Defects in homologous recombination repair behind the human diseases: FA and HBOC

in Endocrine-Related Cancer

Hereditary breast and ovarian cancer (HBOC) syndrome and a rare childhood disorder Fanconi anemia (FA) are caused by homologous recombination (HR) defects, and some of the causative genes overlap. Recent studies in this field have led to the exciting development of PARP inhibitors as novel cancer therapeutics and have clarified important mechanisms underlying genome instability and tumor suppression in HR-defective disorders. In this review, we provide an overview of the basic molecular mechanisms governing HR and DNA crosslink repair, highlighting BRCA2, and the intriguing relationship between HBOC and FA.

Abstract

Hereditary breast and ovarian cancer (HBOC) syndrome and a rare childhood disorder Fanconi anemia (FA) are caused by homologous recombination (HR) defects, and some of the causative genes overlap. Recent studies in this field have led to the exciting development of PARP inhibitors as novel cancer therapeutics and have clarified important mechanisms underlying genome instability and tumor suppression in HR-defective disorders. In this review, we provide an overview of the basic molecular mechanisms governing HR and DNA crosslink repair, highlighting BRCA2, and the intriguing relationship between HBOC and FA.

Introduction

Breast or ovarian cancer is common, and one of the leading causes of mortality in women worldwide (Ferlay et al. 2015, Torre et al. 2015). Although hereditary forms of breast and ovarian cancer (HBOC) are estimated to account for only 5–20% of all cases of breast or ovarian cancer (Nielsen & van Overeem Hansen 2016), HBOC has been recently attracting considerable public attention in many parts of the world, including Japan. A fraction (~25%) of HBOC cases are caused by monoallelic, that is dominant, mutations in either the BRCA1 or BRCA2 genes (Nielsen & van Overeem Hansen 2016), the well-known tumor suppressors that were identified by linkage analysis in the mid-1990s (Miki et al. 1994, Wooster et al. 1995). The lifetime risk of developing breast or ovarian cancer can be up to ~80% in these affected individuals. Mutations in BRCA1/2 also predispose individuals to cancer in other organs such as prostate or pancreas. The BRCA genes are considered to be ‘caretakers’, and they function in homologous recombination (HR) repair, thereby protecting our genome from carcinogenic alterations. Furthermore, cancer genome sequences have revealed an unexpectedly high frequency of HR gene mutations in sporadic cases of high-grade serous ovarian cancer (Cancer Genome Atlas Research Network 2011), highlighting an important role of HR in cancer prevention via genome maintenance.

As the loss of HR activities by the biallelic disruption of either the Brca1 or Brca2 genes in mice results in early embryonic lethality (for example see Ludwig et al. 1997), it was a real surprise to the scientific community that the D’Andrea lab at Harvard University discovered biallelic mutations in BRCA2 in a subset of patients with the rare childhood hematological disorder, Fanconi anemia (FA) (FA-D1 subgroup) in 2002 (Howlett et al. 2002). The combined effects of these biallelic BRCA2/FANCD1 mutations appear to be hypomorphic and somehow compatible with life.

FA is very rare, but still the most prevalent among the inherited forms of bone marrow failure syndrome (Auerbach 2009). It is primarily an autosomal recessive disorder that is clinically characterized by congenital malformations, progressive development of hypoplastic anemia and cancer predisposition that often results in hematological malignancies such as acute myelogenous leukemia (AML) or myelodysplasia (MDS) as well as various solid tumors, especially head and neck squamous carcinoma (Alter 2014). FA was first described in 1927 by the Swiss pediatrician Guido Fanconi (Lobitz & Velleuer 2006). Traute Schroeder and coworkers reported spontaneous chromosomal breakage in FA in 1964 (Schroeder et al. 1964). Ten years later, Sasaki and Tonomura discovered that FA cells are extremely sensitive to ICL-inducing agents such as mitomycin C (MMC) (Sasaki & Tonomura 1973), resulting from defective interstrand crosslink (ICL) repair. It is now generally accepted that an ICL is repaired through consecutive steps of multiple DNA repair activities including the HR mechanism (Fig. 1) (Duxin & Walter 2015).

Figure 1

Download Figure

Figure 1

An overview of the ICL repair pathway. When a replisome collides with an ICL (I), the leading strand initially stalls −20 nucleotides away from the lesion (II). After a second fork converges at the ICL, BRCA1 facilitates the dissociation of the CMG complex (consisting of Cdc45, MCM2-7 and GINS) from chromatin at the stalled fork, allowing the leading strand to approach to the −1 nucleotide adjacent to the ICL (III). In the next step, the SLX4-XPF-ERCC1 complex incises the DNA and unhooks the lesion in a FA pathway-dependent manner (IV). Translesion synthesis (TLS) polymerase (Polζ, or possibly Polκ, Polη, or Polι) extends the leading strand synthesis past the unhooked, ICL-associated nucleotide (V), and HR and the nucleotide excision repair (NER) pathway repair the remaining lesion (VI).

Citation: Endocrine-Related Cancer 23, 10; 10.1530/ERC-16-0221

More recently, it has been recognized that mutation carriers of some of the FA genes (e.g., parents of the FA patients) may actually develop HBOC and that the other HBOC genes (i.e., BRCA1 (Domchek et al. 2013, Sawyer et al. 2015)) can cause an FA-like disorder when biallelically mutated (Bogliolo & Surrallés 2015). Thus, HBOC and the FA genes do overlap to some extent (Tables 1 and 2). In a simplified view, it could be said that near-total loss of (or hypomorphic) HR repair activities causes the FA phenotype (or sometimes FA-like, see the ‘The core HR genes in the FA pathway’ section below), whereas breast and ovarian cancer without constitutive symptoms (i.e., HBOC) is caused when the HR repair activities are disabled to certain levels. However, HR is severely disabled in the HBOC tumors, by loss of heterozygosity (LOH), promoter methylation or other mechanisms. In this review, we provide a brief overview of the current understanding of the molecular mechanism of HR and ICL repairs, highlighting BRCA2, and their relationship with two important human diseases, HBOC and FA.

Table 1

Hereditary breast and ovarian cancer (HBOC) risk genes.

GeneSyndrome with germline mutationsFunctionsCancer typePenetrance
BRCA1HBOCHomologous recombinationBreast and ovarian cancerHigh
BRCA2HBOCHomologous recombinationBreast and ovarian cancerHigh
PTENCowden syndrome, PTEN hamartomaPhosphatidylinositol 3-phosphate, suppresses AKT signalingBreast cancerHigh
TP53Li–Fraumeni syndromeTranscription factor, regulates cell cycle, apoptosis, senescenceBreast and ovarian cancerHigh
CDH1Hereditary diffuse gastric cancer syndromeE-cadherin gene, maintains cell adherenceBreast and ovarian cancerHigh
STK11Peutz–Jeghers syndromeSerine/threonine kinase, regulates cell polarityBreast and ovarian cancerHigh
NBS1Nijmegen breakage syndromeCell cycle checkpoint after DNA damage, member of the MRN complexBreast cancerHigh
NF1Neurofibromatosis type INegative regulator of Ras signalingBreast cancerHigh
ATMAtaxia telangiectasiaPI3 kinase-related kinase, cell cycle checkpoint and DSB repairBreast cancerModerate
CHK2Li–Fraumeni syndromeActivation of cell cycle checkpoint after DNA damageBreast and ovarian cancerModerate
FANCJFanconi anemiaInterstrand crosslink repairBreast and ovarian cancerModerate
FANCMFanconi anemiaInterstrand crosslink repairBreast cancerModerate
PALB2Fanconi anemiaInterstrand crosslink repair, homologous recombinationBreast and ovarian cancerModerate
RAD51CFA-like syndromeInterstrand crosslink repair, homologous recombinationBreast and ovarian cancerModerate
Table 2

Fanconi anemia (FA) and FA-like syndrome genes. There are already 20 distinct genes identified in these syndromes, and all proteins are required for interstrand crosslink (ICL) repair. Heterozygous germline mutations in several genes are also related to HBOC.

GeneSynonymFunctionsSymptomsHeterozygous germline mutation
FANCAComponent of the FA core complexFA pathologies
FANCBComponent of the FA core complexFA pathologies
FANCCComponent of the FA core complexFA pathologies
FANCD1BRCA2HR repair, recruits RAD51 onto DNA, interacts with FANCN, Stalled replication fork protectionFA pathologies, not all patients show bone marrow failureHBOC
FANCD2Ubiquitinated after DNA damage, Stalled replication fork protectionFA pathologies
FANCEComponent of the FA core complexFA pathologies
FANCFComponent of the FA core complexFA pathologies
FANCGXRCC9Component of the FA core complexFA pathologies
FANCIUbiquitinated after DNA damage, required for FA core complex activationFA pathologies
FANCJBACH1, BRIP1ICL repair, HR repair, 3′ to 5′ helicase, interacts with BRCA1FA pathologiesHBOC
FANCLPHF9Component of the FA core complex, E3 ubiquitin ligaseFA pathologies but no cancers
FANCMHefDNA translocase, required for FANCI-D2 ubiquitinationUnknown, the only known patient also has a FANCA mutationHBOC
FANCNPALB2HR repair, interacts with BRCA1 and BRCA2, facilitates BRCA2 functionFA pathologiesHBOC
FANCORAD51CRAD51 paralog, HR repair, RAD51 nucleoprotein filament stabilityFA-like syndrome, no bone marrow failure and cancer
FANCPSLX4Coordinates XPF-ERCC1, interacts with MUS81-EME1 and SLX1 nucleasesFA pathologies
FANCQXPF, ERCC4Endonucleases, associates with ERCC1, ICL unhookingFA pathologiesHBOC
FANCRRAD51HR repair, stalled fork protectionFA-like syndrome, no bone marrow failure and cancer
FANCSBRCA1HR repair, promotes RAD51 recruitment, interacts with FANCNFA-like syndrome, no bone marrow failureHBOC
FANCTUBE2TE2 ubiquitin-conjugating enzyme for FANCD2 complex, interacts with FANCLFA pathologies
FANCUXRCC2RAD51 paralog, HR repair, RAD51 nucleoprotein filament stabilityFA-like syndrome, no bone marrow failure

Genome integrity is maintained by DNA repair

Each day, every cell incurs a large number of DNA lesions that threaten the integrity of the genome. These lesions originate either from an exogenous source (e.g., X-rays or ultraviolet light) or are created endogenously by metabolic pathways (i.e., free radicals or aldehydes) or by programmed cellular activities (i.e., VDJ recombination in developing lymphocytes or meiotic recombination in germ cells) (Hoeijmakers 2001). Alternatively, it is known that replication stress provoked by oncogene activation or fork collision with transcription machineries can induce stalled replication forks that may result in fork collapse in genomic regions such as common fragile sites (Debatisse et al. 2012).

A DNA double-strand break (DSB) is among the most severe insults to the genome, and it can be repaired by two basic mechanisms termed HR or nonhomologous end joining (NHEJ). HR and NHEJ function in a cooperative and overlapping manner, or perhaps paradoxically, can compete with each other (Takata et al. 1998, Prakash et al. 2015). Although NHEJ can function throughout the cell cycle, the HR pathway only functions during S/G2 phase. In essence, NHEJ unites two DNA ends by ligation without any requirement of homology, often after processing (i.e., removal or addition of short stretches of nucleotides) of the ends. Therefore, the repair process can be error-prone. On the other hand, HR functions by a ‘copy-and-paste’ mechanism of genetic information transfer from an intact homologous template to the damaged DNA, and therefore it occurs without sequence alteration. HR normally requires replicated DNA (sister chromatid) as a template, and this is one of the reasons for HR to be restricted to the S and G2 phases of the cell cycle.

In addition to their role in DSB repair, HR proteins have an important role during S phase, where they function in restarting stalled replication forks or protecting stalled replication forks from collapsing due to nucleolytic digestion (Hashimoto et al. 2012, Schlacher et al. 2012). For example, loss of Rad51 in mice (Tsuzuki et al. 1996) or chicken DT40 cells (Sonoda et al. 1998) causes cell lethality, which is accompanied by chromosomal breaks likely due to replication fork collapse. HR is also an integral step during ICL repair, as illustrated in Fig. 1.

Basic molecular mechanisms of HR repair

The mechanisms of HR can be best explained in the context of DSB repair (Fig. 2). To initiate DSB repair through HR, the DSB end needs to be nucleolytically resected to generate 3′ single-stranded DNA (ssDNA). This is quickly coated by the trimeric ssDNA-binding protein complex – replication protein A (RPA). The RPA complex is displaced by RAD51, which is the central player in mammalian HR repair, resulting in the formation of RAD51 nucleoprotein filaments. This reaction is facilitated by mediator proteins including BRCA2 and RAD51 paralogs and can be monitored as the formation of subnuclear small dots (RAD51 foci) by immunohistochemical detection.

Figure 2

Download Figure

Figure 2

Schematic of HR pathway. When a double-strand break (DSB) is generated after DNA replication during S and G2 phase, both strands are resected in the 5′ to 3′ direction to generate 3′ overhangs. Almost immediately, replication protein A (RPA) is loaded onto the single-stranded (ss) DNA, and then replaced by a RAD51 nucleoprotein filament in a process requiring BRCA1–PALB2–BRCA2. RAD51 carries out strand invasion of the sister chromatid by the ssDNA tail and extends the resulting D-loop formation. In synthesis-dependent strand annealing (SDSA), the D-loop structure quickly dissociates from the ssDNA after synthesis of a complementary single strand, and then another strand anneals with a processed ssDNA. An alternate pathway forms a double Holliday junction (dHJ). After second end capture and fill-in synthesis, the Holliday junction is dissociated by the TopIIIα–BLM complex or resolved by resolvase complexes that contain SLX4, MUS81 and GEN1.

Citation: Endocrine-Related Cancer 23, 10; 10.1530/ERC-16-0221

RAD51 is a homolog of Escherichia coli RecA, which mediates the core enzymatic reactions in HR (West 2003). It catalyzes (1) searching for a homologous HR template (homology search) and then (2) pairing of the ssDNA-RAD51 filament with the template DNA (strand invasion and homologous pairing) once the filament encounters the appropriate homologous double-stranded DNA. These reactions result in the formation of a D-loop that consists of heteroduplex DNA coated with RAD51 and displaced ssDNA. The next step is DNA repair synthesis initiated from the invading 3′ ssDNA end. In most instances in mitotic cells, the extended ssDNA is displaced from the template strand and is then annealed by the other processed single-stranded DNA end (i.e., the other end of the DSB). This mechanism is termed ‘synthesis-dependent strand annealing (SDSA) pathway’, and the final product does not contain crossover events (non-crossover). In some cases, the recombination intermediates are converted into a ‘double Holliday junction’ and subsequently resolved by Holliday junction resolvases (e.g., GEN1 or the SLX4 complex) (Garner et al. 2013, Wyatt et al. 2013) (with or without crossover) or the BLM helicase complex (without crossover) (Wu & Hickson 2003). For more complete discussion of HR mechanisms such as their relationship with competing NHEJ (the pathway choice) or Holliday junction resolvases, readers should refer to the recent excellent reviews (Chapman et al. 2012, Sarbajna & West 2014).

Basic molecular mechanisms of ICL repair

An ICL covalently bridges two nucleotides on opposite DNA strands and hampers critical DNA transactions such as DNA replication and transcription. Recent studies from Walter’s lab using Xenopus egg extracts and plasmid DNA harboring an ICL provided a comprehensive and persuasive view on the detailed mechanisms of ICL repair (Fig. 1) (Räschle et al. 2008, Knipscheer et al. 2009, Long et al. 2011, Duxin & Walter 2015).

According to this model, two converging replication forks from opposite directions first stall ~20 bp away (−20) from the ICL, and then leading strand synthesis progresses to the −1 position with respect to the ICL, perhaps after the removal of the CMG helicase. Next, the ICL and ssDNA regions are recognized, leading to the activation of the checkpoint kinase and the FA pathway. The exact mechanism by which these events are accomplished still remains unclear. DNA strands on both sides of the ICL are incised by recruited nucleases, resulting in DSB formation in one of the sister chromatids (unhooking). The DNA replication occurs over the incised ICL by a bypass DNA polymerase specialized in translesion synthesis (TLS), such as REV1 or REV3, perhaps after PCNA monoubiquitination by the RAD18 ubiquitin ligase. The DSB is resected, and the core HR reaction is initiated with the formation of RAD51 filaments. Finally, the short nucleotide fragment that contains the remnant of the ICL is recognized and removed by nucleotide excision repair (NER). How each component of the FA pathway is involved in the ICL repair is described in the following section.

FA, FA pathway and ICL repair

So far, nineteen ‘FA genes’ have been identified and are thought to function in the ICL repair pathway (Kitao & Takata 2011, Kottemann & Smogorzewska 2013, Bogliolo & Surrallés 2015, Ceccaldi et al. 2016). These FA pathway genes are classified into three subgroups by their functional roles in the ICL repair as explained below (Fig. 3).

Figure 3

Download Figure

Figure 3

The mechanism of FA pathway activation. Upon replication fork stalling, FANCM–FAAP24–MHF1/2 complex binding at the ICL lesion activates ATR signaling, followed by the recruitment of the FA core complex. FANCI is phosphorylated by ATR, and subsequently, the FA core complex with UBE2T mediates the monoubiquitination of the FANCI–FANCD2 (D2-I) complex. This modification targets the D2-I complex to chromatin and leads to multiple events that repair the ICL lesion.

