Abstract

Fanconi anemia (FA) represents a paradigm of rare genetic diseases, where the quest for cause and cure has led to seminal discoveries in cancer biology. Although a total of 16 FA genes have been identified thus far, the biochemical function of many of the FA proteins remains to be elucidated. FA is rare, yet the fact that 5 FA genes are in fact familial breast cancer genes and FA gene mutations are found frequently in sporadic cancers suggest wider applicability in hematopoiesis and oncology. Establishing the interaction network involving the FA proteins and their associated partners has revealed an intersection of FA with several DNA repair pathways, including homologous recombination, DNA mismatch repair, nucleotide excision repair, and translesion DNA synthesis. Importantly, recent studies have shown a major involvement of the FA pathway in the tolerance of reactive aldehydes. Moreover, despite improved outcomes in stem cell transplantation in the treatment of FA, many challenges remain in patient care.

Introduction

For many years, Fanconi anemia (FA) was merely acknowledged as a clinical rarity whose biological significance was not appreciated. It was understood that FA was a genetic disease of bone marrow failure, hypersensitivity to cross-linking agents, and high risk of acute myeloid leukemia. Over the last several decades, care for patients with FA has drastically improved with the advent of better blood banking, greater success in stem cell transplant, and increased recognition of the scope of care needed for FA patients. However, only with the identification of the first FA gene, FANCC, 22 years ago did the science begin to catch up with the descriptive studies of how patients and cells derived from these patients behaved.

Now, with the characterization of 16 complementation group genes, including 5 familial breast cancer genes, a better sense of the biology of FA has emerged. First, the pathway is composed of an upstream grouping of 8 proteins termed the core complex, whose primary activity appears to be an E3 ubiquitin ligase executed through one of its member subunits, FANCL. Its primary target is a heterodimer, FANCI-FANCD2 (ID2), which both become monoubiquitinated on DNA damage or during S phase of the cell cycle. Downstream members of the FA pathway are then activated in a way that is much more complex, some entailing physical interactions with FANCD2 but many resulting in coordination of direct DNA repair pathways that have a different web like character rather than the more linear pathway converging on ID2. Although classically termed a disease of interstrand cross-link hypersensitivity, FA is clearly more than that, requiring the coordination of translesion synthesis, homologous recombination, and mismatch repair in an effort to solve the Gordian knot of DNA lesion and replication fork.

In this review, we will lay out the clinical picture of FA along with the current state of care for FA patients. We will also detail the complex web of how the FA pathway orchestrates the DNA damage response to repair DNA lesions.

Clinical features of FA

Studies of FA have led to insights into bone marrow failure syndromes and common cancers. However, FA still holds many mysteries as to how mutations in the FA proteins contribute to the pathophysiology of birth defects, bone marrow failure, and cancer.1-3 

Most FA patients ultimately have bone marrow failure, with ∼90% of patients exhibiting this as their first hematopoietic presentation of disease. Classically, FA patients present with congenital defects, such as malformed or absent thumbs, absent radii, short stature, and microcephaly, and subtle but abnormal facies. A much longer list of less familiar and nonspecific characteristics may be present in patients (Table 1). Strikingly, a significant percentage of all FA patients, up to a third, exhibit none of these features, which is why hematologists, especially pediatric, routinely test for FA in bone marrow failure patients despite the lack of physical findings. The literature is therefore rife with adult patients diagnosed with FA when being treated for head and neck cancer and who exhibit inordinate toxicity as a result.4,5 

Table 1

Physical abnormalities in FA patients

Physical abnormalityPercent of FA patients
Skin discolorations (café au lait) 55 
Hand, arm, and other skeletal abnormalities, including thumb (missing thumb/radius) 51 
Abnormal reproductive organs (hypogenitalia, micropenis) 35 
Microcephaly or microophthalmia 26 
Kidney problems 21 
Low birth weight 11 
Heart defects 
Gastrointestinal problems (bowel) (atresia, imperforate anus) 
Physical abnormalityPercent of FA patients
Skin discolorations (café au lait) 55 
Hand, arm, and other skeletal abnormalities, including thumb (missing thumb/radius) 51 
Abnormal reproductive organs (hypogenitalia, micropenis) 35 
Microcephaly or microophthalmia 26 
Kidney problems 21 
Low birth weight 11 
Heart defects 
Gastrointestinal problems (bowel) (atresia, imperforate anus) 

Data from Alter.125 

When a potential FA patient presents with evidence of a hematopoietic production defect, a bone marrow aspiration and biopsy is in order to confirm bone marrow failure. Typically, a chromosomal breakage assay is also performed using peripheral blood lymphocytes with metaphase analysis in the presence of the DNA cross-linking agent diepoxybutane or mitomycin C (MMC).4,5  FA cells would demonstrate increased breakage and radial formation in the absence and presence of drug. FA diagnosis is aided greatly by sequencing analysis using an algorithm based on the relative unequal distribution of genetic subtypes (Table 2). FA cells are prone to somatic mosaicism as a result of genetic reversion, in which formerly mutant cells acquire additional mutations, resulting in diepoxybutane resistance, most likely as a result of selective pressure in the bone marrow. Such cases along with clinical suspicion necessitate additional testing of fibroblasts obtained through skin biopsy.6,7 

Table 2

Fanconi anemia genes and proteins

GroupGeneChromosomeMW (kDa)MotifsPercent of FA patientsNecessary for FANCD2-ub
FANCA 16q24.3 163 2 NLSs, 5 NESs 60-70 Yes 
FANCB Xp22.31 95 NLS Yes 
FANCC 9q22.3 63 None 14 Yes 
D1 FANCD1/BRCA2 13q12.13 380 8 BRC repeats, HD, 3 OBs, TD No 
D2 FANCD2 3p25.3 155, 162 None Yes 
FANCE 6p21-22 60 2 NLSs Yes 
FANCF 11p15 42 None Yes 
FANCG/XRCC9 9p13 68 7 TPRs 10 Yes 
FANCI/KIAA1794 15q25-26 146 None Yes 
FANCJ/BRIP1/BACH1 17q22-24 130 ATPase, 7 helicase motifs No 
FANCL/PHF9 2p16.1 43 3 WD40s, PHD 0.2 Yes 
FANCM 14q21.3 250 7 helicase motifs, degenerate endonuclease domain, ATPase 0.2 Yes 
FANCN/PALB2 16p12 130 2 WD40s 0.7 No 
FANCO/RAD51C 17q25.1 42 RAD51 paralog/recombinase 0.2 No 
FANCP/SLX4 16p13.3 200 Endonuclease assembly 0.2 No 
FANCQ/ERCC4/XPF 16p13.12 101 Endonuclease 0.5-1.0 No 
GroupGeneChromosomeMW (kDa)MotifsPercent of FA patientsNecessary for FANCD2-ub
FANCA 16q24.3 163 2 NLSs, 5 NESs 60-70 Yes 
FANCB Xp22.31 95 NLS Yes 
FANCC 9q22.3 63 None 14 Yes 
D1 FANCD1/BRCA2 13q12.13 380 8 BRC repeats, HD, 3 OBs, TD No 
D2 FANCD2 3p25.3 155, 162 None Yes 
FANCE 6p21-22 60 2 NLSs Yes 
FANCF 11p15 42 None Yes 
FANCG/XRCC9 9p13 68 7 TPRs 10 Yes 
FANCI/KIAA1794 15q25-26 146 None Yes 
FANCJ/BRIP1/BACH1 17q22-24 130 ATPase, 7 helicase motifs No 
FANCL/PHF9 2p16.1 43 3 WD40s, PHD 0.2 Yes 
FANCM 14q21.3 250 7 helicase motifs, degenerate endonuclease domain, ATPase 0.2 Yes 
FANCN/PALB2 16p12 130 2 WD40s 0.7 No 
FANCO/RAD51C 17q25.1 42 RAD51 paralog/recombinase 0.2 No 
FANCP/SLX4 16p13.3 200 Endonuclease assembly 0.2 No 
FANCQ/ERCC4/XPF 16p13.12 101 Endonuclease 0.5-1.0 No 

HD, helical domain; MW, molecular weight; NES, nuclear export signal; NLS, nuclear localization sequence; OB, oligonucleotide binding; PHD, plant homeo domain; TD, tower domain; TPR, tetratricopeptide repeat.

