas AA, Osborn AJ, Bachant J, Hu F, Werler PJ, Bousset K, Furuya K, Diffley JF, Carr AM, Elledge SJ. Mrc1 transduces signals of DNA replication stress to activate Rad53. Nat Cell Biol. 2001 Nov;3(11):958-65. 11715016
226 - Bjergbaek L, Cobb JA, Tsai-Pflugfelder M, Gasser SM. Mechanistically distinct roles for Sgs1p in checkpoint activation and replication fork maintenance. Embo J. 2005 Jan 26;24(2):405-17. 15616582
227 - Bando M, Katou Y, Komata M, Tanaka H, Itoh T, Sutani T, Shirahige K. Csm3, Tof1, and Mrc1 form a heterotrimeric mediator complex that associates with DNA replication forks. J Biol Chem. 2009 Dec 4;284(49):34355-65. 19819872
228 - Pryce DW, Ramayah S, Jaendling A, McFarlane RJ. Recombination at DNA replication fork barriers is not universal and is differentially regulated by Swi1. Proc Natl Acad Sci U S A. 2009 Mar 24;106(12):4770-5. 19273851