Citation: Endocrine-Related Cancer 23, 10; 10.1530/ERC-16-0221

The FA core complex and the key downstream complex consisting of FANCD2 and FANCI

The first FA group is the ‘ubiquitination module,’ which comprises the E3 ubiquitin ligase complex (termed FA core complex) and its substrates, the FANCD2–FANCI (D2-I) complex (Fig. 3). These genes have been shown to function in HR, mainly based on Jasin’s recombination assay. In this assay, a chromosomal DSB is induced within an integrated recombination substrate by a plasmid-encoded rare restriction enzyme I-SceI, which recognizes a specific 18 bp sequence (Rouet et al. 1994). The HR repair pathway then uses homologous DNA segments placed either upstream or downstream of the DSB, resulting in the expression of the neomycin resistance gene or GFP. In cells lacking these FA genes, the efficiency of HR repair is substantially decreased in chicken DT40 cells (Yamamoto et al. 2003, 2004) or mildly decreased in human cells (Nakanishi et al. 2005). How these proteins function in HR is still under investigation; however, milder HR defects in human cells may indicate that these ‘ubiquitination module’ FA genes do not provide ‘core’ HR functions. It seems more likely that they modulate the function of the core HR machineries (such as BRCA2 or CtIP, see the ‘The nucleases in the FA pathway’ section below) or TLS polymerase (Kim et al. 2012) or they may regulate histone dynamics (Sato et al. 2012). This module is reported to be required for incision/unhooking of the crosslink during ICL repair (Knipscheer et al. 2009, Klein Douwel et al. 2014, Duxin & Walter 2015), and thus functions in the conversion of an ICL to a DSB.

The FA core complex includes FA proteins FANCA, B, C, E, F, G, J, L, M and FA-associated proteins, such as FAAP24, FAAP20 and FAAP100 (Kottemann & Smogorzewska 2013, Bogliolo & Surrallés 2015, Ceccaldi et al. 2016). In response to an ICL and/or a stalled replication fork, the FA core complex is somehow activated downstream of the checkpoint kinase ATR-ATRIP through multiple phosphorylation of FANCI (Ishiai et al. 2008), FANCM (Singh et al. 2013) or FANCA (Collins et al. 2009) and monoubiquitinates FANCD2 at lysine 561, which is a critical activating event in the FA pathway (Garcia-Higuera et al. 2001, Matsushita et al. 2005). Recent studies indicate that ubiquitin-like with PHD and RING finger domain 1 (UHRF1) protein functions as an ICL recognition factor and may participate in these steps (Liang et al. 2015, Tian et al. 2015). Recently, we and two different groups identified UBE2T, which encodes an E2 ubiquitin-conjugating enzyme, as a causative gene for FA (Hira et al. 2015, Rickman et al. 2015, Virts et al. 2015). UBE2T/FANCT is essential for this monoubiquitination event to proceed. FANCI is a paralog of FANCD2 and its binding partner and also undergoes monoubiquitination dependent on the core complex and monoubiquitination of FANCD2 (Smogorzewska et al. 2007, Ishiai et al. 2008). The monoubiquitinated D2-I complex (often referred to as the ‘ID complex’) is recruited and accumulates in foci at damaged chromatin, perhaps by binding to the stalled fork itself (with or without an ICL) (Joo et al. 2011, Liang et al. 2016). Focus formation is likely to function as a platform to recruit the numerous proteins required for homologous recombination (HR) and translesion synthesis (TLS) to the damage site. However, the mechanism by which it orchestrates the repair machinery is still not entirely clear.

FANCM is a human homolog of the archeal helicase/nuclease Hef gene (Komori et al. 2004), which encodes a DNA translocase (Meetei et al. 2005). It is necessary for chromatin loading of the FA core complex (Kim et al. 2008) and checkpoint activation (Huang et al. 2010), and it plays a distinct role in bypass replication past an ICL by promoting ‘traverse’ of the lesion (Huang et al. 2013). Biallelic FANCM mutations were identified in a single patient, but the defects in cells from the patient could not be reversed by the expression of wild-type FANCM (Meetei et al. 2005). This was later found to be due to the presence of simultaneous mutations in FANCA (Singh et al. 2009). Thus, to date, there have been no human FA patients identified with causative mutations solely in FANCM. Furthermore, in the Finnish population, individuals with homozygous loss-of-function FANCMmutations do not show any FA phenotype (Lim et al. 2014), suggesting that FANCM is not a bona fide FA gene, though it clearly contributes to the function of the FA pathway and is a candidate HBOC gene (Kiiski et al. 2014, Peterlongo et al. 2015).

As FANCD2 and FANCI form a dimeric complex that seems quite stable (Sato et al. 2012), and they are mutually dependent on each other for activating monoubiquitination (Smogorzewska et al. 2007, Ishiai et al. 2008), it has been assumed that these proteins should function together. However, it was discovered recently that FANCI, not FANCD2, has an upstream role for foci formation of the core complex components, like FANCA (Castellà et al. 2015). It would be interesting to determine which molecule (FANCM vs FANCI) is furthest upstream in localizing the core complex. Further, ATR-phosphorylated FANCI regulates the replicative helicase MCM complex, thereby suppressing dormant origin firing as a distinct function outside of the FA pathway (Chen et al. 2015). FANCD2 was also found to interact with MCM helicase independently of its monoubiquitination, where it restrains DNA synthesis in stressed cells, attenuating cell proliferation and carcinogenesis (Lossaint et al. 2013).

The nucleases in the FA pathway

One of the essential events in ICL repair is an incision of the crosslink, which is called ‘unhooking’. In the current understanding, this event depends on the monoubiquitinated D2-I complex and is carried out by the nuclease complex SLX4-XPF (Knipscheer et al. 2009, Long et al. 2011, Kim et al. 2013, Hodskinson et al. 2014, Klein Douwel et al. 2014, Duxin & Walter 2015). SLX4/FANCP is a large protein that itself can function as a scaffold and is mutated in FA-P patients (Kottemann & Smogorzewska 2013, Bogliolo & Surrallés 2015, Ceccaldi et al. 2016). SLX4 accumulates in the chromatin containing DNA damage via its tandem UBZ4 domains. It has been reported that SLX4 is recruited by monoubiquitinated FANCD2 (Yamamoto et al. 2011). However, there is a conflicting report (Lachaud et al. 2014), and the UBZ domain is generally considered to bind to K63-linked polyubiquitin (Lachaud et al. 2014). How SLX4 is tethered to the sites of damage and how the D2-I complex affects unhooking are important issues that need to be resolved.

Of note, biallelic mutations in XPF that specifically affect the cellular sensitivity to ICLs but not UV were identified among unclassified FA patients (Bogliolo et al. 2013, Kashiyama et al. 2013). Now this group of patients is termed FA-Q. XPF was originally identified as one of the causative genes for a UV-sensitive disorder – xeroderma pigmentosum (XP). This is an interesting example of distinct phenotypes due to specific mutations affecting different features of a single protein.

Another nuclease FAN1, which associates with monoubiquitinated FANCD2, was also thought to function for ICL repair in an FA pathway-dependent manner. However, quite recently, Lachaud and coworkers demonstrated that FANCD2 monoubiquitination-dependent FAN1 recruitment is dispensable for ICL repair function of FAN1 but is required for DNA replication fork progression and the prevention of chromosome abnormalities (Lachaud et al. 2016).

CtIP is an important nuclease required for end resection of DSBs and has been shown to interact with BRCA1 as well as the MRN complex. We and others have identified CtIP as a novel interactor of FANCD2 (Murina et al. 2014, Unno et al. 2014). MMC-induced CtIP recruitment to damage foci is dependent on the interaction with FANCD2, and this recruitment appears to be required for end resection of the DSB generated after unhooking of the ICL. CtIP depletion mildly sensitizes cells to MMC treatment, consistent with the role of CtIP downstream of FANCD2 (Murina et al. 2014, Unno et al. 2014).

The core HR genes in the FA pathway

The group of HR/FA genes that operate in the repair ICLs includes BRCA2/FANCD1, Brip1/FANCJ, PALB2/FANCN, RAD51C/FANCO and XRCC2/FANCU (Kottemann & Smogorzewska 2013, Bogliolo & Surrallés 2015, Ceccaldi et al. 2016). BRCA1/FANCS (Domchek et al. 2013, Sawyer et al. 2015) and RAD51/FANCR (Ameziane et al. 2015, Wang et al. 2015) genes are recent and surprising additions to this group. Many of these are HBOC genes (Table 1) and RAD51 mediators. Thus, they function as the core HR machinery.

BRCA2/FANCD1 was identified as the first core HR gene implicated in FA (Howlett et al. 2002). As it is a well-known tumor suppressor and HBOC gene, this led to the exciting possibility that mutations in other FA genes would also cause HBOC. However, this prediction turned out to be a bit too simplistic. Genes encoding the core complex components and FANCD2/FANCI are unlikely to be a high-penetrance HBOC gene (Seal et al. 2003, Berwick et al. 2007), although there are some reports indicating FANCM (Kiiski et al. 2014, Peterlongo et al. 2015) or FANCC(Berwick et al. 2007, Thompson et al. 2012) could be considered as candidate HBOC genes.

PALB2/FANCNis the partner and localizer of BRCA2 (Xia et al. 2006), and also binds BRCA1, linking the two critical HR proteins BRCA1 and BRCA2 (Sy et al. 2009, Zhang et al. 2009). Interaction with BRCA1 facilitates the recruitment of PALB2–BRCA2 complex to the DNA damage sites. PALB2 stabilizes BRCA2 and critically regulates the functions of BRCA2 as a mediator for RAD51. A recent study revealed that PALB2–BRCA1 interaction is a regulatory point during cell cycle progression (see the ‘Additional regulators of HR and RAD51 function’ section below) (Orthwein et al. 2015). Strikingly, FA-D1 and FA-N patients develop leukemia and kidney or brain tumors at a very early age, with much higher frequency than other FA complementation groups (Hirsch et al. 2004, Wagner et al. 2004, Reid et al. 2007).

Five RAD51 paralogs (e.g., RAD51B, RAD51C, RAD51D, XRCC2, XRCC3) interact in two distinct complexes (Masson et al. 2001) – RAD51B/C/D/XRCC2 (BCDX2) and RAD51C/XRCC3 (CX3). In addition to BRCA2, they also function as RAD51 mediators. It was confirmed that these two complexes are functionally different from each other (Yonetani et al. 2005). Furthermore, analysis of the C. elegans Rad51 homolog revealed that RAD51 paralogs remodel presynaptic filaments of RAD51 into a stabilized and flexible conformation, which prevents ssDNA degradation by nucleases and RAD51 dissociation (Taylor et al. 2015). Among these paralog genes, RAD51C or XRCC2 mutations were reported in FA-like patients with physical characteristics and chromosome breakage test results similar to FA. Thus, these patients are classified as FA-O (Vaz et al. 2010) or FA-U group (Shamseldin et al. 2012). Cells lacking RAD51 paralog genes generally exhibit a similar phenotype and deficiency in HR (Takata et al. 2000, 2001); therefore, it is possible that humans defective in any of the paralog genes may display a similar FA-like phenotype.

Recently, biallelic mutations in the BRCA1 gene have finally been identified in two patients with early-onset ovarian or breast cancer (Domchek et al. 2013, Sawyer et al. 2015). Diepoxybutane (DEB) induced chromosome breakage test was performed in one of them and found to be positive (Domchek et al. 2013, Sawyer et al. 2015), leading to the designation of FANCS. BRCA1/FANCS is known to target BRCA2/FANCD1 to the DSB site via interaction with PALB2/FANCN (Zhang et al. 2009). It has been reported that one of the BRCA1 interactors, Brip1 helicase, is responsible for the FA-J subgroup. These patients display a typical FA phenotype, albeit not a particularly severe one (Kitao & Takata 2011, Kottemann & Smogorzewska 2013, Bogliolo & Surrallés 2015, Ceccaldi et al. 2016). It was also recently discovered that a monoallelic mutation in RAD51 can give rise to FA-like symptoms by a dominant negative mechanism (termed FA-R subgroup) (Ameziane et al. 2015, Wang et al. 2015).

Interestingly, although FA-D1/N/J patients display the usual constellation of FA symptoms, the patients belonging to FA-O/R/S subgroups do not appear to develop BMF (Bogliolo & Surrallés 2015). Although it is possible that they eventually develop BMF in long-term follow-up, it seems inappropriate to classify these patients as having FA, and at the moment, they should be called FA-like. It will be highly interesting to clarify why these patients do not (or tend not to) develop hematopoietic stem cell failure because patients belonging to FA-D1/N/J/O/R/S all seem to have a similar pathophysiology due to HR deficiency. Of note, a recent study demonstrated that BRCA1 deficiency, specifically in mouse bone marrow, causes hematopoietic defects (Vasanthakumar et al. 2016).

Structure and function of BRCA2 in the HR pathway

General features

The BRCA2 primary structure is depicted in Fig. 4. BRCA2 is a huge protein that encompasses 3418 amino acids in humans. Shahid and coworkers recently revealed the structure of full-length BRCA2 as being a dimer, using cryo-EM 3D reconstruction (Shahid et al. 2014). The sequence conservation among BRCA2 orthologs among various species is mostly limited to the N-terminal, the middle part, and the C-terminal regions; therefore, these regions may be more important for genome maintenance (Takata et al. 2002).

Figure 4

Download Figure

Figure 4

The primary structures of human BRCA1 and BRCA2. BRCA1 consists of 1863 amino acids. The RING domain is in the N-terminus and partly overlaps with the BARD1-binding region. BRCA1 interacts with PALB2 via a coiled-coil region in the C-terminus. The BRCT motif is a phosphorylated protein-binding sequence, and this motif mediates the association of BRCA1 with Abraxas, CtIP and BACH1, which are all known factors in DSB repair. BRCA2 comprises 3418 amino acids. The PALB2 binding region is located in the amino (N) terminus. In the center, BRCA2 harbors eight BRC repeats, which constitute a binding region for RAD51 monomers. In the carboxy (C) terminus, a DNA-binding domain contains three OB folds and a region that interacts with DSS1. The C-terminal region of BRCA2 is also important for the formation of the RAD51 nucleoprotein filament and BRCA2 nuclear localization.

Citation: Endocrine-Related Cancer 23, 10; 10.1530/ERC-16-0221

N-terminus

In the BRCA2 N-terminus, there is a region that binds to PALB2/FANCN (Buisson et al. 2010, Menzel et al. 2011). PALB2 facilitates BRCA2 localization and RAD51 chromatin loading at the damage site. This region of BRCA2 was also reported to bind to EMSY (Hughes-Davies et al. 2003). The EMSY-binding region of BRCA2 is encoded by exon 3, which is known to be deleted in cancer (Hughes-Davies et al. 2003, Cousineau & Belmaaza 2011). The EMSY locus is amplified in sporadic breast cancer (13%) and higher-grade ovarian cancer (17%). At the cellular level, EMSY overexpression leads to defective HR (Cousineau & Belmaaza 2011), consistent with the notion that the EMSY-binding region of BRCA2 is likely to coincide or overlap with the PALB2-interacting region.

BRC repeats

In the middle part (the residues between 990 and 2100), BRCA2 harbors eight tandem BRC repeats, each consisting of about 30 amino acids, which are well conserved across human, mouse, rat and chicken (Takata et al. 2002, Yang et al. 2002). Not only the sequences themselves but also the spacing between them is well conserved across species. These individual repeats are the primary motifs through which BRCA2 binds to RAD51, and they are essential for BRCA2 function in HR as shown by mouse knockout studies (Connor et al. 1997, Ludwig et al. 1997, Friedman et al. 1998, Patel et al. 1998, Jonkers et al. 2001). BRCA2 stimulates RAD51 assembly onto ssDNA, and the BRC repeats are critical for this, perhaps by acting cooperatively. It is still unclear whether each BRC repeat can have a distinct function or they can act in a redundant manner. Supporting the former possibility, BRC missense mutations disrupting the interaction with RAD51 have been identified in breast cancer patients (Pellegrini et al. 2002), and conservation between the repeats in a given species is relatively low. It was also shown that BRC1-4 has a higher affinity to RAD51 monomers than BRC5-8; therefore, BRC repeats may not be functionally equivalent (Carreira & Kowalczykowski 2011). On the other hand, it was shown that an artificial fusion gene consisting of a single BRC repeat and RPA can display HR function (Saeki et al. 2006).

DBD

In the C-terminus of BRCA2, there is a DNA-binding domain (DBD) containing three oligonucleotide-binding (OB) folds, a Tower domain and a helix-turn-helix (HTH) motif. The DBD interacts not only with single-stranded DNA but also with DSS1 (Yang et al. 2002), which is a small 70 amino acid protein identified from the genomic region on chromosome 7q21.3 that was deleted in an inherited developmental syndrome – split hand/foot malformation. Recently, DSS1 has been reported to target BRCA2 to RPA (Zhao et al. 2015), and it functions in the replacement of RPA with RAD51 on resected ssDNA (see the ‘Mediator function of BRCA2 in loading of RAD51 onto ssDNA’ section below).