The vast majority of FA patients present in childhood when hematopoietic disease, be it bone marrow failure or acute myeloid leukemia, predominates. The key treatment of such patients has been stem cell transplantation (SCT).8  Because of the difficult nature of acute myelogenous leukemia therapy, especially in the wake of DNA damage hypersensitivity in FA, the best outcome results from institution of SCT prior to evolution of malignancy or neutropenia-associated infections, including bacterial and fungal. Conversely, application of SCT in the first decade is associated with significant morbidity. Nonetheless, aggressive use of SCT has resulted in an increase in survival of FA patients.9  Use of matched unrelated donors, long associated with increased risk of graft-versus-host disease, has seen marked improvement in recent years even in vulnerable populations such as FA patients. Concomitant use of milder conditioning regimens has lowered toxicity dramatically, thus ameliorating the inherent DNA damage hypersensitivity. Institution of drugs such as fludaribine has lowered the risk of graft failure, which historically has been problematic with a prevalence of up to 10%.10,11  As a result of these general improvements, outcomes for matched related donor transplants are often >80%, whereas those for matched unrelated cases are steadily improving. Late effects of SCT, namely growth delay, endocrinopathies, and second cancers, are magnified in FA patients, for whom such phenomena occur already at increased rates over the general population.

The use of modern blood banking has resulted in supportive care that enables patients to tolerate anemia and thrombocytopenia. The specter of infection remains, however, with respect to neutropenia, despite the use of granulocyte stimulating growth factor, as the use of such a growth factor promotes the evolution of clones that eventually lead to leukemia.12,13  Androgens have also been an adjunct to care and have demonstrated efficacy in FA patients, but virilization and higher risk of liver adenomas have limited their use.

There is good evidence that bone marrow failure in FA patients stems from hematopoietic stem cell (HSC) dysfunction and depletion of the HSC reservoir. The observation that CD34+ cells counts are low in FA patients supports this idea.14,15  The progressive HSC failure in FA patients is linked to the DNA damage response mediated by p53/p21.15  Reports have recently showed that knockdown of FA genes in human embryonic stem cells resulted in defective hematopoiesis, thus implicating the FA pathway in hematopoietic development.16,17  In this same study, FANCA or FANCD2 knockdown caused a significant reduction in the production of HSCs and progenitor cells on in vitro differentiation.

FA genetics

The FA pathway is genetically complex, comprised of 16 complementation groups and associated genes. The encoded proteins have been grouped into 3 categories: (1) the FA core complex, including the E3 ligase, FANCL; (2) the ID2 complex, the substrate for the E3 ubiquitin ligase activity of the core complex; and (3) downstream proteins that possess a DNA repair or damage tolerance function7,18  (Figure 1).

Figure 1

Mechanism of ICL repair in the FA pathway on collision of a replication fork with an ICL. The FA pathway is composed of ≥16 genes (A, B, C, D1, D2, E, F, G, I, K, L, M, N, O, P, and Q). The encoded proteins can be subdivided within the FA pathway into 3 groups: (1) proteins that make up the core complex; (2) the FANCI and FANCD2 proteins, which compose the ID2 complex; and (3) downstream effector proteins. (A) The FA pathway is activated during S phase of the cell cycle or on the detection of ICLs and DNA damage caused by other agents, including endogenous acetaldehydes. The FA core complex is recruited to the damage site through its interaction with the MHF1-MHF2-FANCM complex. (B) The ID2 complex becomes monoubiquitinated and remains associated with the DNA damage. The B-L-100 complex mediates the ubiquitination reaction, with the other 2 core subcomplexes (A-G-20 and C-E-F) playing accessory roles that remain to be elucidated. (C) Specialized endonucleases, in particular XPF/FANCQ-ERCC1 in complex with SLX4/FANCP, incise the DNA. (D) Within chromatin, the monoubiquitinated ID2 complex recruits DNA repair proteins including BRCA1, BRCA2/FANCD1, FANCJ, PALB2/FANCN, and RAD51C/FANCO. (E) Following successful repair, deubiquitination of the ID2 complex by USP1-UAF1 promotes its release from chromatin.

Figure 1

Mechanism of ICL repair in the FA pathway on collision of a replication fork with an ICL. The FA pathway is composed of ≥16 genes (A, B, C, D1, D2, E, F, G, I, K, L, M, N, O, P, and Q). The encoded proteins can be subdivided within the FA pathway into 3 groups: (1) proteins that make up the core complex; (2) the FANCI and FANCD2 proteins, which compose the ID2 complex; and (3) downstream effector proteins. (A) The FA pathway is activated during S phase of the cell cycle or on the detection of ICLs and DNA damage caused by other agents, including endogenous acetaldehydes. The FA core complex is recruited to the damage site through its interaction with the MHF1-MHF2-FANCM complex. (B) The ID2 complex becomes monoubiquitinated and remains associated with the DNA damage. The B-L-100 complex mediates the ubiquitination reaction, with the other 2 core subcomplexes (A-G-20 and C-E-F) playing accessory roles that remain to be elucidated. (C) Specialized endonucleases, in particular XPF/FANCQ-ERCC1 in complex with SLX4/FANCP, incise the DNA. (D) Within chromatin, the monoubiquitinated ID2 complex recruits DNA repair proteins including BRCA1, BRCA2/FANCD1, FANCJ, PALB2/FANCN, and RAD51C/FANCO. (E) Following successful repair, deubiquitination of the ID2 complex by USP1-UAF1 promotes its release from chromatin.

FA patient cells exhibit hypersensitivity to agents that cause interstrand DNA cross-links (ICLs), visualized as chromosomal fragility and radial formation that represent the biological hallmark of the disease. The more recent unraveling of the downstream proteins, namely, BRCA2/FANCD1, BACH1/FANCJ, PALB2/FANCN, RAD51C/FANCO, SLX4/FANCP, and XPF/FANCQ, reveals an intimate link to DNA repair and mainstream cancer biology, including breast cancer.19,20  Such links have revealed opportunities for improved cancer therapy by capitalizing on FA biology, as in the use of polyadenosine ribose polymerase inhibitors in BRCA2 mutant breast cancer.21  This is especially apropos, as FA patients who survive to adulthood display markedly higher rates of nonhematologic cancers, such as breast, head and neck, and squamous cell cancers. In addition, sporadic mutations in FA genes have been reported in many common adult cancers, including pancreatic, lung, gastrointestinal, and squamous cell cancers.22-24  Recent data indicate a direct link between FANCD2 regulation of the transcription of the Tap63 tumor suppressor and squamous cell cancer formation.25 

We focus below on FA protein functions and their interacting partners, followed by a mechanistic model of DNA ICL repair. Such a model may also be more generalizable to replication fork block and collapse. We also summarize recent developments that link the FA pathway to counteracting the genotoxicity of reactive aldehydes that are byproducts of cellular metabolism.

Upstream: the FA core complex

The FA core complex is comprised of 3 subcomplexes: (1) FANCL, FANCB, and FAAP100 (FA-associated protein 100 kDa); (2) FANCA, FANCG, and FAAP20 (FA-associated protein 20 kDa); and (3) FANCC, FANCE, and FANCF26-29  (Figure 1). FANCL is an E3 ubiquitin ligase that in conjunction with the E2 conjugating enzyme UBE2T monoubiquitinates FANCD2 and FANCI at K561 and K523, respectively. Purified FANCL and UBE2T can ubiquitinate both FANCD2 and FANCI site specifically in vitro,30,31  and its activity and substrate specificity are enhanced in the context of the FANCL-FANCB-FAAP100 (the L-B-100) complex.28  In cells, all components of the core complex are required for optimal ID2 ubiquitination, although careful epistasis analysis of mutant cells has revealed that only the L-B-100 complex is absolutely required.28,29  FA proteins in general lack identifiable functional domains and are highly conserved only in vertebrates, so progress has been slow in establishing their biochemical functions. Nevertheless, multiple groups have established a preliminary network of interactions among them32  (Figure 1).