According to the breast cancer information core database, more than 25% of cancer-associated missense mutations map to the C-terminal region (residues 2500–2850) (Szabo et al. 2000), which includes the sequence through which BRCA2 binds to DSS1 (Jeyasekharan et al. 2013). Recent analysis of cancer-associated BRCA2 mutations has led to the identification of a nuclear export signal (NES) in the C-terminal region of BRCA2 (Jeyasekharan et al. 2013). The NES is masked by the interaction with DSS1. Interestingly, a common cancer-associated BRCA2 mutation, D2723H, impairs the binding of BRCA2 to DSS1, leading to its mislocalization to the cytoplasm and disruption of RAD51 loading onto damaged chromatin. Notably, this mutation is likely to decrease RAD51 foci formation even in the presence of normal BRCA2, suggesting that this mutation acts in a dominant-negative manner.

The three OB domains in BRCA2 are structurally very similar to the canonical OB fold, like the one in RPA (Murzin 1993), consisting of a highly curved β-sheet that closes on itself to form a β barrel. Both the OB2 and OB3 folds have the obvious groove that is characteristic of the ssDNA-binding sites of canonical OB folds (Yang et al. 2002). Using electron microscopy, Thorslund and coworkers unveiled that purified human BRCA2 selectively binds to single-stranded DNA in tailed duplexes and replication fork structures (Thorslund et al. 2010). The Tower domain is capable of binding duplex DNA; however, full-length BRCA2 is likely to interact primarily with ssDNA.

C-terminal RAD51 binding site

Esashi and coworkers demonstrated that RAD51 directly interacts with a region near the BRCA2 C-terminal end that has no homology with the BRC repeats. Phosphorylation of this region at Ser3291 by cyclin-dependent kinase (CDK) disrupts the C-terminal BRCA2–RAD51 interaction (Esashi et al. 2005). The level of phosphorylation at this residue is low during S phase when RAD51 activity is high and increases as the cell enters mitosis. DNA damage elicits a block of this phosphorylation, suggesting that this modification can modulate BRCA2 activity. Unlike BRC repeats, the C-terminal RAD51-binding domain selectively interacts with RAD51 oligomers and RAD51 nucleoprotein filaments. This region protects RAD51 nucleoprotein filaments formed on ssDNA from dissociation by the BRC repeats. The FA-D1 patient cell line, EUFA423, expresses truncated BRCA2 lacking the C-terminal 192 amino acid residues, which means that the C-terminal RAD51–binding domain (residues 3265–3330) is lost in this patient. This cell line showed impairments in RAD51 focus formation and HR activity (Wang et al. 2004). In addition, an individual with HBOC has been reported to carry a deletion of the C-terminal 224 residues of BRCA2 (Håkansson et al. 1997). These observations underscore the importance of tumor suppression of the C-terminal RAD51-binding region that regulates RAD51 nucleoprotein filament formation (Esashi et al. 2007, Ayoub et al. 2009).

Mediator function of BRCA2 in loading of RAD51 onto ssDNA

RAD51 protein itself can bind to both ssDNA and dsDNA. In HR, because RAD51 must initially bind to resected ssDNA tails at the DSB, and the ssDNA is quickly coated with RPA, RAD51 requires a targeting factor that mediates its interaction with ssDNA. It has been shown that full-length purified human BRCA2 is able to enhance RAD51 presynaptic assembly on RPA-coated ssDNA, promoting RPA-RAD51 exchange. BRCA2 can stimulate RAD51 ssDNA binding in vitro, while inhibiting the ability of RAD51 to bind dsDNA (Carreira et al. 2009, Shivji et al. 2009). Mechanistically, BRCA2 stabilizes ATP-bound RAD51-ssDNA filaments by blocking ATP hydrolysis (Carreira et al. 2009, Jensen et al. 2010). Unlike yeast Rad52, which plays a dominant mediator role for yeast Rad51, BRCA2 does not bind RPA directly. How, then, does BRCA2 regulate RPA-RAD51 exchange?

A key factor turns out to be DSS1, the small and highly acidic protein that interacts with OB1 of BRCA2 (Yang et al. 2002). Purified human DSS1 in the presence of BRCA2 stimulates RAD51 binding to RPA-covered ssDNA, compared with BRCA2 alone (Liu et al. 2010). In contrast, DSS1 alone does not activate RAD51 binding to RPA-ssDNA. Furthermore, Zhao and coworkers demonstrated that DSS1 targets BRCA2 to RPA, and DSS1 functions as a DNA mimic to promote the removal of RPA from ssDNA, thereby promoting exchange with RAD51 on ssDNA (Zhao et al. 2015).

It is known that there are other molecules involved in RAD51 regulation. These include RAD51AP1 (Wiese et al. 2007) and TONSL/MMS22L (Duro et al. 2010, O’Donnell et al. 2010). Further analysis will shed more light on the possible interplay between these proteins and HR mechanisms underlying disorders like FA or HBOC.

Additional regulators of HR and RAD51 function

XPG, which is affected in xeroderma pigmentosum complementation group G (XP-G), has been reported to form a complex with BRCA2 and DSS1. Trego and coworkers searched for novel XPG partners and unexpectedly found that XPG interacts with BRCA2, RAD51 and PALB2 (Trego et al. 2016). XPG forms foci in S phase, but not in G1 cells. Because XPG depletion caused a decreased presence of RAD51 and BRCA2 in the chromatin fraction, this protein is likely to contribute to HR.

Foci formation by proteins involved in HR, including BRCA1 and RAD51, is tightly regulated during the cell cycle, and they normally accumulate at the site of DNA damage in S and G2 phase, when the cell has sister chromatid DNA. The mechanism by which BRCA1 foci formation is antagonized in G1 phase by proteins that inhibit DNA end resection, such as 53BP1 and RIF1, has been revealed recently (Chapman et al. 2013, Escribano-Diaz et al. 2013, Zimmermann et al. 2013). In addition to this mechanism, it was recently reported that PALB2–BRCA2 cannot bind to BRCA1 specifically in G1 phase owing to the ubiquitination of PALB2. This modification is mediated by the KEAP1–CRL3 ubiquitin ligase, leading to the suppression of HR in G1 (Orthwein et al. 2015). Furthermore, this PALB2 ubiquitination is antagonized by a deubiquitinase USP11. Interestingly, KEAP1 mutations have been reported in breast cancers (Hartikainen et al. 2015).

Genome maintenance and tumor suppression by BRCA2

As discussed previously, monoallelic BRCA2 mutation causes HBOC, whereas biallelic mutations are characteristic of the FA-D1 subgroup, which displays a particularly severe form of FA, with very early onset of leukemia and solid tumors (Hirsch et al. 2004, Wagner et al. 2004). The malignancies observed in FA-D1 patients are not breast or ovarian cancer; however, this is not surprising because these patients are infants whose endocrine and reproductive systems are immature. BRCA1-deficient breast cancers are typically ‘basal-like’ and ‘triple negative’ for epidermal growth factor receptor 2 (HER2), progesterone receptor and estrogen receptor, and they are more recalcitrant to conventional therapy. On the other hand, breast cancer stemming from mutated BRCA2 is clinically categorized as similar to a common sporadic form of breast cancer (Roy et al. 2011). The reason why similar HR deficiencies lead to such distinct clinical entities is still poorly understood (Roy et al. 2011).

In line with the two-hit hypothesis proposed by Knudson (1971), it has been considered that the malignant cells in patients carrying monoallelic BRCA2 mutations obligatorily harbor LOH affecting the wild-type allele, leading to the loss of BRCA2 function. However, a recent study showed that out of 90 BRCA-deficient breast cancers, ten cases did not lose chromosomes that harbored normal copies of BRCA genes and did not show signatures that indicate loss of BRCA functions (Nik-Zainal et al. 2016). Furthermore, loss of mutant alleles can occur in BRCA-associated breast cancer (King et al. 2007). Thus, in addition to the loss of all BRCA function, BRCA haploinsufficiency may also promote carcinogenesis, and some of the cancers arising in the HBOC patients may not be HR deficient. Indeed, BRCA1+/− mutated cells are defective in response to replication stress (Pathania et al. 2014). As studies on mice showed (Ludwig et al. 1997, Jonkers et al. 2001), in carcinogenic steps, the cells tend to lose Tp53 (or an equivalent checkpoint gene) before loss of BRCA/HR function to avoid cell death and/or senescence.

Therapeutic implications of HR defects in HBOC

Defective HR is an important target for chemotherapy in HBOC patients (Konstantinopoulos et al. 2015). Platinum-based chemotherapy is a well-established and widely used modality for cancer treatment. As cisplatin and its derivative carboplatin induce intrastrand and interstrand crosslinks (Deans & West 2011), BRCA-deficient, hence ICL repair-deficient, HBOC cells are naturally sensitive to these drugs (De Picciotto et al. 2016). Indeed, BRCA-mutated HBOC patients appear to have a better short-term prognosis compared with non-BRCA patients, perhaps owing to the better response to chemotherapy (Konstantinopoulos et al. 2015), though this may not be the case for long-term survival (McLaughlin et al. 2013). Based on the discovery that PARP inhibition induces a dramatic cell killing in cells deficient in HR (Bryant et al. 2005, Farmer et al. 2005), an exciting opportunity to develop novel chemotherapeutic drugs has emerged. This is an instance where two distinct but important DNA repair activities are simultaneously inhibited, leading to cell death (synthetic lethality). An initial explanation that this lethality was due to impaired base excision repair, with an increased level of single-strand breaks that are converted to toxic DSBs by replication, is now challenged, and revised models have been proposed (Helleday 2011, Konstantinopoulos et al. 2015).

Resistance to chemotherapy drugs invariably appears after the initial clinical response during prolonged treatment. A number of resistance mechanisms have been proposed. It has been suggested that secondary mutations in BRCA genes that restore the wild-type reading frame, leading to recovered HR activity, are the major mechanisms for the acquired resistance (Edwards et al. 2008, Sakai et al. 2008). This is analogous to the reversion mosaicism in hematopoietic cells sometimes observed in FA patients, which may mitigate progression of bone marrow failure (Soulier et al. 2005). Genome instability due to HR defects and selection may contribute to these phenomena. Another mechanism for acquired resistance in BRCA2-deficient tumors is the increased replication fork stability without restoring HR (Chaudhuri et al. 2016). This could be due to several mechanisms including PTIP deficiency that inhibits the access of MRE11 nuclease to stalled replication forks (Chaudhuri et al. 2016) or loss of the nucleosome remodeling factor CHD4 (Guillemette et al. 2015). In BRCA1-mutated tumors, normal levels of HR activities might be restored by the loss of 53BP1 or REV7/MAD2L2 (Bouwman et al. 2010, Boersma et al. 2015, Xu et al. 2015). In the absence of BRCA1, these genes function to prevent end resection of DSBs, blocking the subsequent HR reaction.

BRCA-deficient tumors may accumulate an enormous number of mutations due to HR defects and genome instability during the carcinogenic process. Thus, these cells may carry higher numbers of tumor-specific peptide antigens that are presented to tumor-infiltrating lymphocytes. This hypothesis has been tested in clinical samples, leading to the conclusion that BRCA1/2-mutated high-grade serous ovarian cancer may be more sensitive to recently developed immune checkpoint inhibitors, such as anti-PD-1 or anti-PD-L1 antibodies (Strickland et al. 2016).

How defects in HR and ICL repair affect hematopoietic stem cells or promote cancer development?

Deficiencies in DNA repair limit the renewal capacity of aging hematopoietic stem cells (Rossi et al. 2007), and FA patients have higher levels of DNA damage in these cells, leading to the upregulation of p53 and cell death/senescence (Ceccaldi et al. 2012). The origin of the endogenous damage in FA is an important issue that needs to be resolved (Garaycoechea & Patel 2014) because this knowledge may allow us to develop a novel strategy for preventing bone marrow failure and cancer. Likewise, it is important to know by what mechanism the genome is destabilized in BRCA mutation carriers.

Endogenous aldehydes and lipid peroxidation products have been proposed as major sources of spontaneous DNA damage in FA (Garaycoechea et al. 2012). Using a mutant cell panel derived from the chicken DT40 cell line, it was shown that cells lacking Fancd2 or Brca2 are particularly sensitive to formaldehyde at concentrations similar to those in normal human serum (Ridpath et al. 2007). Patel and coworkers indicated in a series of papers that aldehyde detoxifying enzymes ALDH2 (which mainly catalyzes acetaldehyde) and ADH5 (which mainly catalyzes formaldehyde) play critical roles in FA model mice in the suppression of bone marrow failure and leukemogenesis (Langevin et al. 2011, Garaycoechea et al. 2012, Oberbeck et al. 2014, Pontel et al. 2015). These results clearly indicate that endogenous aldehydes can damage DNA in hematopoietic stem cells. As East Asians often carry an enzymatically defective ALDH2 variant allele (ALDH2*2), we examined ALDH2 genotypes in our cohort of Japanese FA patients (Hira et al. 2013). In line with the mouse studies, our results indicated that the ALDH2 variant allele accelerates the progression of bone marrow failure in these patients. Strikingly, we identified several FA children who had homozygous ALDH2 mutations. These patients displayed particularly grave symptoms, including an extremely early onset of myelodysplasia (Hira et al. 2013, Yabe et al. 2016). This combined FA–ALDH2 deficiency could be considered to be a distinct disease entity. It will be exciting to see whether ALDH2 can be a drug target to prevent bone marrow failure in FA patients. A compound that stimulates ALDH2 activity has already been developed (Perez-Miller et al. 2010). It will also be interesting to test how ALDH2 status can affect cancer development in HBOC among East Asian populations.

Another source of DNA damage can be DNA replication fork stalling, which likely contributes to genome instability in FA or HBOC. The nascent DNA strand at the blocked fork is protected by RAD51 filaments stabilized by the C-terminal domain of BRCA2 in a manner independent of HR (Schlacher et al. 2011). In the absence of BRCA2, BRCA1 or FANCD2, the stalled fork cannot be protected and is degraded by MRE11 nuclease, leading to the loss of genetic information or genome rearrangements (Schlacher et al. 2012). This mechanism might be important for genome stability and tumor suppression provided by BRCA1/2 or FA genes.

Genes involved in pre-mRNA splicing and in the biogenesis and export of messenger ribonucleoprotein (mRNP) also have an important role for genome stability (Paulsen et al. 2009). R-loops consisting of DNA–RNA hybrids and a displaced single-stranded DNA often arise when transcription is perturbed (e.g., upon collision of transcription bubbles and replication forks). Thus, R-loops may be a chief source of replication stress and cancer-associated genome instability. Bhatia and coworkers demonstrated the accumulation of R-loops in BRCA2-depleted cells (Bhatia et al. 2015). Furthermore, recent studies asked whether the FA pathway coordinates transcription–replication conflicts and is involved in R-loop resolution (García-Rubio et al. 2015, Schwab et al. 2015). Indeed, human and mouse cells deficient in FA gene function accumulate R-loops, indicating that the FA pathway does play a critical role in R-loop resolution. MMC-induced FANCD2 foci levels are reduced by the expression of RNaseH1, which digests RNA in RNA–DNA hybrids. These studies imply that the accumulation of R-loops might contribute to hematopoietic stem cell exhaustion in FA. It will also be interesting to know how R-loops trigger the activation of the FA pathway.

Of note, it has also been proposed that cytokines that are upregulated in FA, such as TNF-α or TGF-β, may directly harm hematopoietic stem cells or modulate DNA damage repair in the stem cell compartment (Du et al. 2014, Zhang et al. 2016). This line of investigation may inform the development of novel therapeutic strategies for FA.

Conclusions

As summarized in this review, there has been a lot of progress toward the mechanistic understanding of HR repair and genome stability in this decade. Furthermore, we have seen an exciting development of PARP inhibitors as novel and promising cancer therapeutics. Detailed knowledge about the pathogenesis of HBOC and FA has been obtained.

Despite this progress, obvious questions are still lingering in the field. It is a true enigma that HR deficiency leads to carcinogenesis in a tissue-specific manner, although such specificity is also often the case for the other hereditary cancer syndromes. From a practical point of view, a large number of variants of unknown significance (VUS) generated from genetic testing of BRCA1 or BRCA2 pose a significant problem in the interpretation of the test results. In the long run, accumulated knowledge about segregation of the genotype and an individual’s cancer susceptibility within families may eventually clarify the significance of VUS. At the moment, careful evaluation of DNA repair capacity in lymphocytes from cases with VUS might be useful (Pathania et al. 2014, Vaclová et al. 2015). It would be particularly useful to construct a collection of isogenic knock-in cells with candidate variants using the CRISPR–CAS9 system (Paquet et al. 2016). Endogenous aldehydes may include at least several molecular species (Xie et al. 2016), and they may induce various types of DNA damage, such as monoadducts, interstrand crosslinks or DNA–protein crosslinks. Which of these actually contributes to FA pathology, and how endogenous aldehydes are generated in cells should be elucidated in the near future.

Declaration of interest

The authors declare that there is no conflict of interest that could be perceived as prejudicing the impartiality of this review.

Funding

Our work is supported by grants from The Ministry of Education, Culture, Sports, Science and Technology (MEXT), a Grant-in-Aid for Scientific Research (KAKENHI) (KAKENHI Grant Numbers 23114010, 24310042, 26550026 and 15H01738) and grants from the Ministry of Health, Labour and Welfare.

Acknowledgements

The authors would like to thank our collaborators and past and current members of our laboratory for their contribution, and Prof. James Hejna (Graduate School of Biostudies, Kyoto University) for critical reading and English editing of the manuscript.

This paper forms part of a thematic review section on 20 Years of BRCA 1 and 2.