X-ray crystal structures suggest that FANCD2 binds 2 interfaces in the core complex, with contact to FANCL and FANCE.33,34  The FANCF structure suggests a flexible protein that links the FANCC-FANCE and FANCA-FANCG subcomplexes and mediates recruitment of the entire complex to sites of damage and FANCD2.29,35,36 

Another core complex protein FANCA stimulates the activity of MUS81-EME1, a structure-specific nuclease involved in ICL repair37  (see below). FANCA also binds core complex members FANCG38  and FAAP20,39-41  which is necessary for normal FANCD2 ubiquitination and foci formation, mediated via its UBZ ubiquitin-binding domain. The UBZ domain also has been reported to mediate the interaction between FAAP20 and the translesion synthesis (TLS) protein REV1.42 

Nexus: the FANCI-FANCD2 (ID2) complex

Monoubiquitination of FANCD2 and FANCI signals activation of the FA DNA repair network and is required for ICL resistance, which is manifest by replication fork collapse. Formation of the heterodimeric ID2 complex is necessary for FANCD2 monoubiquitination and localization of the complex to chromatin at DNA damage foci. ID2 ubiquitination occurs mainly in S-phase.43-46 

The precise molecular function of FANCD2 and FANCI remains elusive, but biochemical analyses revealed that FANCD2, FANCI, and the ID2 complex possess DNA binding activity.45  Structural analysis of the ID2 complex suggests conformational changes inducible by interaction with DNA as well as core complex subcomplexes, such as L-B-100.47  An interplay of phosphorylation and presence of the entire array of core complex members is necessary for full ubiquitination of both members of the ID2 complex.30,31,48 

The removal of ubiquitin from the ID2 complex on completion of replication fork repair and restart is catalyzed by the USP1-UAF1 deubiquitinase (DUB), which also targets other proteins such as ubiquitinated PCNA.49,50  USP1, the catalytic component, is stimulated by heterodimerization with the accessory protein UAF1.51,52  Negative regulation of this DUB occurs on DNA damage signaling via suppression of USP1 transcription, in addition to cleavage and proteolytic degradation of the USP1 protein.53,54  Because the timely deubiquitination of the ID2 complex is necessary for the functional integrity of the FA pathway, USP1-UAF1 has emerged as an important target for developing small molecule inhibitors that can potentiate the cytotoxic effect of ICL-inducing chemotherapeutics.55  In addition, both murine and chicken cells deficient for either UAF1 or USP1 exhibit a defect in DNA double-strand break repair by homologous recombination (HR),56-58  suggesting a HR role for USP1-UAF1 as well.

FANCM and its binding partners: regulation of replication restart

FANCM is a core complex member with an ATP-dependent DNA translocase activity. FANCM is required for resistance to ICLs via mediation of (1) recruitment of the FA core complex to chromatin59 ; (2) regression of stalled DNA replication forks; (3) traversal of the DNA replication machinery across an ICL60 ; and (4) efficient DNA damage signaling via the ATR kinase.61-63  Notably, the DNA translocase activity is not required for FANCD2 ubiquitination.64  Such activities are deemed necessary for the ability of replication to traverse an ICL using replication fork regression.65  Regression facilitates bypass of lesions in the template DNA strand,66  but how it promotes ICL removal remains unclear.

Replication restart also requires the BLM helicase, which is deficient in Bloom’s syndrome, yet another way that FA interacts with a distinct repair pathway. Bloom’s syndrome is marked by cancer susceptibility and by cells that display increased sister chromatid exchange and exhibits several FA-like features. BLM regulates HR by promoting the formation of noncrossovers during homologous recombination, and it also catalyzes replication fork regression.67  Importantly, FANCM and BLM form a complex, being tethered via RMI1-RMI2, and their interaction is required for resistance to ICLs.36,68 

The MHF complex (MHF1-MHF2-FAAP24), structurally similar to histones, is important for FANCM stability, recruitment of the FA core complex to damaged chromatin, FANCD2 ubiquitination, and cellular resistance to ICLs.69,70  The MHF complex also stimulates the DNA binding and translocase activities of FANCM.71,72  Based on the crystal structure of the binary complex of MHF and DNA, it has been suggested that the MHF complex serves to anchor FANCM at a DNA junction to promote replication fork regression and DNA branch migration reactions.70  FAAP24 also binds DNA and interacts with FANCM directly. The DNA binding activity of FAAP24 is required for resistance to ICLs, as well as for FANCD2 ubiquitination.73 

Downstream: effectors of the FA pathway

Biallelic mutations in BRCA2/FANCD1, BACH1/FANCJ, PALB2/FANCN, RAD51C/FANCO, SLX4/FANCP, and XPF/FANCQ, which all play a role in known DNA repair reactions, can lead to FA.74  Because a deficiency in these proteins does not affect FANCD2 ubiquitination, they are characterized as downstream of FANCD2 in ICL removal.46  Conversely, FANCD2 has been found in complex and in repair foci with several of these proteins, suggesting a more direct functional link.

BRCA2, PALB2, and RAD51C (3 known familial breast cancer gene products) all have defined functions in HR.75  Specifically, BRCA2 and PALB2 associate with RAD51, the recombinase that catalyzes the HR reaction, and stimulate its activity.75,76  A paralog of RAD51, RAD51C, has been identified as a FA gene, FANCO, and is found in multiple subcomplexes, all with defined roles in HR.77-79  A subcomplex of RAD51B-RAD51C enhances RAD51-mediated DNA strand exchange in vitro.80 

BACH1/FANCJ, another familial breast cancer and FA-J complementation group protein, is a DNA helicase that translocates on ssDNA with a 5′ to 3′ polarity. FANCJ also binds to the DNA mismatch repair protein MLH1,81  which interacts with FAN1 (see below) and FANCD2,82  supporting a role for mismatch repair in FA.

SLX4 acts as a nuclear scaffold to enhance the activity of 3 structure-specific nucleases previously implicated in ICL repair and other DNA repair pathways.83,84  These nucleases are (1) XPF/FANCQ-ERCC1 that functions in nucleotide excision repair85,86 ; (2) MUS81-EME1; and (3) SLX1. Each of these nucleases possesses a substrate specificity consistent with a role in the unhooking and removal of ICLs.87,88  Recent studies involving the use of Xenopus cell-free extracts have provided biochemical evidence for a major role of XPF-ERCC1-SLX4 in ICL removal.89 

FAN1, which possesses 5′ flap endonuclease and 5′ exonuclease activities, was recently identified as a binding partner of monoubiquitinated FANCD2.90  Abrogation of FAN1 nuclease activity results in hypersensitivity to ICL agents. It has been suggested that FAN1 is recruited by ubiquitinated FANCD2 to help mediate ICL incision and removal.90  Interestingly, mutations in FAN1 do not cause FA but underlie the kidney disease karyomegalic interstitial nephritis.91 

MUS81-EME1 and SLX1-SLX4 have been shown to cooperatively cleave the Holliday junction, an important DNA intermediate in HR.92  However, this function may be independent of the role of these nucleases in ICL removal.93 

Translesion DNA synthesis polymerases and FA

ICL unhooking at a stalled replication fork results in a double-strand break in 1 chromatid and a single-stranded gap harboring the ICL adduct on the other (Figure 1). Repair of the strand break occurs via HR,94  whereas gap filling in the lesion-containing DNA requires TLS, being catalyzed by a specialized DNA polymerase. Several TLS polymerases have been implicated in this regard, although for replication-dependent ICL repair, the most important appears to be the REV1-Polζ complex.95  In ICL repair, a mechanism that relies on interaction of REV1 with FAAP20, an FA core complex member, is thought to mediate the recruitment of REV1-Polζ.42 

Cell extracts use to examine the removal of a site-specific cross-link has led to a mechanistic model for ICL repair when 2 convergent replication forks encounter the DNA cross-link96  (Figure 1). In this system, replication fork stalling occurs momentarily ∼20 to 40 nucleotides from the lesion, followed by disassembly of the machinery, and then by polymerase stalling again at 1 nucleotide from the lesion. At this stage, unhooking incisions, requiring XPF/ERCC1 and SLX4,89  occur on the parental strand opposite to that of the approaching leading strand (Figure 1). Finally, nucleotide insertion across the adducted base by REV1-Polζ occurs. Following the unhooking and TLS events, coordinated completion of repair requires actions of HR and mismatch repair.

FA pathway and cytokinesis

Several reports have suggested that FA proteins play important roles during M phase, especially in cytokinesis.97-100  First, FANCD2 and FANCI were found to localize at ultrafine anaphase bridges between segregating sister chromatids; these bridges increase in frequency on replication stress.99,100  The number of anaphase bridges, which are normally decorated with BLM and FANCM, increases in FA-deficient cells, leading to a higher frequency of cytokinesis failure, binucleated cells, as well as supernumerary centrosomes. In addition, several FA proteins localize to centrosomes and the mitotic spindle.98,101 

DNA damage response and FA: ATM, ATR, and CHK1

The FA pathway influences DNA damage signaling through the ataxia telangiectasia mutated (ATM), ataxia telangiectasia and Rad3-related (ATR),102  and Chk1 kinases. Many of the FA proteins are phosphorylated in response to DNA damage and during distinct phases of the cell cycle. Similar to ATM-mutant cells, FANCD2-deficient cells display a defect in S phase checkpoint response after ionizing radiation (IR) exposure.103  Normally, FANCD2 is phosphorylated by ATM on serine 222 on IR treatment, and this event is required for establishing the intra-S checkpoint and a proper cellular response to DSBs.103  These results thus reveal a direct interaction between the ATM and FA pathways.

The FA pathway also intersects with ATR-mediated checkpoint signaling. Following exposure to a DNA cross-linking agent, FANCD2 and ATR colocalize in nuclear foci.104  Phosphorylation of FANCD2 on threonine 691 and serine 717 by ATR is required for FANCD2 monoubiquitination and correction of MMC sensitivity in FANCD2-deficient cells.105  ATR also phosphorylates FANCG106  and FANCI.48  FANCG phosphorylation by ATR is important for the functional interaction of FANCG with BRCA2/FANCD1107 . Interestingly, downregulation of FANCM or FAAP24 negatively affects ATR-mediated checkpoint signaling.62  Clinically, Seckel’s syndrome, stemming from biallelic mutations in ATR, resembles FA in the exhibition of chromosome fragility, developmental delay, growth retardation, and cancer susceptibility.

CHK1, a substrate of both ATM and ATR, phosphorylates FANCE on threonine 346 and serine 374, and these modifications are required for the cellular resistance to MMC without affecting FANCD2 monoubiquitination and focus formation.102  CHK1 is also implicated in the serine 331 phosphorylation of FANCD2, which is required for resistance to MMC and FANCD2 monoubiquitination.108  Increased CHK1 activity is associated with early marrow failure, and downregulation of such activity may promote later leukemogenesis.109 

Function of the FA pathway in alleviating the genotoxicity of acetaldehyde

Recent work has implicated aldehyde metabolism as a primary cause of the FA phenotype.110,111  Chicken DT40 cells, ablated for one of several FA genes, including FANCB, FANCC, FANCL, and BRCA2/FANCD1, are hypersensitive to acetaldehyde.110  In addition, these FA gene knockouts are synthetic lethal with mutations in the formaldehyde catabolism gene ADH5, indicating that cells become sensitized to aldehyde toxicity when the FA pathway is defective.110  Notably, mice doubly deficient in Fancd2 and aldehyde dehydrogenase (Aldh2) are prone to ethanol-induced bone marrow failure compared with wild-type mice or the single mutants.110  The bone marrow failure in Fancd2−/−:Aldh2−/− mice correlates with the accumulation of damaged DNA within the hematopoietic stem and progenitor cell pool.111  A genotypic analysis of a group of Japanese FA patients revealed that ALDH2 deficiency dramatically accelerates bone marrow failure and increases the frequency of malformation in some tissues, providing additional evidence that reactive aldehydes play an important role in the pathogenesis of FA.112  However, it remains unclear if aldehydes cause FA-associated genotoxicity. Aldehydes may make FA patients clinically worse, but no evidence yet exists that antialdehyde therapy could benefit them, although the possibility is intriguing.

FA pathway, cytokine sensitivity, and oxidative stress

FA patients exhibit altered expression levels of growth factors and cytokines, including tumor necrosis factor-α (TNF-α), which is involved in the initiation of apoptosis. Studies using Fanca−/−, Fancc−/−, and Fancg−/− mouse models have demonstrated that FA cells are hypersensitive to TNF-α, and this sensitivity contributes to bone marrow failure in FA.113  Expression of Fancc cDNA in Fancc−/− stem cells prevents the formation of leukemic clonal outgrowths, implying that FANCC is crucial for proper cellular response to TNF-α. FANCD2 represses the transcription of TNF-α by binding to its promoter region,114  which may explain why FA patients have elevated TNF-α levels. Oxidative DNA damage level is persistently higher in HSC or progenitor cells from TNF-α–injected Fancc−/− mice, further supporting the notion that FA proteins protect against reactive oxygen species-induced DNA damage.115 

Hematopoietic stem cells with mutant FANCC are hypersensitive to interferon-γ,116  and interferon-γ stimulates increased apoptosis in the mutant setting.117  Reconstitution of the myeloid compartment appears to depend on interferon response pathways.118  FANCC also functions as a negative regulator of cytokine-induced apoptosis by modulating the activity of PKR, a growth inhibitory kinase and key effector of apoptosis.119 

Original descriptions of FA cell hypersensitivity to oxygen have led to the hypothesis that FA harbors primary defects in management of oxidative stress, and more recent evidence supports this idea. For example, FANCA and FANCG are sensitive to redox conditioning, such that hydrogen peroxide treatment triggers complex formation of these FA proteins.120  The antioxidant tempol displayed tumor onset and protective effects against oxidative damage in Fancd2−/− mice. Low oxygen has recently been shown to stimulate ID2 ubiquitination in an ATR-dependent manner.121 

The FA pathway has also been linked to mitochondrial dysfunction. A recent study has revealed excessive formation of mitochondrial reactive oxygen species in FA (FA-A, -C, and -D2) cells,122  and mice doubly deficient in Fancc and superoxide dismutase display bone marrow hypocellularity, which is not present in mice with either of the mutations.123  FA-deficient cells exhibit better growth characteristics under hypoxic conditions, and the use of low oxygen tension allows the generation of FA-deficient induced pluripotent stem cell lines.124 

Summary

Although a tremendous amount of information has emerged in the 22 years since the cloning of the first FA gene, FANCC, much remains to be learned regarding how DNA repair, DNA damage checkpoints, and associated processes intersect within the FA pathway. Even though FA is rare, its general relevance to cancer biology is exemplified by ≥5 FA genes being familial breast cancer genes and the identification of somatic FA gene mutations in many common cancers.

Many challenges lie ahead in devising an effective treatment of FA. Although stem cell transplant overcomes the ravages of acute myelogenous leukemia, patients remain prone to many solid tumors, including breast, head and neck, genitourinary tumors, and second cancers after transplant. Second, stem cell transplant in FA patients is still fraught with complications, and less toxic, more efficacious modalities would be welcome. Third, as noted earlier, many common adult cancers exhibit FA gene mutations, suggesting that dysfunction in the FA pathway can contribute to oncogenic genomic instability. Such avenues of investigation promise to enhance medical care of FA and non-FA patients alike. For example, the elaboration of polyadenosine ribose polymerase inhibitors in BRCA1 and BRCA2, both central to FA biology, have led to clinical trials that help target tumors with mutations in these proteins. Understanding the biology of FA will lead to greater opportunities for therapeutic advantage.

Authorship

Contribution: All authors contributed to the writing and editing of the manuscript.

Conflict-of-interest disclosure: The authors declare no competing financial interests.

Correspondence: Gary M. Kupfer, LMP2073, Section of Pediatric Hematology-Oncology, Yale School of Medicine, 333 Cedar St, New Haven, CT 06510; e-mail: gary.kupfer@yale.edu.