The Guest Editors for this section were Kokichi Sugano and William Foulkes.

References

  • AlterBP2014Fanconi anemia and the development of leukemia. Best Practice & Research. Clinical Haematology27214221. (doi:10.1016/
j.beha.2014.10.002)

  • AmezianeNMayPHaitjemaAvan de VrugtHJvan Rossum-FikkertSERisticDWilliamsGJBalkJRockxDLiH2015A novel Fanconi anaemia subtype associated with a dominant-negative mutation in RAD51. Nature Communications68829. (doi:10.1038/ncomms9829)

  • AuerbachAD2009Fanconi anemia and its diagnosis. Mutation Research668410. (doi:10.1016/j.mrfmmm.2009.01.013)

  • AyoubNRajendraESuXJeyasekharanADMahenRVenkitaramanAR2009The carboxyl terminus of Brca2 links the disassembly of Rad51 complexes to mitotic entry. Current Biology1910751085. (doi:10.1016/j.cub.2009.05.057)

  • BerwickMSatagopanJMBen-PoratLCarlsonAMahKHenryRDiottiRMiltonKPujaraKLandersT2007Genetic heterogeneity among Fanconi anemia heterozygotes and risk of cancer. Cancer Research6795919596. (doi:10.1158/0008-5472.CAN-07-1501)

  • BhatiaVBarrosoSIGarcía-RubioMLTuminiEHerrera-MoyanoEAguileraA2015BRCA2 prevents R-loop accumulation and associates with TREX-2 mRNA export factor PCID2. Nature511362365. (doi:10.1038/nature13374)

  • BoersmaVMoattiNSegura-BayonaSPeuscherMHvan der TorreJWeversBAOrthweinADurocherDJacobsJJL2015MAD2L2 controls DNA repair at telomeres and DNA breaks by inhibiting 5′ end resection. Nature521537540. (doi:10.1038/nature14216)

  • BoglioloMSurrallésJ2015Fanconi anemia: a model disease for studies on human genetics and advanced therapeutics. Current Opinion in Genetics & Development333240. (doi:10.1016/j.gde.2015.07.002)

  • BoglioloMSchusterBStoepkerCDerkuntBSuYRaamsATrujilloJPMinguillónJRamírezMJPujolR2013Mutations in ERCC4, encoding the DNA-repair endonuclease XPF, cause Fanconi anemia. American Journal of Human Genetics92800806. (doi:10.1016/j.ajhg.2013.04.002)

  • BouwmanPAlyAEscandellJMPieterseMBartkovaJvan der GuldenHHiddinghSThanasoulaMKulkarniAYangQ201053BP1 loss rescues BRCA1 deficiency and is associated with triple-negative and BRCA-mutated breast cancers. Nature Structural & Molecular Biology17688695. (doi:10.1038/nsmb.1831)

  • BryantHESchultzNThomasHDParkerKMFlowerDLopezEKyleSMeuthMCurtinNJHelledayT2005Specific killing of BRCA2-deficient tumours with inhibitors of poly(ADP-ribose) polymerase. Nature434913917. (doi:10.1038/nature03443)

  • BuissonRDion-CôtéA-MCoulombeYLaunayHCaiHStasiakAZStasiakAXiaBMassonJ-Y2010Cooperation of breast cancer proteins PALB2 and piccolo BRCA2 in stimulating homologous recombination. Nature Structural & Molecular Biology1712471254. (doi:10.1038/nsmb.1915)

  • Cancer Genome Atlas Research Network2011Integrated genomic analyses of ovarian carcinoma. Nature474609615. (doi:10.1038/nature10166)

  • CarreiraAKowalczykowskiSC2011Two classes of BRC repeats in BRCA2 promote RAD51 nucleoprotein filament function by distinct mechanisms. PNAS1081044810453. (doi:10.1073/pnas.1106971108)

  • CarreiraAHilarioJAmitaniIBaskinRJShivjiMKKVenkitaramanARKowalczykowskiSC2009The BRC repeats of BRCA2 modulate the DNA-binding selectivity of RAD51. Cell13610321043. (doi:10.1016/j.cell.2009.02.019)

  • CastellàMJacquemontCThompsonELYeoJECheungRSHuangJ-WSobeckAHendricksonEATaniguchiT2015FANCI regulates recruitment of the FA core complex at sites of DNA damage independently of FANCD2. PLoS Genetics11e1005563. (doi:10.1371/journal.pgen.1005563)

  • CeccaldiRParmarKMoulyEDelordMKimJMRegairazMPlaMVasquezNZhangQ-SPondarreC2012Bone marrow failure in Fanconi anemia is triggered by an exacerbated p53/p21 DNA damage response that impairs hematopoietic stem and progenitor cells. Cell Stem Cell113649. (doi:10.1016/j.stem.2012.05.013)

  • CeccaldiRSarangiPD’AndreaAD2016The Fanconi anaemia pathway: new players and new functions. Nature Reviews Molecular Cell Biology17337349. (doi:10.1038/nrm.2016.48)

  • ChapmanJRTaylorMRGBoultonSJ2012Playing the end game: DNA double-strand break repair pathway choice. Molecular Cell47497510. (doi:10.1016/j.molcel.2012.07.029)

  • ChapmanJRBarralPVannierJ-BBorelVStegerMTomas-LobaASartoriAAAdamsIRBatistaFDBoultonSJ2013RIF1 is essential for 53BP1-dependent nonhomologous end joining and suppression of DNA double-strand break resection. Molecular Cell49858871. (doi:10.1016/j.molcel.2013.01.002)

  • ChaudhuriARCallenEDingXGogolaEDuarteAALeeJ-EWongNLafargaVCalvoJAPanzarinoNJ2016Replication fork stability confers chemoresistance in BRCA-deficient cells. Nature535382387. (doi:10.1038/nature18325)

  • ChenY-HJonesMJKYinYCristSBColnaghiLSimsRJRothenbergEJallepalliPVHuangTT2015ATR-mediated phosphorylation of FANCI regulates dormant origin firing in response to replication stress. Molecular Cell58323338. (doi:10.1016/j.molcel.2015.02.031)

  • CollinsNBWilsonJBBushTThomashevskiARobertsKJJonesNJKupferGM2009ATR-dependent phosphorylation of FANCA on serine 1449 after DNA damage is important for FA pathway function. Blood11321812190. (doi:10.1182/blood-2008-05-154294)

  • ConnorFBertwistleDMeePJRossGMSwiftSGrigorievaETybulewiczVLAshworthA1997Tumorigenesis and a DNA repair defect in mice with a truncating Brca2 mutation. Nature Genetics17423430. (doi:10.1038/ng1297-423)

  • CousineauIBelmaazaA2011EMSY overexpression disrupts the BRCA2/RAD51 pathway in the DNA-damage response: implications for chromosomal instability/recombination syndromes as checkpoint diseases. Molecular Genetics and Genomics285325340. (doi:10.1007/s00438-011-0612-5)

  • De PicciottoNCacheuxWRothAChappuisPOLabidi-GalySI2016Ovarian cancer: status of homologous recombination pathway as a predictor of drug response. Critical Reviews in Oncology/Hematology1015059. (doi:10.1016/j.critrevonc.2016.02.014)

  • DeansAJWestSC2011DNA interstrand crosslink repair and cancer. Nature Reviews Cancer11467480. (doi:10.1038/nrc3088)

  • DebatisseMLe TallecBLetessierADutrillauxBBrisonO2012Common fragile sites: mechanisms of instability revisited. Trends in Genetics282232. (doi:10.1016/j.tig.2011.10.003)

  • DomchekSMTangJStopferJLilliDRHamelNTischkowitzMMonteiroANAMessickTEPowersJYonkerA2013Biallelic deleterious BRCA1 mutations in a woman with early-onset ovarian cancer. Cancer Discovery3399405. (doi:10.1158/2159-8290.CD-12-0421)

  • DuWErdenOPangQ2014TNF-α signaling in Fanconi anemia. 52211. (doi:10.1016/j.bcmd.2013.06.005)

  • DuroELundinCAskKSanchez-PulidoLMacArtneyTJTothRPontingCPGrothAHelledayTRouseJ2010Identification of the MMS22L-TONSL complex that promotes homologous recombination. Molecular Cell40632644. (doi:10.1016/j.molcel.2010.10.023)

  • DuxinJPWalterJC2015What is the DNA repair defect underlying Fanconi anemia?Current Opinion in Cell Biology374960. (doi:10.1016/j.ceb.2015.09.002)

  • EdwardsSLBroughRLordCJNatrajanRVatchevaRLevineDABoydJReis-FilhoJSAshworthA2008Resistance to therapy caused by intragenic deletion in BRCA2. Nature45111111115. (doi:10.1038/nature06548)

  • EsashiFChristNGannonJLiuYHuntTJasinMWestSC2005CDK-dependent phosphorylation of BRCA2 as a regulatory mechanism for recombinational repair. Nature434598604. (doi:10.1038/nature03404)

  • EsashiFGalkinVEYuXEgelmanEHWestSC2007Stabilization of RAD51 nucleoprotein filaments by the C-terminal region of BRCA2. Nature Structural & Molecular Biology14468474. (doi:10.1038/nsmb1245)

  • Escribano-DiazCOrthweinAFradet-TurcotteAXingMYoungJTFTká010DJCookMARosebrockAPMunroMCannyMD2013A cell cycle-dependent regulatory circuit composed of 53BP1-RIF1 and BRCA1-CtIP controls DNA repair pathway choice. Molecular Cell49872883. (doi:10.1016/j.molcel.2013.01.001)

  • FarmerHMcCabeNLordCJTuttANJJohnsonDARichardsonTBSantarosaMDillonKJHicksonIKnightsC2005Targeting the DNA repair defect in BRCA mutant cells as a therapeutic strategy. Nature434917921. (doi:10.1038/nature03445)

  • FerlayJSoerjomataramIDikshitREserSMathersCRebeloMParkinDMFormanDBrayF2015Cancer incidence and mortality worldwide: sources, methods and major patterns in GLOBOCAN 2012. International Journal of Cancer136E359E386. (doi:10.1002/ijc.29210)

  • FriedmanLSThistlethwaiteFCPatelKJYuVPLeeHVenkitaramanARAbelKJCarltonMBHunterSMColledgeWH1998Thymic lymphomas in mice with a truncating mutation in Brca2. Cancer Research5813381343.

  • GaraycoecheaJIPatelKJ2014Why does the bone marrow fail in Fanconi anemia?Blood1232634. (doi:10.1182/blood-2013-09-427740)

  • GaraycoecheaJICrossanGPLangevinFDalyMArendsMJPatelKJ2012Genotoxic consequences of endogenous aldehydes on mouse haematopoietic stem cell function. Nature489571575. (doi:10.1038/nature11368)

  • Garcia-HigueraITaniguchiTGanesanSMeynMSTimmersCHejnaJGrompeMD’AndreaAD2001Interaction of the Fanconi anemia proteins and BRCA1 in a common pathway. Molecular Cell7249262. (doi:10.1016/S1097-2765(01)00173-3)

  • García-RubioMLPérez-CaleroCBarrosoSITuminiEHerrera-MoyanoERosadoIVAguileraA2015The Fanconi anemia pathway protects genome integrity from R-loops. PLoS Genetics11e1005674. (doi:10.1371/journal.pgen.1005674)

  • GarnerEKimYLachFPKottemannMCSmogorzewskaA2013Human GEN1 and the SLX4-associated nucleases MUS81 and SLX1 are essential for the resolution of replication-induced Holliday junctions. Cell Reports5207215. (doi:10.1016/j.celrep.2013.08.041)

  • GuillemetteSSerraRWPengMHayesJAKonstantinopoulosPAGreenMRCantorSB2015Resistance to therapy in BRCA2 mutant cells due to loss of the nucleosome remodeling factor CHD4. Genes & Development29489494. (doi:10.1101/gad.256214.114)

  • HartikainenJMTengströmMWinqvistRJukkola-VuorinenAPylkäsKKosmaV-MSoiniYMannermaaA2015KEAP1 Genetic polymorphisms associate with breast cancer risk and survival outcomes. Clinical Cancer Research2115911601. (doi:10.1158/
1078-0432.CCR-14-1887)

  • HashimotoYPudduFCostanzoV2012RAD51- and MRE11-dependent reassembly of uncoupled CMG helicase complex at collapsed replication forks. Nature Structural & Molecular Biology191724. (doi:10.1038/nsmb.2177)

  • HåkanssonSJohannssonOJohanssonUSellbergGLomanNGerdesAMHolmbergEDahlNPandisNKristofferssonU1997Moderate frequency of BRCA1 and BRCA2 germ-line mutations in Scandinavian familial breast cancer. American Journal of Human Genetics6010681078.

  • HelledayT2011The underlying mechanism for the PARP and BRCA synthetic lethality: clearing up the misunderstandings. Molecular Oncology5387393. (doi:10.1016/j.molonc.2011.07.001)

  • HiraAYabeHYoshidaKOkunoYShiraishiYChibaKTanakaHMiyanoSNakamuraJKojimaS2013Variant ALDH2 is associated with accelerated progression of bone marrow failure in Japanese Fanconi anemia patients. Blood12232063209. (doi:10.1182/blood-2013-06-507962)

  • HiraAYoshidaKSatoKOkunoYShiraishiYChibaKTanakaHMiyanoSShimamotoATaharaH2015Mutations in the gene encoding the E2 conjugating enzyme UBE2T cause Fanconi anemia. American Journal of Human Genetics9610011007. (doi:10.1016/j.ajhg.2015.04.022)

  • HirschBShimamuraAMoreauLBaldingerSHag-alshiekhMBostromBSencerSD’AndreaAD2004Association of biallelic BRCA2/FANCD1 mutations with spontaneous chromosomal instability and solid tumors of childhood. Blood10325542559. (doi:10.1182/blood-2003-06-1970)

  • HodskinsonMRGSilhanJCrossanGPGaraycoecheaJIMukherjeeSJohnsonCMSchärerODPatelKJ2014Mouse SLX4 is a tumor suppressor that stimulates the activity of the nuclease XPF-ERCC1 in DNA crosslink repair. Molecular Cell54472484. (doi:10.1016/
j.molcel.2014.03.014)

  • HoeijmakersJH2001Genome maintenance mechanisms for preventing cancer. Nature411366374. (doi:10.1038/35077232)

  • HowlettNGTaniguchiTOlsonSCoxBWaisfiszQDe Die-SmuldersCPerskyNGrompeMJoenjeHPalsG2002Biallelic inactivation of BRCA2 in Fanconi anemia. Science297606609. (doi:10.1126/science.1073834)

  • HuangMKimJMShiotaniBYangKZouLD’AndreaAD2010The FANCM/FAAP24 complex is required for the DNA interstrand crosslink-induced checkpoint response. Molecular Cell39259268. (doi:10.1016/j.molcel.2010.07.005)

  • HuangJLiuSBellaniMAThazhathveetilAKLingCde WinterJPWangYWangWSeidmanMM2013The DNA translocase FANCM/MHF promotes replication traverse of DNA interstrand crosslinks. Molecular Cell52434446. (doi:10.1016/j.molcel.2013.09.021)

  • Hughes-DaviesLHuntsmanDRuasMFuksFByeJChinS-FMilnerJBrownLAHsuFGilksB2003EMSY links the BRCA2 pathway to sporadic breast and ovarian cancer. Cell115523535. (doi:10.1016/S0092-8674(03)00930-9)

  • IshiaiMKitaoHSmogorzewskaATomidaJKinomuraAUchidaESaberiAKinoshitaEKinoshita-KikutaEKoikeT2008FANCI phosphorylation functions as a molecular switch to turn on the Fanconi anemia pathway. Nature Structural & Molecular Biology1511381146. (doi:10.1038/nsmb.1504)

  • JensenRBCarreiraAKowalczykowskiSC2010Purified human BRCA2 stimulates RAD51-mediated recombination. Nature467678683. (doi:10.1038/nature09399)

  • JeyasekharanADLiuYHattoriHPisupatiVJonsdottirABRajendraELeeMSundaramoorthyESchlachterSKaminskiCF2013A cancer-associated BRCA2 mutation reveals masked nuclear export signals controlling localization. Nature Structural & Molecular Biology2011911198. (doi:10.1038/nsmb.2666)

  • JonkersJMeuwissenRvan der GuldenHPeterseHvan der ValkMBernsA2001Synergistic tumor suppressor activity of BRCA2 and p53 in a conditional mouse model for breast cancer. Nature Genetics29418425. (doi:10.1038/ng747)

  • JooWXuGPerskyNSSmogorzewskaARudgeDGBuzovetskyOElledgeSJPavletichNP2011Structure of the FANCI-FANCD2 complex: insights into the Fanconi anemia DNA repair pathway. Science333312316. (doi:10.1126/science.1205805)

  • KashiyamaKNakazawaYPilzDTGuoCShimadaMSasakiKFawcettHWingJFLewinSOCarrL2013Malfunction of nuclease ERCC1-XPF results in diverse clinical manifestations and causes Cockayne syndrome, xeroderma pigmentosum, and Fanconi anemia. American Journal of Human Genetics92807819. (doi:10.1016/j.ajhg.2013.04.007)

  • KiiskiJIPelttariLMKhanSFreysteinsdottirESReynisdottirIHartSNShimelisHVilskeSKallioniemiASchleutkerJ2014Exome sequencing identifies FANCM as a susceptibility gene for triple-negative breast cancer. PNAS1111517215177. (doi:10.1073/pnas.1407909111)