References

References
1
Bagby
 
GC
Alter
 
BP
Fanconi anemia.
Semin Hematol
2006
, vol. 
43
 
3
(pg. 
147
-
156
)
2
D’Andrea
 
AD
Dahl
 
N
Guinan
 
EC
Shimamura
 
A
Marrow failure.
Hematology (Am Soc Hematol Educ Program)
2002
 
58-72
3
Chirnomas
 
SD
Kupfer
 
GM
The inherited bone marrow failure syndromes.
Pediatr Clin North Am
2013
, vol. 
60
 
6
(pg. 
1291
-
1310
)
4
Green
 
AM
Kupfer
 
GM
Fanconi anemia.
Hematol Oncol Clin North Am
2009
, vol. 
23
 
2
(pg. 
193
-
214
)
5
Alter
 
BP
Diagnosis, genetics, and management of inherited bone marrow failure syndromes.
Hematology (Am Soc Hematol Educ Program)
2007
 
29-39
6
Khincha
 
PP
Savage
 
SA
Genomic characterization of the inherited bone marrow failure syndromes.
Semin Hematol
2013
, vol. 
50
 
4
(pg. 
333
-
347
)
7
Kee
 
Y
D’Andrea
 
AD
Molecular pathogenesis and clinical management of Fanconi anemia.
J Clin Invest
2012
, vol. 
122
 
11
(pg. 
3799
-
3806
)
8
Gluckman
 
E
Wagner
 
JE
Hematopoietic stem cell transplantation in childhood inherited bone marrow failure syndrome.
Bone Marrow Transplant
2008
, vol. 
41
 
2
(pg. 
127
-
132
)
9
Farzin
 
A
Davies
 
SM
Smith
 
FO
, et al. 
Matched sibling donor haematopoietic stem cell transplantation in Fanconi anaemia: an update of the Cincinnati Children’s experience.
Br J Haematol
2007
, vol. 
136
 
4
(pg. 
633
-
640
)
10
Chaudhury
 
S
Auerbach
 
AD
Kernan
 
NA
, et al. 
Fludarabine-based cytoreductive regimen and T-cell-depleted grafts from alternative donors for the treatment of high-risk patients with Fanconi anaemia.
Br J Haematol
2008
, vol. 
140
 
6
(pg. 
644
-
655
)
11
Bitan
 
M
Or
 
R
Shapira
 
MY
, et al. 
Fludarabine-based reduced intensity conditioning for stem cell transplantation of Fanconi anemia patients from fully matched related and unrelated donors.
Biol Blood Marrow Transplant
2006
, vol. 
12
 
7
(pg. 
712
-
718
)
12
Myers
 
KC
Davies
 
SM
Hematopoietic stem cell transplantation for bone marrow failure syndromes in children.
Biol Blood Marrow Transplant
2009
, vol. 
15
 
3
(pg. 
279
-
292
)
13
MacMillan
 
ML
Wagner
 
JE
Haematopoeitic cell transplantation for Fanconi anaemia - when and how?
Br J Haematol
2010
, vol. 
149
 
1
(pg. 
14
-
21
)
14
Kelly
 
PF
Radtke
 
S
von Kalle
 
C
, et al. 
Stem cell collection and gene transfer in Fanconi anemia.
Mol Ther
2007
, vol. 
15
 
1
(pg. 
211
-
219
)
15
Ceccaldi
 
R
Parmar
 
K
Mouly
 
E
, et al. 
Bone marrow failure in Fanconi anemia is triggered by an exacerbated p53/p21 DNA damage response that impairs hematopoietic stem and progenitor cells.
Cell Stem Cell
2012
, vol. 
11
 
1
(pg. 
36
-
49
)
16
Tulpule
 
A
Lensch
 
MW
Miller
 
JD
, et al. 
Knockdown of Fanconi anemia genes in human embryonic stem cells reveals early developmental defects in the hematopoietic lineage.
Blood
2010
, vol. 
115
 
17
(pg. 
3453
-
3462
)
17
Kamimae-Lanning
 
AN
Goloviznina
 
NA
Kurre
 
P
Fetal origins of hematopoietic failure in a murine model of Fanconi anemia.
Blood
2013
, vol. 
121
 
11
(pg. 
2008
-
2012
)
18
Garaycoechea
 
JI
Patel
 
KJ
Why does the bone marrow fail in Fanconi anemia?
Blood
2014
, vol. 
123
 
1
(pg. 
26
-
34
)
19
Masserot
 
C
Peffault de Latour
 
R
Rocha
 
V
, et al. 
Head and neck squamous cell carcinoma in 13 patients with Fanconi anemia after hematopoietic stem cell transplantation.
Cancer
2008
, vol. 
113
 
12
(pg. 
3315
-
3322
)
20
Lowy
 
DR
Gillison
 
ML
A new link between Fanconi anemia and human papillomavirus-associated malignancies.
J Natl Cancer Inst
2003
, vol. 
95
 
22
(pg. 
1648
-
1650
)
21
Lee
 
JM
Ledermann
 
JA
Kohn
 
EC
 
PARP Inhibitors for BRCA1/2 mutation-associated and BRCA-like malignancies. Ann Oncol. 2014;25(1):32-40
22
Kennedy
 
RD
D’Andrea
 
AD
The Fanconi Anemia/BRCA pathway: new faces in the crowd.
Genes Dev
2005
, vol. 
19
 
24
(pg. 
2925
-
2940
)
23
Levitus
 
M
Joenje
 
H
de Winter
 
JP
The Fanconi anemia pathway of genomic maintenance.
Cell Oncol
2006
, vol. 
28
 
1-2
(pg. 
3
-
29
)
24
Valeri
 
A
Martínez
 
S
Casado
 
JA
Bueren
 
JA
Fanconi anaemia: from a monogenic disease to sporadic cancer.
Clin Transl Oncol
2011
, vol. 
13
 
4
(pg. 
215
-
221
)
25
Park
 
E
Kim
 
H
Kim
 
JM
, et al. 
FANCD2 activates transcription of TAp63 and suppresses tumorigenesis.
Mol Cell
2013
, vol. 
50
 
6
(pg. 
908
-
918
)
26
Ling
 
C
Ishiai
 
M
Ali
 
AM
, et al. 
FAAP100 is essential for activation of the Fanconi anemia-associated DNA damage response pathway.
EMBO J
2007
, vol. 
26
 
8
(pg. 
2104
-
2114
)
27
Medhurst
 
AL
Laghmani
 
H
Steltenpool
 
J
, et al. 
Evidence for subcomplexes in the Fanconi anemia pathway.
Blood
2006
, vol. 
108
 
6
(pg. 
2072
-
2080
)
28
Rajendra
 
E
Oestergaard
 
VH
Langevin
 
F
, et al. 
The genetic and biochemical basis of FANCD2 monoubiquitination.
Mol Cell
2014
, vol. 
54
 
5
(pg. 
858
-
869
)
29
Huang
 
YL
Justin
 
WC
Lowery
 
M
, et al. 
Modularized functions of the Fanconi anemia core complex.
Cell Reports
2014
, vol. 
7
 
6
(pg. 
1849
-
1857
)
30
Sato
 
K
Toda
 
K
Ishiai
 
M
Takata
 
M
Kurumizaka
 
H
DNA robustly stimulates FANCD2 monoubiquitylation in the complex with FANCI.
Nucleic Acids Res
2012
, vol. 
40
 
10
(pg. 
4553
-
4561
)
31
Longerich
 
SKY
Tsai
 
M-S
Hlaing
 
A
Kupfer
 
GM
Sung
 
P
Regulation of FANCD2 and FANCI ubiquitination by their interaction and by DNA.
Nucleic Acids Res
2014
, vol. 
42
 
9
(pg. 
5657
-
5670
)
32
Hodson
 
C
Walden
 
H
 
Towards a molecular understanding of the fanconi anemia core complex. Anemia. doi: 10.1155/2012/926787
33
Hodson
 
C
Cole
 
AR
Lewis
 
LP
Miles
 
JA
Purkiss
 
A
Walden
 
H
Structural analysis of human FANCL, the E3 ligase in the Fanconi anemia pathway.
J Biol Chem
2011
, vol. 
286
 
37
(pg. 
32628
-
32637
)
34
Nookala
 
RK
Hussain
 
S
Pellegrini
 
L
Insights into Fanconi Anaemia from the structure of human FANCE.
Nucleic Acids Res
2007
, vol. 
35
 
5
(pg. 
1638
-
1648
)
35
Kowal
 
P
Gurtan
 
AM
Stuckert
 
P
D’Andrea
 
AD
Ellenberger
 
T
Structural determinants of human FANCF protein that function in the assembly of a DNA damage signaling complex.
J Biol Chem
2007
, vol. 
282
 
3
(pg. 
2047
-
2055
)
36
Deans
 
AJ
West
 
SC
FANCM connects the genome instability disorders Bloom’s Syndrome and Fanconi Anemia.
Mol Cell
2009
, vol. 
36
 
6
(pg. 
943
-
953
)
37
Benitez
 
A
Yuan
 
F
Nakajima
 
S
, et al. 
Damage-dependent regulation of MUS81-EME1 by Fanconi anemia complementation group A protein.
Nucleic Acids Res
2014
, vol. 
42
 