  • KimJMKeeYGurtanAD’AndreaAD2008Cell cycle-dependent chromatin loading of the Fanconi anemia core complex by FANCM/FAAP24. Blood11152155222. (doi:10.1182/blood-2007-09-113092)

  • KimHYangKDejsuphongDD’AndreaAD2012Regulation of Rev1 by the Fanconi anemia core complex. Nature Structural & Molecular Biology19164170. (doi:10.1038/nsmb.2222)

  • KimYSpitzGSVeturiULachFPAuerbachADSmogorzewskaA2013Regulation of multiple DNA repair pathways by the Fanconi anemia protein SLX4. Blood1215463. (doi:10.1182/blood-2012-07-441212)

  • KingTALiWBrogiEYeeCJGemignaniMLOlveraNLevineDANortonLRobsonMEOffitK2007Heterogenic loss of the wild-type BRCA allele in human breast tumorigenesis. Annals of Surgical Oncology1425102518. (doi:10.1245/s10434-007-9372-1)

  • KitaoHTakataM2011Fanconi anemia: a disorder defective in the DNA damage response. International Journal of Hematology93417424. (doi:10.1007/s12185-011-0777-z)

  • Klein DouwelDBoonenRACMLongDTSzypowskaAARäschleMWalterJCKnipscheerP2014XPF-ERCC1 acts in Unhooking DNA interstrand crosslinks in cooperation with FANCD2 and FANCP/SLX4. Molecular Cell54460471. (doi:10.1016/j.molcel.2014.03.015)

  • KnipscheerPRäschleMSmogorzewskaAEnoiuMHoTVSchärerODElledgeSJWalterJC2009The Fanconi anemia pathway promotes replication-dependent DNA interstrand cross-link repair. Science32616981701. (doi:10.1126/science.1182372)

  • KnudsonAG1971Mutation and cancer: statistical study of retinoblastoma. PNAS68820823. (doi:10.1073/pnas.68.4.820)

  • KomoriKHidakaMHoriuchiTFujikaneRShinagawaHIshinoY2004Cooperation of the N-terminal helicase and C-terminal endonuclease activities of Archaeal Hef protein in processing stalled replication forks. Journal of Biological Chemistry2795317553185. (doi:10.1074/jbc.M409243200)

  • KonstantinopoulosPACeccaldiRShapiroGID’AndreaAD2015Homologous recombination deficiency: exploiting the fundamental vulnerability of ovarian cancer. Cancer Discovery511371154. (doi:10.1158/2159-8290.CD-15-0714)

  • KottemannMCSmogorzewskaA2013Fanconi anaemia and the repair of Watson and Crick DNA crosslinks. Nature493356363. (doi:10.1038/nature11863)

  • LachaudCCastorDHainKMuñozIWilsonJMacArtneyTJSchindlerDRouseJ2014Distinct functional roles for the two SLX4 ubiquitin-binding UBZ domains mutated in Fanconi anemia. Journal of Cell Science12728112817. (doi:10.1242/jcs.146167)

  • LachaudCMorenoAMarchesiFTothRBlowJJRouseJ2016Ubiquitinated Fancd2 recruits Fan1 to stalled replication forks to prevent genome instability. Science351846849. (doi:10.1126/science.aad5634)

  • LangevinFCrossanGPRosadoIVArendsMJPatelKJ2011Fancd2 counteracts the toxic effects of naturally produced aldehydes in mice. Nature4755358. (doi:10.1038/nature10192)

  • LiangC-CZhanBYoshikawaYHaasWGygiSPCohnMA2015UHRF1 is a sensor for DNA interstrand crosslinks and recruits FANCD2 to initiate the Fanconi anemia pathway. Cell Reports1019471956. (doi:10.1016/j.celrep.2015.02.053)

  • LiangC-CLiZLopez-MartinezDNicholsonWVVénien-BryanCCohnMA2016The FANCD2-FANCI complex is recruited to DNA interstrand crosslinks before monoubiquitination of FANCD2. Nature Communications712124. (doi:10.1038/ncomms12124)

  • LimETWürtzPHavulinnaASPaltaPTukiainenTRehnströmKEskoTMägiRInouyeMLappalainenT2014Distribution and medical impact of loss-of-function variants in the Finnish founder population. PLoS Genetics10e1004494. (doi:10.1371/journal.pgen.1004494)

  • LiuJDotyTGibsonBHeyerW-D2010Human BRCA2 protein promotes RAD51 filament formation on RPA-covered single-stranded DNA. Nature Structural & Molecular Biology1712601262. (doi:10.1038/nsmb.1904)

  • LobitzSVelleuerE2006Guido Fanconi (1892–1979): a jack of all trades. Nature Reviews. Cancer6893898. (doi:10.1038/nrc2009)

  • LongDTRäschleMJoukovVWalterJC2011Mechanism of RAD51-dependent DNA interstrand cross-link repair. Science3338487. (doi:10.1126/science.1204258)

  • LossaintGLarroqueMRibeyreCBecNLarroqueCDécailletCGariKConstantinouA2013FANCD2 binds MCM proteins and controls replisome function upon activation of s phase checkpoint signaling. Molecular Cell51678690. (doi:10.1016/j.molcel.2013.07.023)

  • LudwigTChapmanDLPapaioannouVEEfstratiadisA1997Targeted mutations of breast cancer susceptibility gene homologs in mice: lethal phenotypes of Brca1, Brca2, Brca1/Brca2, Brca1/p53, and Brca2/p53 nullizygous embryos. Genes & Development1112261241. (doi:10.1101/gad.11.10.1226)

  • MassonJYTarsounasMCStasiakAZStasiakAShahRMcIlwraithMJBensonFEWestSC2001Identification and purification of two distinct complexes containing the five RAD51 paralogs. Genes & Development1532963307. (doi:10.1101/gad.947001)

  • MatsushitaNKitaoHIshiaiMNagashimaNHiranoSOkawaKOhtaTYuDSMcHughPJHicksonID2005A FancD2-monoubiquitin fusion reveals hidden functions of Fanconi anemia core complex in DNA repair. Molecular Cell19841847. (doi:10.1016/j.molcel.2005.08.018)

  • McLaughlinJRRosenBMoodyJPalTFanIShawPARischHASellersTASunPNarodSA2013Long-term ovarian cancer survival associated with mutation in BRCA1 or BRCA2. Journal of the National Cancer Institute105141148. (doi:10.1093/jnci/djs494)

  • MeeteiARMedhurstALLingCXueYSinghTRBierPSteltenpoolJStoneSDokalIMathewCG2005A human ortholog of archaeal DNA repair protein Hef is defective in Fanconi anemia complementation group M. Nature Genetics37958963. (doi:10.1038/ng1626)

  • MenzelTNähse-KumpfVKousholtANKleinDKLund-AndersenCLeesMJohansenJVSyljuåsenRGSørensenCS2011A genetic screen identifies BRCA2 and PALB2 as key regulators of G2 checkpoint maintenance. EMBO Reports12705712. (doi:10.1038/embor.2011.99)

  • MikiYSwensenJShattuck-EidensDFutrealPHarshmanKTavtigianSLiuQCochranCBennettLDingW1994A strong candidate for the breast and ovarian cancer susceptibility gene BRCA1. Science2666671. (doi:10.1126/science.7545954)

  • MurinaOAesch vonCKarakusUFerrettiLPBolckHAHänggiKSartoriAA2014FANCD2 and CtIP cooperate to repair DNA interstrand crosslinks. Cell Reports710301038. (doi:10.1016/j.celrep.2014.03.069)

  • MurzinAG1993OB(oligonucleotide/oligosaccharide binding)-fold: common structural and functional solution for non-homologous sequences. EMBO Journal12861867.

  • NakanishiKYangY-GPierceAJTaniguchiTDigweedMD’AndreaADWangZ-QJasinM2005Human Fanconi anemia monoubiquitination pathway promotes homologous DNA repair. PNAS10211101115. (doi:10.1073/pnas.0407796102)

  • NielsenFCvan Overeem HansenT2016Hereditary breast and ovarian cancer: new genes in confined pathways. Nature Reviews Cancer16599612. (doi:10.1038/nrc.2016.72)

  • Nik-ZainalSDaviesHStaafJRamakrishnaMGlodzikDZouXMartincorenaIAlexandrovLBMartinSWedgeDC2016Landscape of somatic mutations in 560 breast cancer whole-genome sequences. Nature5344754. (doi:10.1038/nature17676)

  • O’DonnellLPanierSWildenhainJTkachJMAl-HakimALandryM-CEscribano-DiazCSzilardRKYoungJTFMunroM2010The MMS22L-TONSL complex mediates recovery from replication stress and homologous recombination. Molecular Cell40619631. (doi:10.1016/j.molcel.2010.10.024)

  • OberbeckNLangevinFKingGde WindNCrossanGPPatelKJ2014Maternal aldehyde elimination during pregnancy preserves the fetal genome. Molecular Cell55807817. (doi:10.1016/
j.molcel.2014.07.010)

  • OrthweinANoordermeerSMWilsonMDLandrySEnchevRISherkerAMunroMPinderJSalsmanJDellaireG2015A mechanism for the suppression of homologous recombination in G1 cells. Nature528422426. (doi:10.1038/nature16142)

  • PaquetDKwartDChenASproulAJacobSTeoSOlsenKMGreggANoggleSTessier-LavigneM2016Efficient introduction of specific homozygous and heterozygous mutations using CRISPR/Cas9. Nature533125129. (doi:10.1038/nature17664)

  • PatelKJYuVPLeeHCorcoranAThistlethwaiteFCEvansMJColledgeWHFriedmanLSPonderBAVenkitaramanAR1998Involvement of Brca2 in DNA repair. Molecular Cell1347357. (doi:10.1016/S1097-2765(00)80035-0)

  • PathaniaSBadeSLe GuillouMBurkeKReedRBowman-ColinCSuYTingDTPolyakKRichardsonAL2014BRCA1 haploinsufficiency for replication stress suppression in primary cells. Nature Communications55496. (doi:10.1038/ncomms6496)

  • PaulsenRDSoniDVWollmanRHahnATYeeM-CGuanAHesleyJAMillerSCCromwellEFSolow-CorderoDE2009A genome-wide siRNA screen reveals diverse cellular processes and pathways that mediate genome stability. Molecular Cell35228239. (doi:10.1016/j.molcel.2009.06.021)

  • PellegriniLYuDSLoTAnandSLeeMBlundellTLVenkitaramanAR2002Insights into DNA recombination from the structure of a RAD51-BRCA2 complex. Nature420287293. (doi:10.1038/nature01230)

  • Perez-MillerSYounusHVanamRChenC-HMochly-RosenDHurleyTD2010Alda-1 is an agonist and chemical chaperone for the common human aldehyde dehydrogenase 2 variant. Nature Structural & Molecular Biology17159164. (doi:10.1038/nsmb.1737)

  • PeterlongoPCatucciIColomboMCalecaLMucakiEBoglioloMMarínMDamiolaFBernardLPensottiV2015FANCM c.5791C>T nonsense mutation (rs144567652) induces exon skipping, affects DNA repair activity and is a familial breast cancer risk factor. Human Molecular Genetics2453455355. (doi:10.1093/hmg/ddv251)

  • PontelLBRosadoIVBurgos-BarraganGGaraycoecheaJIYuRArendsMJChandrasekaranGBroeckerVWeiWLiuL2015Endogenous formaldehyde is a hematopoietic stem cell genotoxin and metabolic carcinogen. Molecular Cell60177188. (doi:10.1016/
j.molcel.2015.08.020)

  • PrakashRZhangYFengWJasinM2015Homologous recombination and human health: the roles of BRCA1, BRCA2, and associated proteins. Cold Spring Harbor Perspectives in Biology7a016600. (doi:10.1101/cshperspect.a016600)

  • RäschleMKnipsheerPEnoiuMAngelovTSunJGriffithJDEllenbergerTESchärerODWalterJC2008Mechanism of replication-coupled DNA interstrand crosslink repair. Cell134969980. (doi:10.1016/j.cell.2008.08.030)

  • ReidSSchindlerDHanenbergHBarkerKHanksSKalbRNevelingKKellyPSealSFreundM2007Biallelic mutations in PALB2 cause Fanconi anemia subtype FA-N and predispose to childhood cancer. Nature Genetics39162164. (doi:10.1038/ng1947)

  • RickmanKALachFPAbhyankarADonovanFXSanbornEMKennedyJASougnezCGabrielSBElementoOChandrasekharappaSC2015Deficiency of UBE2T, the E2 ubiquitin ligase necessary for FANCD2 and FANCI ubiquitination, causes FA-T subtype of Fanconi anemia. Cell Reports123541. (doi:10.1016/j.celrep.2015.06.014)

  • RidpathJRNakamuraATanoKLukeAMSonodaEArakawaHBuersteddeJ-MGillespieDAFSaleJEYamazoeM2007Cells deficient in the FANC/BRCA pathway are hypersensitive to plasma levels of formaldehyde. Cancer Research671111711122. (doi:10.1158/0008-5472.CAN-07-3028)

  • RossiDJBryderDSeitaJNussenzweigAHoeijmakersJWeissmanIL2007Deficiencies in DNA damage repair limit the function of haematopoietic stem cells with age. Nature447725729. (doi:10.1038/nature05862)

  • RouetPSmihFJasinM1994Expression of a site-specific endonuclease stimulates homologous recombination in mammalian cells. PNAS9160646068. (doi:10.1073/pnas.91.13.6064)

  • RoyRChunJPowellSN2011BRCA1 and BRCA2: different roles in a common pathway of genome protection. Nature Reviews Cancer126878. (doi:10.1038/nrc3181)

  • SaekiHSiaudNChristNWiegantWWvan BuulPPWHanMZdzienickaMZStarkJMJasinM2006Suppression of the DNA repair defects of BRCA2-deficient cells with heterologous protein fusions. PNAS10387688773. (doi:10.1073/pnas.0600298103)

  • SakaiWSwisherEMKarlanBYAgarwalMKHigginsJFriedmanCVillegasEJacquemontCFarrugiaDJCouchFJ2008Secondary mutations as a mechanism of cisplatin resistance in BRCA2-mutated cancers. Nature45111161120. (doi:10.1038/nature06633)

  • SarbajnaSWestSC2014Holliday junction processing enzymes as guardians of genome stability. Trends in Biochemical Sciences39409419. (doi:10.1016/j.tibs.2014.07.003)

  • SasakiMSTonomuraA1973A high susceptibility of Fanconi’s anemia to chromosome breakage by DNA cross-linking agents. Cancer Research3318291836.

  • SatoKIshiaiMTodaKFurukoshiSOsakabeATachiwanaHTakizawaYKagawaWKitaoHDohmaeN2012Histone chaperone activity of Fanconi anemia proteins, FANCD2 and FANCI, is required for DNA crosslink repair. EMBO Journal3135243536. (doi:10.1038/emboj.2012.197)

  • SawyerSLTianLKahkonenMSchwartzentruberJKircherMMajewskiJDymentDAInnesAM2015Biallelic mutations in BRCA1 cause a new Fanconi anemia subtype. Cancer Discovery5135142. (doi:10.1158/2159-8290.CD-14-1156)

  • SchlacherKChristNSiaudNEgashiraAWuHJasinM2011Double-strand break repair-independent role for BRCA2 in blocking stalled replication fork degradation by MRE11. Cell145529542. (doi:10.1016/j.cell.2011.03.041)

  • SchlacherKWuHJasinM2012A distinct replication fork protection pathway connects Fanconi anemia tumor suppressors to RAD51-BRCA1/2. Cancer Cell22106116. (doi:10.1016/j.ccr.2012.05.015)

  • SchroederTMAnschützFKnoppA1964[Spontaneous chromosome aberrations in familial panmyelopathy]. Humangenetik1194196.

  • SchwabRANieminuszczyJShahFLangtonJMartinezDLLiangC-CCohnMAGibbonsRJDeansAJNiedzwiedzW2015The Fanconi anemia pathway maintains genome stability by coordinating replication and transcription. Molecular Cell60351361. (doi:10.1016/j.molcel.2015.09.012)

  • SealSBarfootRJayatilakeHSmithPRenwickABascombeLMcGuffogLEvansDGEcclesDEastonDF2003Evaluation of Fanconi anemia genes in familial breast cancer predisposition. Cancer Research6385968599.

  • ShahidTSorokaJKongEHMalivertLMcilwraithMJPapeTWestSCZhangX2014Structure and mechanism of action of the BRCA2 breast cancer tumor suppressor. Nature Structural & Molecular Biology21962968. (doi:10.1038/nsmb.2899)

  • ShamseldinHEElfakiMAlkurayaFS2012Exome sequencing reveals a novel Fanconi group defined by XRCC2 mutation. Journal of Medical Genetics49184186. (doi:10.1136/jmedgenet-2011-100585)

  • ShivjiMKKMukundSRRajendraEChenSShortJMSavillJKlenermanDVenkitaramanAR2009The BRC repeats of human BRCA2 differentially regulate RAD51 binding on single- versus double-stranded DNA to stimulate strand exchange. PNAS1061325413259. (doi:10.1073/pnas.0906208106)

  • SinghTRBakkerSTAgarwalSJansenMGrassmanEGodthelpBCAliAMDuC-HRooimansMAFanQ2009Impaired FANCD2 monoubiquitination and hypersensitivity to camptothecin uniquely characterize Fanconi anemia complementation group M. Blood114174180. (doi:10.1182/blood-2009-02-207811)

  • SinghTRAliAMParamasivamMPradhanAWahengbamKSeidmanMMMeeteiAR2013ATR-dependent phosphorylation of FANCM at serine 1045 is essential for FANCM functions. Cancer Research7343004310. (doi:10.1158/0008-5472.CAN-12-3976)

  • SmogorzewskaAMatsuokaSVinciguerraPMcDonaldERHurovKELuoJBallifBAGygiSPHofmannKD’AndreaAD2007Identification of the FANCI protein, a monoubiquitinated FANCD2 paralog required for DNA repair. Cell129289301. (doi:10.1016/
j.cell.2007.03.009)

  • SonodaESasakiMSBuersteddeJ-MBezzubovaOShinoharaAOgawaHTakataMYamaguchi-IwaiYTakedaS1998Rad51-deficient vertebrate cells accumulate chromosomal breaks prior to cell death. EMBO Journal17598608. (doi:10.1093/emboj/17.2.598)

  • SoulierJLeblancTLargheroJDastotHShimamuraAGuardiolaPEsperouHFerryCJubertCFeugeasJ-P2005Detection of somatic mosaicism and classification of Fanconi anemia patients by analysis of the FA/BRCA pathway. Blood10513291336. (doi:10.1182/blood-2004-05-1852)

  • StricklandKCHowittBEShuklaSARodigSRitterhouseLLLiuJFGarberJEChowdhuryDWuCJD’AndreaAD2016Association and prognostic significance of BRCA1/2-mutation status with neoantigen load, number of tumor-infiltrating lymphocytes and expression of PD-1/PD-L1 in high grade serous ovarian cancer. Oncotarget71358713598. (doi:10.18632/oncotarget.7277)

  • SySMHHuenMSYChenJ2009PALB2 is an integral component of the BRCA complex required for homologous recombination repair. PNAS10671557160. (doi:10.1073/pnas.0811159106)

  • SzaboCMasielloARyanJFBrodyLC2000The breast cancer information core: database design, structure, and scope. Human Mutation16123131. (doi:10.1002/1098-1004(200008)16:2<123::aid-humu4>3.0.co;2-y)

  • TakataMSasakiMSSonodaEMorrisonCHashimotoMUtsumiHYamaguchi-IwaiYShinoharaATakedaS1998Homologous recombination and non-homologous end-joining pathways of DNA double-strand break repair have overlapping roles in the maintenance of chromosomal integrity in vertebrate cells. EMBO Journal1754975508. (doi:10.1093/emboj/17.18.5497)

  • TakataMSasakiMSSonodaEFukushimaTMorrisonCAlbalaJSSwagemakersSMKanaarRThompsonLHTakedaS2000The Rad51 paralog Rad51B promotes homologous recombinational repair. Molecular and Cellular Biology2064766482. (doi:10.1128/MCB.20.17.6476-6482.2000)

  • TakataMSasakiMSTachiiriSFukushimaTSonodaESchildDThompsonLHTakedaS2001Chromosome instability and defective recombinational repair in knockout mutants of the five Rad51 paralogs. Molecular and Cellular Biology2128582866. (doi:10.1128/MCB.21.8.2858-2866.2001)

  • TakataMTachiiriSFujimoriAThompsonLHMikiYHiraokaMTakedaSYamazoeM2002Conserved domains in the chicken homologue of BRCA2. Oncogene2111301134. (doi:10.1038/sj.onc.1205168)

  • TaylorMRGŠpírekMChaurasiyaKRWardJDCarzanigaRYuXEgelmanEHCollinsonLMRuedaDKrejciL2015Rad51 paralogs remodel pre-synaptic Rad51 filaments to stimulate homologous recombination. Cell162271286. (doi:10.1016/
j.cell.2015.06.015)

  • ThompsonERDoyleMARylandGLRowleySMChoongDYHTothillRWThorne HBarnesDRLiJ2012Exome sequencing identifies rare deleterious mutations in DNA repair genes FANCC and BLM as potential breast cancer susceptibility alleles. PLoS Genetics8e1002894. (doi:10.1371/journal.pgen.1002894)

  • ThorslundTMcilwraithMJComptonSALekomtsevSPetronczkiMGriffithJDWestSC2010The breast cancer tumor suppressor BRCA2 promotes the specific targeting of RAD51 to single-stranded DNA. Nature Structural & Molecular Biology1712631265. (doi:10.1038/nsmb.1905)

  • TianYParamasivamMGhosalGChenDShenXHuangYAkhterSLegerskiRChenJSeidmanMM2015UHRF1 Contributes to DNA damage repair as a lesion recognition factor and nuclease scaffold. Cell Reports1019571966. (doi:10.1016/j.celrep.2015.03.038)

  • TorreLABrayFSiegelRLFerlayJ2015Global cancer statistics, 2012. CA: A Cancer Journal for Clinicians6587108. (doi:10.3322/caac.21262)

  • TregoKSGroesserTDavalosARParplysACZhaoWNelsonMRHlaingAShihBRydbergBPluthJM2016Non-catalytic roles for XPG with BRCA1 and BRCA2 in homologous recombination and genome stability. Molecular Cell61535546. (doi:10.1016/j.molcel.2015.12.026)

  • TsuzukiTFujiiYSakumiKTominagaYNakaoKSekiguchiMMatsushiroAYoshimuraYMoritaT1996Targeted disruption of the Rad51 gene leads to lethality in embryonic mice. PNAS9362366240. (doi:10.1073/pnas.93.13.6236)

  • UnnoJItayaATaokaMSatoKTomidaJSakaiWSugasawaKIshiaiMIkuraTIsobeT2014FANCD2 binds CtIP and regulates DNA-end resection during DNA interstrand crosslink repair. Cell Reports710391047. (doi:10.1016/j.celrep.2014.04.005)

  • VaclováTGómez-LópezGSetiénFBuenoJMGMacíasJABarrosoAUriosteMEstellerMBenitezJOsorioA2015DNA repair capacity is impaired in healthy BRCA1 heterozygous mutation carriers. Breast Cancer Research and Treatment152271282. (doi:10.1007/s10549-015-3459-3)

  • VasanthakumarAArnovitzSMarquezRLeporeJRafidiGAsomAWeatherlyMDavisEMNeistadtBDuszynskiR2016Brca1 deficiency causes bone marrow failure and spontaneous hematologic malignancies in mice. Blood127310313. (doi:10.1182/blood-2015-03-635599)

  • VazFHanenbergHSchusterBBarkerKWiekCErvenVNevelingKEndtDKestertonIAutoreF2010Mutation of the RAD51C gene in a Fanconi anemia-like disorder. Nature Genetics42406409. (doi:10.1038/ng.570)

  • VirtsELJankowskaAMacKayCGlaasMFWiekCKelichSLLottmannNKennedyFMMarchalCLehnertE2015AluY-mediated germline deletion, duplication and somatic stem cell reversion in UBE2T defines a new subtype of Fanconi anemia. Human Molecular Genetics2450935108. (doi:10.1093/hmg/ddv227)

  • WagnerJETolarJLevranOSchollTDeffenbaughASatagopanJBen-PoratLMahKBatishSDKutlerDI2004Germline mutations in BRCA2: shared genetic susceptibility to breast cancer, early onset leukemia, and Fanconi anemia. Blood10332263229. (doi:10.1182/blood-2003-09-3138)

  • WangXAndreassenPRD’AndreaAD2004Functional interaction of monoubiquitinated FANCD2 and BRCA2/FANCD1 in chromatin. Molecular and Cellular Biology2458505862. (doi:10.1128/MCB.24.13.5850-5862.2004)

  • WangATKimTWagnerJEContiBALachFPHuangALMolinaHSanbornEMZierhutHCornesBK2015A dominant mutation in human RAD51 reveals its function in DNA interstrand crosslink repair independent of homologous recombination. Molecular Cell59478490. (doi:10.1016/j.molcel.2015.07.009)

  • WestSC2003Molecular views of recombination proteins and their control. Nature Reviews Molecular Cell Biology4435445. (doi:10.1038/nrm1127)

  • WieseCDrayEGroesserTSan FilippoJShiICollinsDWTsaiM-SWilliamsGJRydbergBSungP2007Promotion of homologous recombination and genomic stability by RAD51AP1 via RAD51 recombinase enhancement. Molecular Cell28482490. (doi:10.1016/j.molcel.2007.08.027)

  • WoosterRBignellGLancasterJSwiftSSealSMangionJCollinsNGregorySGumbsCMicklemG1995Identification of the breast cancer susceptibility gene BRCA2. Nature378789792. (doi:10.1038/378789a0)

  • WuLHicksonID2003The Bloom’s syndrome helicase suppresses crossing over during homologous recombination. Nature426870874. (doi:10.1038/nature02253)

  • WyattHDMSarbajnaSMatosJWestSC2013Coordinated actions of SLX1-SLX4 and MUS81-EME1 for holliday junction resolution in human cells. Molecular Cell52234247. (doi:10.1016/
j.molcel.2013.08.035)

  • XiaBShengQNakanishiKOhashiAWuJChristNLiuXJasinMCouchFJLivingstonDM2006Control of BRCA2 cellular and clinical functions by a nuclear partner, PALB2. Molecular Cell22719729. (doi:10.1016/j.molcel.2006.05.022)

  • XieM-ZShoulkamyMISalemAMHObaSGodaMNakanoTIdeH2016Aldehydes with high and low toxicities inactivate cells by damaging distinct cellular targets. Mutation Research7864151. (doi:10.1016/j.mrfmmm.2016.02.005)

  • XuGChapmanJRBrandsmaIYuanJMistrikMBouwmanPBartkovaJGogolaEWarmerdamDBarazasM2015REV7 counteracts DNA double-strand break resection and affects PARP inhibition. Nature521541544. (doi:10.1038/nature14328)

  • YabeMYabeHMorimotoTFukumuraAOhtsuboKKoikeTYoshidaKOgawaSItoEOkunoY2016The phenotype and clinical course of Japanese Fanconi anaemia infants is influenced by patient, but not maternal ALDH2 genotype. British Journal of Haematology[in press]. (doi:10.1111/bjh.14243)

  • YamamotoKIshiaiMMatsushitaNArakawaHLamerdinJEBuersteddeJ-MTanimotoMHaradaMThompsonLHTakataM2003Fanconi anemia FANCG protein in mitigating radiation- and enzyme-induced DNA double-strand breaks by homologous recombination in vertebrate cells. Molecular and Cellular Biology2354215430. (doi:10.1128/MCB.23.15.5421-5430.2003)

  • YamamotoKHiranoSIshiaiMMorishimaKKitaoHNamikoshiKKimuraMMatsushitaNArakawaHBuersteddeJ-M2004Fanconi anemia protein FANCD2 promotes immunoglobulin gene conversion and DNA repair through a mechanism related to homologous recombination. Molecular and Cellular Biology253443. (doi:10.1128/MCB.25.1.34-43.2005)

  • YamamotoKNKobayashiSTsudaMKurumizakaHTakataMKonoKJiricnyJTakedaSHirotaK2011Involvement of SLX4 in interstrand cross-link repair is regulated by the Fanconi anemia pathway. PNAS10864926496. (doi:10.1073/pnas.1018487108)

  • YangHJeffreyPDMillerJKinnucanESunYThomaNHZhengNChenP-LLeeW-HPavletichNP2002BRCA2 function in DNA binding and recombination from a BRCA2-DSS1-ssDNA structure. Science29718371848. (doi:10.1126/science.
297.5588.1837)

  • YonetaniYHocheggerHSonodaEShinyaSYoshikawaHTakedaSYamazoeM2005Differential and collaborative actions of Rad51 paralog proteins in cellular response to DNA damage. Nucleic Acids Research3345444552. (doi:10.1093/nar/gki766)

  • ZhangFMaJWuJYeLCaiHXiaBYuX2009PALB2 links BRCA1 and BRCA2 in the DNA-damage response. Current Biology19524529. (doi:10.1016/j.cub.2009.02.018)

  • ZhangHKozonoDEO’ConnorKWVidal-CardenasSRousseauAHamiltonAMoreauLGaudianoEFGreenbergerJBagbyG2016TGF-β inhibition rescues hematopoietic stem cell defects and bone marrow failure in Fanconi anemia. Cell Stem Cell18668681. (doi:10.1016/j.stem.2016.03.002)

  • ZhaoWVaithiyalingamSSan FilippoJMaranonDGJimenez-SainzJFontenayGVKwonYLeungSGLuLJensenRB2015Promotion of BRCA2-dependent homologous recombination by DSS1 via RPA targeting and DNA mimicry. Molecular Cell59176187. (doi:10.1016/j.molcel.2015.05.032)

  • ZimmermannMLottersbergerFBuonomoSBSfeirAde LangeT201353BP1 regulates DSB repair using Rif1 to control 5′ end resection. Science339700704. (doi:10.1126/science.1231573)

 

An official journal of

Society for Endocrinology

Sections

Figures

  • View in gallery

    An overview of the ICL repair pathway. When a replisome collides with an ICL (I), the leading strand initially stalls −20 nucleotides away from the lesion (II). After a second fork converges at the ICL, BRCA1 facilitates the dissociation of the CMG complex (consisting of Cdc45, MCM2-7 and GINS) from chromatin at the stalled fork, allowing the leading strand to approach to the −1 nucleotide adjacent to the ICL (III). In the next step, the SLX4-XPF-ERCC1 complex incises the DNA and unhooks the lesion in a FA pathway-dependent manner (IV). Translesion synthesis (TLS) polymerase (Polζ, or possibly Polκ, Polη, or Polι) extends the leading strand synthesis past the unhooked, ICL-associated nucleotide (V), and HR and the nucleotide excision repair (NER) pathway repair the remaining lesion (VI).

  • View in gallery

    Schematic of HR pathway. When a double-strand break (DSB) is generated after DNA replication during S and G2 phase, both strands are resected in the 5′ to 3′ direction to generate 3′ overhangs. Almost immediately, replication protein A (RPA) is loaded onto the single-stranded (ss) DNA, and then replaced by a RAD51 nucleoprotein filament in a process requiring BRCA1–PALB2–BRCA2. RAD51 carries out strand invasion of the sister chromatid by the ssDNA tail and extends the resulting D-loop formation. In synthesis-dependent strand annealing (SDSA), the D-loop structure quickly dissociates from the ssDNA after synthesis of a complementary single strand, and then another strand anneals with a processed ssDNA. An alternate pathway forms a double Holliday junction (dHJ). After second end capture and fill-in synthesis, the Holliday junction is dissociated by the TopIIIα–BLM complex or resolved by resolvase complexes that contain SLX4, MUS81 and GEN1.

  • View in gallery

    The mechanism of FA pathway activation. Upon replication fork stalling, FANCM–FAAP24–MHF1/2 complex binding at the ICL lesion activates ATR signaling, followed by the recruitment of the FA core complex. FANCI is phosphorylated by ATR, and subsequently, the FA core complex with UBE2T mediates the monoubiquitination of the FANCI–FANCD2 (D2-I) complex. This modification targets the D2-I complex to chromatin and leads to multiple events that repair the ICL lesion.

  • View in gallery

    The primary structures of human BRCA1 and BRCA2. BRCA1 consists of 1863 amino acids. The RING domain is in the N-terminus and partly overlaps with the BARD1-binding region. BRCA1 interacts with PALB2 via a coiled-coil region in the C-terminus. The BRCT motif is a phosphorylated protein-binding sequence, and this motif mediates the association of BRCA1 with Abraxas, CtIP and BACH1, which are all known factors in DSB repair. BRCA2 comprises 3418 amino acids. The PALB2 binding region is located in the amino (N) terminus. In the center, BRCA2 harbors eight BRC repeats, which constitute a binding region for RAD51 monomers. In the carboxy (C) terminus, a DNA-binding domain contains three OB folds and a region that interacts with DSS1. The C-terminal region of BRCA2 is also important for the formation of the RAD51 nucleoprotein filament and BRCA2 nuclear localization.