3
(pg. 
1671
-
1683
)
38
Garcia-Higuera
 
I
Kuang
 
Y
Denham
 
J
D’Andrea
 
AD
The fanconi anemia proteins FANCA and FANCG stabilize each other and promote the nuclear accumulation of the Fanconi anemia complex.
Blood
2000
, vol. 
96
 
9
(pg. 
3224
-
3230
)
39
Ali
 
AM
Pradhan
 
A
Singh
 
TR
, et al. 
FAAP20: a novel ubiquitin-binding FA nuclear core-complex protein required for functional integrity of the FA-BRCA DNA repair pathway.
Blood
2012
, vol. 
119
 
14
(pg. 
3285
-
3294
)
40
Leung
 
JW
Wang
 
Y
Fong
 
KW
Huen
 
MS
Li
 
L
Chen
 
J
Fanconi anemia (FA) binding protein FAAP20 stabilizes FA complementation group A (FANCA) and participates in interstrand cross-link repair.
Proc Natl Acad Sci USA
2012
, vol. 
109
 
12
(pg. 
4491
-
4496
)
41
Yan
 
Z
Guo
 
R
Paramasivam
 
M
, et al. 
A ubiquitin-binding protein, FAAP20, links RNF8-mediated ubiquitination to the Fanconi anemia DNA repair network.
Mol Cell
2012
, vol. 
47
 
1
(pg. 
61
-
75
)
42
Kim
 
H
Yang
 
K
Dejsuphong
 
D
D’Andrea
 
AD
Regulation of Rev1 by the Fanconi anemia core complex.
Nat Struct Mol Biol
2012
, vol. 
19
 
2
(pg. 
164
-
170
)
43
Schlacher
 
K
Wu
 
H
Jasin
 
M
A distinct replication fork protection pathway connects Fanconi anemia tumor suppressors to RAD51-BRCA1/2.
Cancer Cell
2012
, vol. 
22
 
1
(pg. 
106
-
116
)
44
Wang
 
LC
Stone
 
S
Hoatlin
 
ME
Gautier
 
J
Fanconi anemia proteins stabilize replication forks.
DNA Repair (Amst)
2008
, vol. 
7
 
12
(pg. 
1973
-
1981
)
45
Garner
 
E
Smogorzewska
 
A
Ubiquitylation and the Fanconi anemia pathway.
FEBS Lett
2011
, vol. 
585
 
18
(pg. 
2853
-
2860
)
46
Moldovan
 
GL
D’Andrea
 
AD
How the fanconi anemia pathway guards the genome.
Annu Rev Genet
2009
, vol. 
43
 (pg. 
223
-
249
)
47
Joo
 
W
Xu
 
G
Persky
 
NS
, et al. 
Structure of the FANCI-FANCD2 complex: insights into the Fanconi anemia DNA repair pathway.
Science
2011
, vol. 
333
 
6040
(pg. 
312
-
316
)
48
Ishiai
 
M
Kitao
 
H
Smogorzewska
 
A
, et al. 
FANCI phosphorylation functions as a molecular switch to turn on the Fanconi anemia pathway.
Nat Struct Mol Biol
2008
, vol. 
15
 
11
(pg. 
1138
-
1146
)
49
Nijman
 
SM
Huang
 
TT
Dirac
 
AM
, et al. 
The deubiquitinating enzyme USP1 regulates the Fanconi anemia pathway.
Mol Cell
2005
, vol. 
17
 
3
(pg. 
331
-
339
)
50
Smogorzewska
 
A
Matsuoka
 
S
Vinciguerra
 
P
, et al. 
Identification of the FANCI protein, a monoubiquitinated FANCD2 paralog required for DNA repair.
Cell
2007
, vol. 
129
 
2
(pg. 
289
-
301
)
51
Cohn
 
MA
Kowal
 
P
Yang
 
K
, et al. 
A UAF1-containing multisubunit protein complex regulates the Fanconi anemia pathway.
Mol Cell
2007
, vol. 
28
 
5
(pg. 
786
-
797
)
52
Villamil
 
MA
Liang
 
Q
Chen
 
J
, et al. 
Serine phosphorylation is critical for the activation of ubiquitin-specific protease 1 and its interaction with WD40-repeat protein UAF1.
Biochemistry
2012
, vol. 
51
 
45
(pg. 
9112
-
9123
)
53
Huang
 
TT
Nijman
 
SM
Mirchandani
 
KD
, et al. 
Regulation of monoubiquitinated PCNA by DUB autocleavage.
Nat Cell Biol
2006
, vol. 
8
 
4
(pg. 
339
-
347
)
54
Piatkov
 
KI
Colnaghi
 
L
Békés
 
M
Varshavsky
 
A
Huang
 
TT
The auto-generated fragment of the Usp1 deubiquitylase is a physiological substrate of the N-end rule pathway.
Mol Cell
2012
, vol. 
48
 
6
(pg. 
926
-
933
)
55
Liang
 
Q
Dexheimer
 
TS
Zhang
 
P
, et al. 
A selective USP1-UAF1 inhibitor links deubiquitination to DNA damage responses.
Nat Chem Biol
2014
, vol. 
10
 
4
(pg. 
298
-
304
)
56
Murai
 
J
Yang
 
K
Dejsuphong
 
D
Hirota
 
K
Takeda
 
S
D’Andrea
 
AD
The USP1/UAF1 complex promotes double-strand break repair through homologous recombination.
Mol Cell Biol
2011
, vol. 
31
 
12
(pg. 
2462
-
2469
)
57
Park
 
E
Kim
 
JM
Primack
 
B
, et al. 
Inactivation of Uaf1 causes defective homologous recombination and early embryonic lethality in mice.
Mol Cell Biol
2013
, vol. 
33
 
22
(pg. 
4360
-
4370
)
58
Kim
 
JM
Parmar
 
K
Huang
 
M
, et al. 
Inactivation of murine Usp1 results in genomic instability and a Fanconi anemia phenotype.
Dev Cell
2009
, vol. 
16
 
2
(pg. 
314
-
320
)
59
Kim
 
JM
Kee
 
Y
Gurtan
 
A
D'Andrea
 
AD
Cell cycle-dependent chromatin loading of the Fanconi anemia core complex by FANCM/FAAP24.
Blood
2008
, vol. 
111
 
10
(pg. 
5215
-
5222
)
60
Huang
 
J
Liu
 
S
Bellani
 
MA
, et al. 
The DNA translocase FANCM/MHF promotes replication traverse of DNA interstrand crosslinks.
Mol Cell
2013
, vol. 
52
 
3
(pg. 
434
-
446
)
61
Blackford
 
AN
Schwab
 
RA
Nieminuszczy
 
J
Deans
 
AJ
West
 
SC
Niedzwiedz
 
W
The DNA translocase activity of FANCM protects stalled replication forks.
Hum Mol Genet
2012
, vol. 
21
 
9
(pg. 
2005
-
2016
)
62
Collis
 
SJ
Ciccia
 
A
Deans
 
AJ
, et al. 
FANCM and FAAP24 function in ATR-mediated checkpoint signaling independently of the Fanconi anemia core complex.
Mol Cell
2008
, vol. 
32
 
3
(pg. 
313
-
324
)
63
Huang
 
M
Kim
 
JM
Shiotani
 
B
Yang
 
K
Zou
 
L
D’Andrea
 
AD
The FANCM/FAAP24 complex is required for the DNA interstrand crosslink-induced checkpoint response.
Mol Cell
2010
, vol. 
39
 
2
(pg. 
259
-
268
)
64
Rosado
 
IV
Niedzwiedz
 
W
Alpi
 
AF
Patel
 
KJ
The Walker B motif in avian FANCM is required to limit sister chromatid exchanges but is dispensable for DNA crosslink repair.
Nucleic Acids Res
2009
, vol. 
37
 
13
(pg. 
4360
-
4370
)
65
Ray Chaudhuri
 
A
Hashimoto
 
Y
Herrador
 
R
, et al. 
Topoisomerase I poisoning results in PARP-mediated replication fork reversal.
Nat Struct Mol Biol
2012
, vol. 
19
 
4
(pg. 
417
-
423
)
66
Yeeles
 
JT
Poli
 
J
Marians
 
KJ
Pasero
 
P
Rescuing stalled or damaged replication forks.
Cold Spring Harb Perspect Biol
2013
, vol. 
5
 
5
pg. 
a012815
 
67
Manthei
 
KA
Keck
 
JL
The BLM dissolvasome in DNA replication and repair.
Cell Mol Life Sci
2013
, vol. 
70
 
21
(pg. 
4067
-
4084
)
68
Hoadley
 
KA
Xue
 
Y
Ling
 
C
Takata
 
M
Wang
 
W
Keck
 
JL
Defining the molecular interface that connects the Fanconi anemia protein FANCM to the Bloom syndrome dissolvasome.
Proc Natl Acad Sci USA
2012
, vol. 
109
 
12
(pg. 
4437
-
4442
)
69
Tao
 
Y
Jin
 
C
Li
 
X
, et al. 
The structure of the FANCM-MHF complex reveals physical features for functional assembly.
Nat Commun
2012
, vol. 
3
 pg. 
782
 
70
Zhao
 
Q
Saro
 
D
Sachpatzidis
 
A
, et al. 
The MHF complex senses branched DNA by binding a pair of crossover DNA duplexes.
Nat Commun
2014
, vol. 
5
 pg. 
2987
 
71
Singh
 
TR
Saro
 
D
Ali
 
AM
, et al. 
MHF1-MHF2, a histone-fold-containing protein complex, participates in the Fanconi anemia pathway via FANCM.
Mol Cell
2010
, vol. 
37
 
6
(pg. 
879
-
886
)
72
Yan
 
Z
Delannoy
 
M
Ling
 
C
, et al. 
A histone-fold complex and FANCM form a conserved DNA-remodeling complex to maintain genome stability.
Mol Cell
2010
, vol. 
37
 
6
(pg. 
865
-
878
)
73
Ciccia
 
A
McDonald
 
N
West
 
SC
Structural and functional relationships of the XPF/MUS81 family of proteins.
Annu Rev Biochem
2008
, vol. 
77
 (pg. 
259
-
287
)
74
Deans
 
AJ
West
 
SC
DNA interstrand crosslink repair and cancer.
Nat Rev Cancer
2011
, vol. 
11
 
7
(pg. 
467
-
480
)
75
San Filippo
 
J
Sung
 
P
Klein
 
H
Mechanism of eukaryotic homologous recombination.
Annu Rev Biochem
2008
, vol. 
77
 (pg. 
229
-
257
)
76
Dray
 
E
Etchin
 
J
Wiese
 
C
, et al. 
Enhancement of RAD51 recombinase activity by the tumor suppressor PALB2.
Nat Struct Mol Biol
2010
, vol. 
17
 
10
(pg. 
1255
-
1259
)
77
Somyajit
 
K
Subramanya
 
S
Nagaraju
 
G
RAD51C: a novel cancer susceptibility gene is linked to Fanconi anemia and breast cancer.
Carcinogenesis
2010
, vol. 
31
 
12
(pg. 
2031
-
2038
)
78
Vaz
 
F
Hanenberg
 
H
Schuster
 
B
, et al. 
Mutation of the RAD51C gene in a Fanconi anemia-like disorder.
Nat Genet
2010
, vol. 
42
 
5
(pg. 
406
-
409
)
79
Suwaki
 
N
Klare
 
K
Tarsounas
 
M
RAD51 paralogs: roles in DNA damage signalling, recombinational repair and tumorigenesis.
Semin Cell Dev Biol
2011
, vol. 
22
 
8
(pg. 
898
-
905
)
80
Sigurdsson
 
S
Van Komen
 
S
Bussen
 
W
Schild
 
D
Albala
 
JS
Sung
 
P
Mediator function of the human Rad51B-Rad51C complex in Rad51/RPA-catalyzed DNA strand exchange.
Genes Dev
2001
, vol. 
15
 
24
(pg. 
3308
-
3318
)
81
Peng
 
M
Litman
 
R
Xie
 
J
Sharma
 
S
Brosh
 
RM
Cantor
 
SB
The FANCJ/MutLalpha interaction is required for correction of the cross-link response in FA-J cells.
EMBO J
2007
, vol. 
26
 
13
(pg. 
3238
-
3249
)
82
Williams
 
SA
Wilson
 
JB
Clark
 
AP
, et al. 
Functional and physical interaction between the mismatch repair and FA-BRCA pathways.
Hum Mol Genet
2011
, vol. 
20
 
22
(pg. 
4395
-
4410
)
83
Kim
 
Y
Lach
 
FP
Desetty
 
R
Hanenberg
 
H
Auerbach
 
AD
Smogorzewska
 
A
Mutations of the SLX4 gene in Fanconi anemia.
Nat Genet
2011
, vol. 
43
 
2
(pg. 
142
-
146
)
84
Stoepker
 
C
Hain
 
K
Schuster
 
B
, et al. 
SLX4, a coordinator of structure-specific endonucleases, is mutated in a new Fanconi anemia subtype.
Nat Genet
2011
, vol. 
43
 
2
(pg. 
138
-
141
)
85
Bogliolo
 
M
Schuster
 
B
Stoepker
 
C
, et al. 
Mutations in ERCC4, encoding the DNA-repair endonuclease XPF, cause Fanconi anemia.
Am J Hum Genet
2013
, vol. 
92
 
5
(pg. 
800
-
806
)
86
Kashiyama
 
K
Nakazawa
 
Y
Pilz
 
DT
, et al. 
Malfunction of nuclease ERCC1-XPF results in diverse clinical manifestations and causes Cockayne syndrome, xeroderma pigmentosum, and Fanconi anemia.
Am J Hum Genet
2013
, vol. 
92
 
5
(pg. 
807
-
819
)
87
Cybulski
 
KE
Howlett
 
NG
FANCP/SLX4: a Swiss army knife of DNA interstrand crosslink repair.
Cell Cycle
2011
, vol. 
10
 
11
(pg. 
1757
-
1763
)
88
Sengerová
 
B
Wang
 
AT
McHugh
 
PJ
Orchestrating the nucleases involved in DNA interstrand cross-link (ICL) repair.
Cell Cycle
2011
, vol. 
10
 
23
(pg. 
3999
-
4008
)
89
Klein Douwel
 
D
Boonen
 
RA
Long
 
DT
, et al. 
XPF-ERCC1 acts in Unhooking DNA interstrand crosslinks in cooperation with FANCD2 and FANCP/SLX4.
Mol Cell
2014
, vol. 
54
 
3
(pg. 
460
-
471
)
90
Huang
 
M
D’Andrea
 
AD
A new nuclease member of the FAN club.
Nat Struct Mol Biol
2010
, vol. 
17
 
8
(pg. 
926
-
928
)
91
Zhou
 
W
Otto
 
EA
Cluckey
 
A
, et al. 
FAN1 mutations cause karyomegalic interstitial nephritis, linking chronic kidney failure to defective DNA damage repair.
Nat Genet
2012
, vol. 
44
 
8
(pg. 
910
-
915
)
92
Brill
 
SJ
Linking the Enzymes that Unlink DNA.
Mol Cell
2013
, vol. 
52
 
2
(pg. 
159
-
160
)
93
Castor
 
D
Nair
 
N
Déclais
 
AC
, et al. 
Cooperative control of holliday junction resolution and DNA repair by the SLX1 and MUS81-EME1 nucleases.
Mol Cell
2013
, vol. 
52
 
2
(pg. 
221
-
233
)
94
Long
 
DT
Räschle
 
M
Joukov
 
V
Walter
 
JC
Mechanism of RAD51-dependent DNA interstrand cross-link repair.
Science
2011
, vol. 
333
 
6038
(pg. 
84
-
87
)
95
Sharma
 
S
Helchowski
 
CM
Canman
 
CE
The roles of DNA polymerase ζ and the Y family DNA polymerases in promoting or preventing genome instability.
Mutat Res
2013
, vol. 
743-744
 (pg. 
97
-
110
)
96
Zhang
 
J
Walter
 
JC
Mechanism and regulation of incisions during DNA interstrand cross-link repair.
DNA Repair (Amst)
2014
, vol. 
19
 (pg. 
135
-
142
)
97
Nalepa
 
G
Clapp
 
DW
Fanconi anemia and the cell cycle: new perspectives on aneuploidy.
F1000Prime Rep
2014
, vol. 
6
 pg. 
23
 