References

AlterBP2014Fanconi anemia and the development of leukemia. Best Practice & Research. Clinical Haematology27214221. (doi:10.1016/
j.beha.2014.10.002)

AmezianeNMayPHaitjemaAvan de VrugtHJvan Rossum-FikkertSERisticDWilliamsGJBalkJRockxDLiH2015A novel Fanconi anaemia subtype associated with a dominant-negative mutation in RAD51. Nature Communications68829. (doi:10.1038/ncomms9829)

AuerbachAD2009Fanconi anemia and its diagnosis. Mutation Research668410. (doi:10.1016/j.mrfmmm.2009.01.013)

AyoubNRajendraESuXJeyasekharanADMahenRVenkitaramanAR2009The carboxyl terminus of Brca2 links the disassembly of Rad51 complexes to mitotic entry. Current Biology1910751085. (doi:10.1016/j.cub.2009.05.057)

BerwickMSatagopanJMBen-PoratLCarlsonAMahKHenryRDiottiRMiltonKPujaraKLandersT2007Genetic heterogeneity among Fanconi anemia heterozygotes and risk of cancer. Cancer Research6795919596. (doi:10.1158/0008-5472.CAN-07-1501)

BhatiaVBarrosoSIGarcía-RubioMLTuminiEHerrera-MoyanoEAguileraA2015BRCA2 prevents R-loop accumulation and associates with TREX-2 mRNA export factor PCID2. Nature511362365. (doi:10.1038/nature13374)

BoersmaVMoattiNSegura-BayonaSPeuscherMHvan der TorreJWeversBAOrthweinADurocherDJacobsJJL2015MAD2L2 controls DNA repair at telomeres and DNA breaks by inhibiting 5′ end resection. Nature521537540. (doi:10.1038/nature14216)

BoglioloMSurrallésJ2015Fanconi anemia: a model disease for studies on human genetics and advanced therapeutics. Current Opinion in Genetics & Development333240. (doi:10.1016/j.gde.2015.07.002)

BoglioloMSchusterBStoepkerCDerkuntBSuYRaamsATrujilloJPMinguillónJRamírezMJPujolR2013Mutations in ERCC4, encoding the DNA-repair endonuclease XPF, cause Fanconi anemia. American Journal of Human Genetics92800806. (doi:10.1016/j.ajhg.2013.04.002)

BouwmanPAlyAEscandellJMPieterseMBartkovaJvan der GuldenHHiddinghSThanasoulaMKulkarniAYangQ201053BP1 loss rescues BRCA1 deficiency and is associated with triple-negative and BRCA-mutated breast cancers. Nature Structural & Molecular Biology17688695. (doi:10.1038/nsmb.1831)

BryantHESchultzNThomasHDParkerKMFlowerDLopezEKyleSMeuthMCurtinNJHelledayT2005Specific killing of BRCA2-deficient tumours with inhibitors of poly(ADP-ribose) polymerase. Nature434913917. (doi:10.1038/nature03443)

BuissonRDion-CôtéA-MCoulombeYLaunayHCaiHStasiakAZStasiakAXiaBMassonJ-Y2010Cooperation of breast cancer proteins PALB2 and piccolo BRCA2 in stimulating homologous recombination. Nature Structural & Molecular Biology1712471254. (doi:10.1038/nsmb.1915)

Cancer Genome Atlas Research Network2011Integrated genomic analyses of ovarian carcinoma. Nature474609615. (doi:10.1038/nature10166)

CarreiraAKowalczykowskiSC2011Two classes of BRC repeats in BRCA2 promote RAD51 nucleoprotein filament function by distinct mechanisms. PNAS1081044810453. (doi:10.1073/pnas.1106971108)

CarreiraAHilarioJAmitaniIBaskinRJShivjiMKKVenkitaramanARKowalczykowskiSC2009The BRC repeats of BRCA2 modulate the DNA-binding selectivity of RAD51. Cell13610321043. (doi:10.1016/j.cell.2009.02.019)

CastellàMJacquemontCThompsonELYeoJECheungRSHuangJ-WSobeckAHendricksonEATaniguchiT2015FANCI regulates recruitment of the FA core complex at sites of DNA damage independently of FANCD2. PLoS Genetics11e1005563. (doi:10.1371/journal.pgen.1005563)

CeccaldiRParmarKMoulyEDelordMKimJMRegairazMPlaMVasquezNZhangQ-SPondarreC2012Bone marrow failure in Fanconi anemia is triggered by an exacerbated p53/p21 DNA damage response that impairs hematopoietic stem and progenitor cells. Cell Stem Cell113649. (doi:10.1016/j.stem.2012.05.013)

CeccaldiRSarangiPD’AndreaAD2016The Fanconi anaemia pathway: new players and new functions. Nature Reviews Molecular Cell Biology17337349. (doi:10.1038/nrm.2016.48)

ChapmanJRTaylorMRGBoultonSJ2012Playing the end game: DNA double-strand break repair pathway choice. Molecular Cell47497510. (doi:10.1016/j.molcel.2012.07.029)

ChapmanJRBarralPVannierJ-BBorelVStegerMTomas-LobaASartoriAAAdamsIRBatistaFDBoultonSJ2013RIF1 is essential for 53BP1-dependent nonhomologous end joining and suppression of DNA double-strand break resection. Molecular Cell49858871. (doi:10.1016/j.molcel.2013.01.002)

ChaudhuriARCallenEDingXGogolaEDuarteAALeeJ-EWongNLafargaVCalvoJAPanzarinoNJ2016Replication fork stability confers chemoresistance in BRCA-deficient cells. Nature535382387. (doi:10.1038/nature18325)

ChenY-HJonesMJKYinYCristSBColnaghiLSimsRJRothenbergEJallepalliPVHuangTT2015ATR-mediated phosphorylation of FANCI regulates dormant origin firing in response to replication stress. Molecular Cell58323338. (doi:10.1016/j.molcel.2015.02.031)

CollinsNBWilsonJBBushTThomashevskiARobertsKJJonesNJKupferGM2009ATR-dependent phosphorylation of FANCA on serine 1449 after DNA damage is important for FA pathway function. Blood11321812190. (doi:10.1182/blood-2008-05-154294)

ConnorFBertwistleDMeePJRossGMSwiftSGrigorievaETybulewiczVLAshworthA1997Tumorigenesis and a DNA repair defect in mice with a truncating Brca2 mutation. Nature Genetics17423430. (doi:10.1038/ng1297-423)

CousineauIBelmaazaA2011EMSY overexpression disrupts the BRCA2/RAD51 pathway in the DNA-damage response: implications for chromosomal instability/recombination syndromes as checkpoint diseases. Molecular Genetics and Genomics285325340. (doi:10.1007/s00438-011-0612-5)

De PicciottoNCacheuxWRothAChappuisPOLabidi-GalySI2016Ovarian cancer: status of homologous recombination pathway as a predictor of drug response. Critical Reviews in Oncology/Hematology1015059. (doi:10.1016/j.critrevonc.2016.02.014)

DeansAJWestSC2011DNA interstrand crosslink repair and cancer. Nature Reviews Cancer11467480. (doi:10.1038/nrc3088)

DebatisseMLe TallecBLetessierADutrillauxBBrisonO2012Common fragile sites: mechanisms of instability revisited. Trends in Genetics282232. (doi:10.1016/j.tig.2011.10.003)

DomchekSMTangJStopferJLilliDRHamelNTischkowitzMMonteiroANAMessickTEPowersJYonkerA2013Biallelic deleterious BRCA1 mutations in a woman with early-onset ovarian cancer. Cancer Discovery3399405. (doi:10.1158/2159-8290.CD-12-0421)

DuWErdenOPangQ2014TNF-α signaling in Fanconi anemia. 52211. (doi:10.1016/j.bcmd.2013.06.005)

DuroELundinCAskKSanchez-PulidoLMacArtneyTJTothRPontingCPGrothAHelledayTRouseJ2010Identification of the MMS22L-TONSL complex that promotes homologous recombination. Molecular Cell40632644. (doi:10.1016/j.molcel.2010.10.023)

DuxinJPWalterJC2015What is the DNA repair defect underlying Fanconi anemia?Current Opinion in Cell Biology374960. (doi:10.1016/j.ceb.2015.09.002)

EdwardsSLBroughRLordCJNatrajanRVatchevaRLevineDABoydJReis-FilhoJSAshworthA2008Resistance to therapy caused by intragenic deletion in BRCA2. Nature45111111115. (doi:10.1038/nature06548)

EsashiFChristNGannonJLiuYHuntTJasinMWestSC2005CDK-dependent phosphorylation of BRCA2 as a regulatory mechanism for recombinational repair. Nature434598604. (doi:10.1038/nature03404)

EsashiFGalkinVEYuXEgelmanEHWestSC2007Stabilization of RAD51 nucleoprotein filaments by the C-terminal region of BRCA2. Nature Structural & Molecular Biology14468474. (doi:10.1038/nsmb1245)

Escribano-DiazCOrthweinAFradet-TurcotteAXingMYoungJTFTká010DJCookMARosebrockAPMunroMCannyMD2013A cell cycle-dependent regulatory circuit composed of 53BP1-RIF1 and BRCA1-CtIP controls DNA repair pathway choice. Molecular Cell49872883. (doi:10.1016/j.molcel.2013.01.001)

FarmerHMcCabeNLordCJTuttANJJohnsonDARichardsonTBSantarosaMDillonKJHicksonIKnightsC2005Targeting the DNA repair defect in BRCA mutant cells as a therapeutic strategy. Nature434917921. (doi:10.1038/nature03445)

FerlayJSoerjomataramIDikshitREserSMathersCRebeloMParkinDMFormanDBrayF2015Cancer incidence and mortality worldwide: sources, methods and major patterns in GLOBOCAN 2012. International Journal of Cancer136E359E386. (doi:10.1002/ijc.29210)

FriedmanLSThistlethwaiteFCPatelKJYuVPLeeHVenkitaramanARAbelKJCarltonMBHunterSMColledgeWH1998Thymic lymphomas in mice with a truncating mutation in Brca2. Cancer Research5813381343.

GaraycoecheaJIPatelKJ2014Why does the bone marrow fail in Fanconi anemia?Blood1232634. (doi:10.1182/blood-2013-09-427740)

GaraycoecheaJICrossanGPLangevinFDalyMArendsMJPatelKJ2012Genotoxic consequences of endogenous aldehydes on mouse haematopoietic stem cell function. Nature489571575. (doi:10.1038/nature11368)

Garcia-HigueraITaniguchiTGanesanSMeynMSTimmersCHejnaJGrompeMD’AndreaAD2001Interaction of the Fanconi anemia proteins and BRCA1 in a common pathway. Molecular Cell7249262. (doi:10.1016/S1097-2765(01)00173-3)

García-RubioMLPérez-CaleroCBarrosoSITuminiEHerrera-MoyanoERosadoIVAguileraA2015The Fanconi anemia pathway protects genome integrity from R-loops. PLoS Genetics11e1005674. (doi:10.1371/journal.pgen.1005674)

GarnerEKimYLachFPKottemannMCSmogorzewskaA2013Human GEN1 and the SLX4-associated nucleases MUS81 and SLX1 are essential for the resolution of replication-induced Holliday junctions. Cell Reports5207215. (doi:10.1016/j.celrep.2013.08.041)

GuillemetteSSerraRWPengMHayesJAKonstantinopoulosPAGreenMRCantorSB2015Resistance to therapy in BRCA2 mutant cells due to loss of the nucleosome remodeling factor CHD4. Genes & Development29489494. (doi:10.1101/gad.256214.114)

HartikainenJMTengströmMWinqvistRJukkola-VuorinenAPylkäsKKosmaV-MSoiniYMannermaaA2015KEAP1 Genetic polymorphisms associate with breast cancer risk and survival outcomes. Clinical Cancer Research2115911601. (doi:10.1158/
1078-0432.CCR-14-1887)

HashimotoYPudduFCostanzoV2012RAD51- and MRE11-dependent reassembly of uncoupled CMG helicase complex at collapsed replication forks. Nature Structural & Molecular Biology191724. (doi:10.1038/nsmb.2177)

HåkanssonSJohannssonOJohanssonUSellbergGLomanNGerdesAMHolmbergEDahlNPandisNKristofferssonU1997Moderate frequency of BRCA1 and BRCA2 germ-line mutations in Scandinavian familial breast cancer. American Journal of Human Genetics6010681078.

HelledayT2011The underlying mechanism for the PARP and BRCA synthetic lethality: clearing up the misunderstandings. Molecular Oncology5387393. (doi:10.1016/j.molonc.2011.07.001)

HiraAYabeHYoshidaKOkunoYShiraishiYChibaKTanakaHMiyanoSNakamuraJKojimaS2013Variant ALDH2 is associated with accelerated progression of bone marrow failure in Japanese Fanconi anemia patients. Blood12232063209. (doi:10.1182/blood-2013-06-507962)

HiraAYoshidaKSatoKOkunoYShiraishiYChibaKTanakaHMiyanoSShimamotoATaharaH2015Mutations in the gene encoding the E2 conjugating enzyme UBE2T cause Fanconi anemia. American Journal of Human Genetics9610011007. (doi:10.1016/j.ajhg.2015.04.022)

HirschBShimamuraAMoreauLBaldingerSHag-alshiekhMBostromBSencerSD’AndreaAD2004Association of biallelic BRCA2/FANCD1 mutations with spontaneous chromosomal instability and solid tumors of childhood. Blood10325542559. (doi:10.1182/blood-2003-06-1970)

HodskinsonMRGSilhanJCrossanGPGaraycoecheaJIMukherjeeSJohnsonCMSchärerODPatelKJ2014Mouse SLX4 is a tumor suppressor that stimulates the activity of the nuclease XPF-ERCC1 in DNA crosslink repair. Molecular Cell54472484. (doi:10.1016/
j.molcel.2014.03.014)

HoeijmakersJH2001Genome maintenance mechanisms for preventing cancer. Nature411366374. (doi:10.1038/35077232)

HowlettNGTaniguchiTOlsonSCoxBWaisfiszQDe Die-SmuldersCPerskyNGrompeMJoenjeHPalsG2002Biallelic inactivation of BRCA2 in Fanconi anemia. Science297606609. (doi:10.1126/science.1073834)

HuangMKimJMShiotaniBYangKZouLD’AndreaAD2010The FANCM/FAAP24 complex is required for the DNA interstrand crosslink-induced checkpoint response. Molecular Cell39259268. (doi:10.1016/j.molcel.2010.07.005)

HuangJLiuSBellaniMAThazhathveetilAKLingCde WinterJPWangYWangWSeidmanMM2013The DNA translocase FANCM/MHF promotes replication traverse of DNA interstrand crosslinks. Molecular Cell52434446. (doi:10.1016/j.molcel.2013.09.021)

Hughes-DaviesLHuntsmanDRuasMFuksFByeJChinS-FMilnerJBrownLAHsuFGilksB2003EMSY links the BRCA2 pathway to sporadic breast and ovarian cancer. Cell115523535. (doi:10.1016/S0092-8674(03)00930-9)

IshiaiMKitaoHSmogorzewskaATomidaJKinomuraAUchidaESaberiAKinoshitaEKinoshita-KikutaEKoikeT2008FANCI phosphorylation functions as a molecular switch to turn on the Fanconi anemia pathway. Nature Structural & Molecular Biology1511381146. (doi:10.1038/nsmb.1504)

JensenRBCarreiraAKowalczykowskiSC2010Purified human BRCA2 stimulates RAD51-mediated recombination. Nature467678683. (doi:10.1038/nature09399)

JeyasekharanADLiuYHattoriHPisupatiVJonsdottirABRajendraELeeMSundaramoorthyESchlachterSKaminskiCF2013A cancer-associated BRCA2 mutation reveals masked nuclear export signals controlling localization. Nature Structural & Molecular Biology2011911198. (doi:10.1038/nsmb.2666)

JonkersJMeuwissenRvan der GuldenHPeterseHvan der ValkMBernsA2001Synergistic tumor suppressor activity of BRCA2 and p53 in a conditional mouse model for breast cancer. Nature Genetics29418425. (doi:10.1038/ng747)

JooWXuGPerskyNSSmogorzewskaARudgeDGBuzovetskyOElledgeSJPavletichNP2011Structure of the FANCI-FANCD2 complex: insights into the Fanconi anemia DNA repair pathway. Science333312316. (doi:10.1126/science.1205805)

KashiyamaKNakazawaYPilzDTGuoCShimadaMSasakiKFawcettHWingJFLewinSOCarrL2013Malfunction of nuclease ERCC1-XPF results in diverse clinical manifestations and causes Cockayne syndrome, xeroderma pigmentosum, and Fanconi anemia. American Journal of Human Genetics92807819. (doi:10.1016/j.ajhg.2013.04.007)

KiiskiJIPelttariLMKhanSFreysteinsdottirESReynisdottirIHartSNShimelisHVilskeSKallioniemiASchleutkerJ2014Exome sequencing identifies FANCM as a susceptibility gene for triple-negative breast cancer. PNAS1111517215177. (doi:10.1073/pnas.1407909111)

KimJMKeeYGurtanAD’AndreaAD2008Cell cycle-dependent chromatin loading of the Fanconi anemia core complex by FANCM/FAAP24. Blood11152155222. (doi:10.1182/blood-2007-09-113092)

KimHYangKDejsuphongDD’AndreaAD2012Regulation of Rev1 by the Fanconi anemia core complex. Nature Structural & Molecular Biology19164170. (doi:10.1038/nsmb.2222)

KimYSpitzGSVeturiULachFPAuerbachADSmogorzewskaA2013Regulation of multiple DNA repair pathways by the Fanconi anemia protein SLX4. Blood1215463. (doi:10.1182/blood-2012-07-441212)

KingTALiWBrogiEYeeCJGemignaniMLOlveraNLevineDANortonLRobsonMEOffitK2007Heterogenic loss of the wild-type BRCA allele in human breast tumorigenesis. Annals of Surgical Oncology1425102518. (doi:10.1245/s10434-007-9372-1)

KitaoHTakataM2011Fanconi anemia: a disorder defective in the DNA damage response. International Journal of Hematology93417424. (doi:10.1007/s12185-011-0777-z)

Klein DouwelDBoonenRACMLongDTSzypowskaAARäschleMWalterJCKnipscheerP2014XPF-ERCC1 acts in Unhooking DNA interstrand crosslinks in cooperation with FANCD2 and FANCP/SLX4. Molecular Cell54460471. (doi:10.1016/j.molcel.2014.03.015)