98
Vinciguerra
 
P
Godinho
 
SA
Parmar
 
K
Pellman
 
D
D'Andrea
 
AD
Cytokinesis failure occurs in Fanconi anemia pathway-deficient murine and human bone marrow hematopoietic cells.
J Clin Invest
2010
, vol. 
120
 
11
(pg. 
3834
-
3842
)
99
Naim
 
V
Rosselli
 
F
The FANC pathway and BLM collaborate during mitosis to prevent micro-nucleation and chromosome abnormalities.
Nat Cell Biol
2009
, vol. 
11
 
6
(pg. 
761
-
768
)
100
Chan
 
KL
Palmai-Pallag
 
T
Ying
 
S
Hickson
 
ID
Replication stress induces sister-chromatid bridging at fragile site loci in mitosis.
Nat Cell Biol
2009
, vol. 
11
 
6
(pg. 
753
-
760
)
101
Nalepa
 
G
Enzor
 
R
Sun
 
Z
, et al. 
Fanconi anemia signaling network regulates the spindle assembly checkpoint.
J Clin Invest
2013
, vol. 
123
 
9
(pg. 
3839
-
3847
)
102
Wang
 
X
Kennedy
 
RD
Ray
 
K
Stuckert
 
P
Ellenberger
 
T
D'Andrea
 
AD
Chk1-mediated phosphorylation of FANCE is required for the Fanconi anemia/BRCA pathway.
Mol Cell Biol
2007
, vol. 
27
 
8
(pg. 
3098
-
3108
)
103
Taniguchi
 
T
Garcia-Higuera
 
I
Xu
 
B
, et al. 
Convergence of the Fanconi anemia and ataxia telangiectasia signaling pathways.
Cell
2002
, vol. 
109
 
4
(pg. 
459
-
472
)
104
Andreassen
 
PR
D'Andrea
 
AD
Taniguchi
 
T
ATR couples FANCD2 monoubiquitination to the DNA-damage response.
Genes Dev
2004
, vol. 
18
 
16
(pg. 
1958
-
1963
)
105
Ho
 
GPH
Margossian
 
S
Taniguchi
 
T
D'Andrea
 
AD
Phosphorylation of FANCD2 on two novel sites is required for mitomycin C resistance.
Mol Cell Biol
2006
, vol. 
26
 
18
(pg. 
7005
-
7015
)
106
Wilson
 
JB
Yamamoto
 
K
Marriott
 
AS
, et al. 
FANCG promotes formation of a newly identified protein complex containing BRCA2, FANCD2 and XRCC3.
Oncogene
2008
, vol. 
27
 
26
(pg. 
3641
-
3652
)
107
Wilson
 
JB
Blom
 
E
Cunningham
 
R
Xiao
 
Y
Kupfer
 
GM
Jones
 
NJ
Several tetratricopeptide repeat (TPR) motifs of FANCG are required for assembly of the BRCA2/D1-D2-G-X3 complex, FANCD2 monoubiquitylation and phleomycin resistance.
Mutat Res
2010
, vol. 
689
 
1-2
(pg. 
12
-
20
)
108
Zhi
 
G
Wilson
 
JB
Chen
 
X
, et al. 
Fanconi anemia complementation group FANCD2 protein serine 331 phosphorylation is important for fanconi anemia pathway function and BRCA2 interaction.
Cancer Res
2009
, vol. 
69
 
22
(pg. 
8775
-
8783
)
109
Ceccaldi
 
R
Briot
 
D
Larghero
 
J
, et al. 
Spontaneous abrogation of the G2DNA damage checkpoint has clinical benefits but promotes leukemogenesis in Fanconi anemia patients.
J Clin Invest
2011
, vol. 
121
 
1
(pg. 
184
-
194
)
110
Rosado
 
IV
Langevin
 
F
Crossan
 
GP
Takata
 
M
Patel
 
KJ
Formaldehyde catabolism is essential in cells deficient for the Fanconi anemia DNA-repair pathway.
Nat Struct Mol Biol
2011
, vol. 
18
 
12
(pg. 
1432
-
1434
)
111
Garaycoechea
 
JI
Crossan
 
GP
Langevin
 
F
Daly
 
M
Arends
 
MJ
Patel
 
KJ
Genotoxic consequences of endogenous aldehydes on mouse haematopoietic stem cell function.
Nature
2012
, vol. 
489
 
7417
pg. 
571
 
112
Hira
 
A
Yabe
 
H
Yoshida
 
K
, et al. 
Variant ALDH2 is associated with accelerated progression of bone marrow failure in Japanese Fanconi anemia patients.
Blood
2013
, vol. 
122
 
18
(pg. 
3206
-
3209
)
113
Li
 
XX
Le Beau
 
MM
Ciccone
 
S
, et al. 
Ex vivo culture of Fancc(−/−) stem/progenitor cells predisposes cells to undergo apoptosis, and surviving stem/progenitor cells display cytogenetic abnormalities and an increased risk of malignancy.
Blood
2005
, vol. 
105
 
9
(pg. 
3465
-
3471
)
114
Matsushita
 
N
Endo
 
Y
Sato
 
K
, et al. 
Direct inhibition of TNF-alpha promoter activity by Fanconi anemia protein FANCD2.
PLoS ONE
 
doi: 10.1371/journal.pone.0023324
115
Zhang
 
X
Sejas
 
DP
Qiu
 
Y
Williams
 
DA
Pang
 
Q
Inflammatory ROS promote and cooperate with the Fanconi anemia mutation for hematopoietic senescence.
J Cell Sci
2007
, vol. 
120
 
9
(pg. 
1572
-
1583
)
116
Fagerlie
 
S
Lensch
 
MW
Pang
 
QS
Bagby
 
GC
The Fanconi anemia group C gene product: Signaling functions in hematopoietic cells.
Exp Hematol
2001
, vol. 
29
 
12
(pg. 
1371
-
1381
)
117
Rathbun
 
RK
Christianson
 
TA
Faulkner
 
GR
, et al. 
Interferon-gamma-induced apoptotic responses of Fanconi anemia group C hematopoietic progenitor cells involve caspase 8-dependent activation of caspase 3 family members.
Blood
2000
, vol. 
96
 
13
(pg. 
4204
-
4211
)
118
Hu
 
L
Huang
 
W
Hjort
 
E
Eklund
 
EA
Increased Fanconi C expression contributes to the emergency granulopoiesis response.
J Clin Invest
2013
, vol. 
123
 
9
(pg. 
3952
-
3966
)
119
Pang
 
QS
Christianson
 
TA
Keeble
 
W
Koretsky
 
T
Bagby
 
GC
The anti-apoptotic function of Hsp70 in the interferon-inducible double-stranded RNA-dependent protein kinase-mediated death signaling pathway requires the Fanconi anemia protein, FANCC.
J Biol Chem
2002
, vol. 
277
 
51
(pg. 
49638
-
49643
)
120
Park
 
SJ
Ciccone
 
SLM
Beck
 
BD
, et al. 
Oxidative stress/damage induces multimerization and interaction of Fanconi anemia proteins.
J Biol Chem
2004
, vol. 
279
 
29
(pg. 
30053
-
30059
)
121
Scanlon
 
SE
Glazer
 
PM
Hypoxic stress facilitates acute activation and chronic downregulation of Fanconi anemia proteins.
Mol Cancer Res
2014
122
Kumari
 
U
Ya Jun
 
W
Huat Bay
 
B
Lyakhovich
 
A
Evidence of mitochondrial dysfunction and impaired ROS detoxifying machinery in Fanconi anemia cells.
Oncogene
2014
, vol. 
33
 
2
(pg. 
165
-
172
)
123
Hadjur
 
S
Ung
 
K
Wadsworth
 
L
, et al. 
Defective hematopoiesis and hepatic steatosis in mice with combined deficiencies of the genes encoding Fancc and Cu/Zn superoxide dismutase.
Blood
2001
, vol. 
98
 
4
(pg. 
1003
-
1011
)
124
Muller
 
LU
Milsom
 
MD
Harris
 
CE
, et al. 
Overcoming reprogramming resistance of Fanconi anemia cells.
Blood
2012
, vol. 
119
 
23
(pg. 
5449
-
5457
)
125
Alter
 
B
 
Diagnostic evaluation of FA. In: Owen J, Frohnmayer L, Eiler ME, eds. Fanconi Anemia Standards for Clinical Care. 2nd Ed. London: The Fanconi Anemia Research Fund; 2003:3-17