KnipscheerPRäschleMSmogorzewskaAEnoiuMHoTVSchärerODElledgeSJWalterJC2009The Fanconi anemia pathway promotes replication-dependent DNA interstrand cross-link repair. Science32616981701. (doi:10.1126/science.1182372)

KnudsonAG1971Mutation and cancer: statistical study of retinoblastoma. PNAS68820823. (doi:10.1073/pnas.68.4.820)

KomoriKHidakaMHoriuchiTFujikaneRShinagawaHIshinoY2004Cooperation of the N-terminal helicase and C-terminal endonuclease activities of Archaeal Hef protein in processing stalled replication forks. Journal of Biological Chemistry2795317553185. (doi:10.1074/jbc.M409243200)

KonstantinopoulosPACeccaldiRShapiroGID’AndreaAD2015Homologous recombination deficiency: exploiting the fundamental vulnerability of ovarian cancer. Cancer Discovery511371154. (doi:10.1158/2159-8290.CD-15-0714)

KottemannMCSmogorzewskaA2013Fanconi anaemia and the repair of Watson and Crick DNA crosslinks. Nature493356363. (doi:10.1038/nature11863)

LachaudCCastorDHainKMuñozIWilsonJMacArtneyTJSchindlerDRouseJ2014Distinct functional roles for the two SLX4 ubiquitin-binding UBZ domains mutated in Fanconi anemia. Journal of Cell Science12728112817. (doi:10.1242/jcs.146167)

LachaudCMorenoAMarchesiFTothRBlowJJRouseJ2016Ubiquitinated Fancd2 recruits Fan1 to stalled replication forks to prevent genome instability. Science351846849. (doi:10.1126/science.aad5634)

LangevinFCrossanGPRosadoIVArendsMJPatelKJ2011Fancd2 counteracts the toxic effects of naturally produced aldehydes in mice. Nature4755358. (doi:10.1038/nature10192)

LiangC-CZhanBYoshikawaYHaasWGygiSPCohnMA2015UHRF1 is a sensor for DNA interstrand crosslinks and recruits FANCD2 to initiate the Fanconi anemia pathway. Cell Reports1019471956. (doi:10.1016/j.celrep.2015.02.053)

LiangC-CLiZLopez-MartinezDNicholsonWVVénien-BryanCCohnMA2016The FANCD2-FANCI complex is recruited to DNA interstrand crosslinks before monoubiquitination of FANCD2. Nature Communications712124. (doi:10.1038/ncomms12124)

LimETWürtzPHavulinnaASPaltaPTukiainenTRehnströmKEskoTMägiRInouyeMLappalainenT2014Distribution and medical impact of loss-of-function variants in the Finnish founder population. PLoS Genetics10e1004494. (doi:10.1371/journal.pgen.1004494)

LiuJDotyTGibsonBHeyerW-D2010Human BRCA2 protein promotes RAD51 filament formation on RPA-covered single-stranded DNA. Nature Structural & Molecular Biology1712601262. (doi:10.1038/nsmb.1904)

LobitzSVelleuerE2006Guido Fanconi (1892–1979): a jack of all trades. Nature Reviews. Cancer6893898. (doi:10.1038/nrc2009)

LongDTRäschleMJoukovVWalterJC2011Mechanism of RAD51-dependent DNA interstrand cross-link repair. Science3338487. (doi:10.1126/science.1204258)

LossaintGLarroqueMRibeyreCBecNLarroqueCDécailletCGariKConstantinouA2013FANCD2 binds MCM proteins and controls replisome function upon activation of s phase checkpoint signaling. Molecular Cell51678690. (doi:10.1016/j.molcel.2013.07.023)

LudwigTChapmanDLPapaioannouVEEfstratiadisA1997Targeted mutations of breast cancer susceptibility gene homologs in mice: lethal phenotypes of Brca1, Brca2, Brca1/Brca2, Brca1/p53, and Brca2/p53 nullizygous embryos. Genes & Development1112261241. (doi:10.1101/gad.11.10.1226)

MassonJYTarsounasMCStasiakAZStasiakAShahRMcIlwraithMJBensonFEWestSC2001Identification and purification of two distinct complexes containing the five RAD51 paralogs. Genes & Development1532963307. (doi:10.1101/gad.947001)

MatsushitaNKitaoHIshiaiMNagashimaNHiranoSOkawaKOhtaTYuDSMcHughPJHicksonID2005A FancD2-monoubiquitin fusion reveals hidden functions of Fanconi anemia core complex in DNA repair. Molecular Cell19841847. (doi:10.1016/j.molcel.2005.08.018)

McLaughlinJRRosenBMoodyJPalTFanIShawPARischHASellersTASunPNarodSA2013Long-term ovarian cancer survival associated with mutation in BRCA1 or BRCA2. Journal of the National Cancer Institute105141148. (doi:10.1093/jnci/djs494)

MeeteiARMedhurstALLingCXueYSinghTRBierPSteltenpoolJStoneSDokalIMathewCG2005A human ortholog of archaeal DNA repair protein Hef is defective in Fanconi anemia complementation group M. Nature Genetics37958963. (doi:10.1038/ng1626)

MenzelTNähse-KumpfVKousholtANKleinDKLund-AndersenCLeesMJohansenJVSyljuåsenRGSørensenCS2011A genetic screen identifies BRCA2 and PALB2 as key regulators of G2 checkpoint maintenance. EMBO Reports12705712. (doi:10.1038/embor.2011.99)

MikiYSwensenJShattuck-EidensDFutrealPHarshmanKTavtigianSLiuQCochranCBennettLDingW1994A strong candidate for the breast and ovarian cancer susceptibility gene BRCA1. Science2666671. (doi:10.1126/science.7545954)

MurinaOAesch vonCKarakusUFerrettiLPBolckHAHänggiKSartoriAA2014FANCD2 and CtIP cooperate to repair DNA interstrand crosslinks. Cell Reports710301038. (doi:10.1016/j.celrep.2014.03.069)

MurzinAG1993OB(oligonucleotide/oligosaccharide binding)-fold: common structural and functional solution for non-homologous sequences. EMBO Journal12861867.

NakanishiKYangY-GPierceAJTaniguchiTDigweedMD’AndreaADWangZ-QJasinM2005Human Fanconi anemia monoubiquitination pathway promotes homologous DNA repair. PNAS10211101115. (doi:10.1073/pnas.0407796102)

NielsenFCvan Overeem HansenT2016Hereditary breast and ovarian cancer: new genes in confined pathways. Nature Reviews Cancer16599612. (doi:10.1038/nrc.2016.72)

Nik-ZainalSDaviesHStaafJRamakrishnaMGlodzikDZouXMartincorenaIAlexandrovLBMartinSWedgeDC2016Landscape of somatic mutations in 560 breast cancer whole-genome sequences. Nature5344754. (doi:10.1038/nature17676)

O’DonnellLPanierSWildenhainJTkachJMAl-HakimALandryM-CEscribano-DiazCSzilardRKYoungJTFMunroM2010The MMS22L-TONSL complex mediates recovery from replication stress and homologous recombination. Molecular Cell40619631. (doi:10.1016/j.molcel.2010.10.024)

OberbeckNLangevinFKingGde WindNCrossanGPPatelKJ2014Maternal aldehyde elimination during pregnancy preserves the fetal genome. Molecular Cell55807817. (doi:10.1016/
j.molcel.2014.07.010)

OrthweinANoordermeerSMWilsonMDLandrySEnchevRISherkerAMunroMPinderJSalsmanJDellaireG2015A mechanism for the suppression of homologous recombination in G1 cells. Nature528422426. (doi:10.1038/nature16142)

PaquetDKwartDChenASproulAJacobSTeoSOlsenKMGreggANoggleSTessier-LavigneM2016Efficient introduction of specific homozygous and heterozygous mutations using CRISPR/Cas9. Nature533125129. (doi:10.1038/nature17664)

PatelKJYuVPLeeHCorcoranAThistlethwaiteFCEvansMJColledgeWHFriedmanLSPonderBAVenkitaramanAR1998Involvement of Brca2 in DNA repair. Molecular Cell1347357. (doi:10.1016/S1097-2765(00)80035-0)

PathaniaSBadeSLe GuillouMBurkeKReedRBowman-ColinCSuYTingDTPolyakKRichardsonAL2014BRCA1 haploinsufficiency for replication stress suppression in primary cells. Nature Communications55496. (doi:10.1038/ncomms6496)

PaulsenRDSoniDVWollmanRHahnATYeeM-CGuanAHesleyJAMillerSCCromwellEFSolow-CorderoDE2009A genome-wide siRNA screen reveals diverse cellular processes and pathways that mediate genome stability. Molecular Cell35228239. (doi:10.1016/j.molcel.2009.06.021)

PellegriniLYuDSLoTAnandSLeeMBlundellTLVenkitaramanAR2002Insights into DNA recombination from the structure of a RAD51-BRCA2 complex. Nature420287293. (doi:10.1038/nature01230)

Perez-MillerSYounusHVanamRChenC-HMochly-RosenDHurleyTD2010Alda-1 is an agonist and chemical chaperone for the common human aldehyde dehydrogenase 2 variant. Nature Structural & Molecular Biology17159164. (doi:10.1038/nsmb.1737)

PeterlongoPCatucciIColomboMCalecaLMucakiEBoglioloMMarínMDamiolaFBernardLPensottiV2015FANCM c.5791C>T nonsense mutation (rs144567652) induces exon skipping, affects DNA repair activity and is a familial breast cancer risk factor. Human Molecular Genetics2453455355. (doi:10.1093/hmg/ddv251)

PontelLBRosadoIVBurgos-BarraganGGaraycoecheaJIYuRArendsMJChandrasekaranGBroeckerVWeiWLiuL2015Endogenous formaldehyde is a hematopoietic stem cell genotoxin and metabolic carcinogen. Molecular Cell60177188. (doi:10.1016/
j.molcel.2015.08.020)

PrakashRZhangYFengWJasinM2015Homologous recombination and human health: the roles of BRCA1, BRCA2, and associated proteins. Cold Spring Harbor Perspectives in Biology7a016600. (doi:10.1101/cshperspect.a016600)

RäschleMKnipsheerPEnoiuMAngelovTSunJGriffithJDEllenbergerTESchärerODWalterJC2008Mechanism of replication-coupled DNA interstrand crosslink repair. Cell134969980. (doi:10.1016/j.cell.2008.08.030)

ReidSSchindlerDHanenbergHBarkerKHanksSKalbRNevelingKKellyPSealSFreundM2007Biallelic mutations in PALB2 cause Fanconi anemia subtype FA-N and predispose to childhood cancer. Nature Genetics39162164. (doi:10.1038/ng1947)

RickmanKALachFPAbhyankarADonovanFXSanbornEMKennedyJASougnezCGabrielSBElementoOChandrasekharappaSC2015Deficiency of UBE2T, the E2 ubiquitin ligase necessary for FANCD2 and FANCI ubiquitination, causes FA-T subtype of Fanconi anemia. Cell Reports123541. (doi:10.1016/j.celrep.2015.06.014)

RidpathJRNakamuraATanoKLukeAMSonodaEArakawaHBuersteddeJ-MGillespieDAFSaleJEYamazoeM2007Cells deficient in the FANC/BRCA pathway are hypersensitive to plasma levels of formaldehyde. Cancer Research671111711122. (doi:10.1158/0008-5472.CAN-07-3028)

RossiDJBryderDSeitaJNussenzweigAHoeijmakersJWeissmanIL2007Deficiencies in DNA damage repair limit the function of haematopoietic stem cells with age. Nature447725729. (doi:10.1038/nature05862)

RouetPSmihFJasinM1994Expression of a site-specific endonuclease stimulates homologous recombination in mammalian cells. PNAS9160646068. (doi:10.1073/pnas.91.13.6064)

RoyRChunJPowellSN2011BRCA1 and BRCA2: different roles in a common pathway of genome protection. Nature Reviews Cancer126878. (doi:10.1038/nrc3181)

SaekiHSiaudNChristNWiegantWWvan BuulPPWHanMZdzienickaMZStarkJMJasinM2006Suppression of the DNA repair defects of BRCA2-deficient cells with heterologous protein fusions. PNAS10387688773. (doi:10.1073/pnas.0600298103)

SakaiWSwisherEMKarlanBYAgarwalMKHigginsJFriedmanCVillegasEJacquemontCFarrugiaDJCouchFJ2008Secondary mutations as a mechanism of cisplatin resistance in BRCA2-mutated cancers. Nature45111161120. (doi:10.1038/nature06633)

SarbajnaSWestSC2014Holliday junction processing enzymes as guardians of genome stability. Trends in Biochemical Sciences39409419. (doi:10.1016/j.tibs.2014.07.003)

SasakiMSTonomuraA1973A high susceptibility of Fanconi’s anemia to chromosome breakage by DNA cross-linking agents. Cancer Research3318291836.

SatoKIshiaiMTodaKFurukoshiSOsakabeATachiwanaHTakizawaYKagawaWKitaoHDohmaeN2012Histone chaperone activity of Fanconi anemia proteins, FANCD2 and FANCI, is required for DNA crosslink repair. EMBO Journal3135243536. (doi:10.1038/emboj.2012.197)

SawyerSLTianLKahkonenMSchwartzentruberJKircherMMajewskiJDymentDAInnesAM2015Biallelic mutations in BRCA1 cause a new Fanconi anemia subtype. Cancer Discovery5135142. (doi:10.1158/2159-8290.CD-14-1156)

SchlacherKChristNSiaudNEgashiraAWuHJasinM2011Double-strand break repair-independent role for BRCA2 in blocking stalled replication fork degradation by MRE11. Cell145529542. (doi:10.1016/j.cell.2011.03.041)

SchlacherKWuHJasinM2012A distinct replication fork protection pathway connects Fanconi anemia tumor suppressors to RAD51-BRCA1/2. Cancer Cell22106116. (doi:10.1016/j.ccr.2012.05.015)

SchroederTMAnschützFKnoppA1964[Spontaneous chromosome aberrations in familial panmyelopathy]. Humangenetik1194196.

SchwabRANieminuszczyJShahFLangtonJMartinezDLLiangC-CCohnMAGibbonsRJDeansAJNiedzwiedzW2015The Fanconi anemia pathway maintains genome stability by coordinating replication and transcription. Molecular Cell60351361. (doi:10.1016/j.molcel.2015.09.012)

SealSBarfootRJayatilakeHSmithPRenwickABascombeLMcGuffogLEvansDGEcclesDEastonDF2003Evaluation of Fanconi anemia genes in familial breast cancer predisposition. Cancer Research6385968599.

ShahidTSorokaJKongEHMalivertLMcilwraithMJPapeTWestSCZhangX2014Structure and mechanism of action of the BRCA2 breast cancer tumor suppressor. Nature Structural & Molecular Biology21962968. (doi:10.1038/nsmb.2899)

ShamseldinHEElfakiMAlkurayaFS2012Exome sequencing reveals a novel Fanconi group defined by XRCC2 mutation. Journal of Medical Genetics49184186. (doi:10.1136/jmedgenet-2011-100585)

ShivjiMKKMukundSRRajendraEChenSShortJMSavillJKlenermanDVenkitaramanAR2009The BRC repeats of human BRCA2 differentially regulate RAD51 binding on single- versus double-stranded DNA to stimulate strand exchange. PNAS1061325413259. (doi:10.1073/pnas.0906208106)

SinghTRBakkerSTAgarwalSJansenMGrassmanEGodthelpBCAliAMDuC-HRooimansMAFanQ2009Impaired FANCD2 monoubiquitination and hypersensitivity to camptothecin uniquely characterize Fanconi anemia complementation group M. Blood114174180. (doi:10.1182/blood-2009-02-207811)

SinghTRAliAMParamasivamMPradhanAWahengbamKSeidmanMMMeeteiAR2013ATR-dependent phosphorylation of FANCM at serine 1045 is essential for FANCM functions. Cancer Research7343004310. (doi:10.1158/0008-5472.CAN-12-3976)

SmogorzewskaAMatsuokaSVinciguerraPMcDonaldERHurovKELuoJBallifBAGygiSPHofmannKD’AndreaAD2007Identification of the FANCI protein, a monoubiquitinated FANCD2 paralog required for DNA repair. Cell129289301. (doi:10.1016/
j.cell.2007.03.009)

SonodaESasakiMSBuersteddeJ-MBezzubovaOShinoharaAOgawaHTakataMYamaguchi-IwaiYTakedaS1998Rad51-deficient vertebrate cells accumulate chromosomal breaks prior to cell death. EMBO Journal17598608. (doi:10.1093/emboj/17.2.598)

SoulierJLeblancTLargheroJDastotHShimamuraAGuardiolaPEsperouHFerryCJubertCFeugeasJ-P2005Detection of somatic mosaicism and classification of Fanconi anemia patients by analysis of the FA/BRCA pathway. Blood10513291336. (doi:10.1182/blood-2004-05-1852)

StricklandKCHowittBEShuklaSARodigSRitterhouseLLLiuJFGarberJEChowdhuryDWuCJD’AndreaAD2016Association and prognostic significance of BRCA1/2-mutation status with neoantigen load, number of tumor-infiltrating lymphocytes and expression of PD-1/PD-L1 in high grade serous ovarian cancer. Oncotarget71358713598. (doi:10.18632/oncotarget.7277)

SySMHHuenMSYChenJ2009PALB2 is an integral component of the BRCA complex required for homologous recombination repair. PNAS10671557160. (doi:10.1073/pnas.0811159106