scholarly journals Evidence for subcomplexes in the Fanconi anemia pathway

Blood ◽  
2006 ◽  
Vol 108 (6) ◽  
pp. 2072-2080 ◽  
Author(s):  
Annette L. Medhurst ◽  
El Houari Laghmani ◽  
Jurgen Steltenpool ◽  
Miriam Ferrer ◽  
Chantal Fontaine ◽  
...  
Keyword(s):  
Fanconi Anemia ◽  
Protein Interactions ◽  
Congenital Abnormalities ◽  
Bone Marrow Failure ◽  
Direct Interaction ◽  
Molecular Architecture ◽  
Cross Link ◽  
Core Complex ◽  
Downstream Target ◽  
Nuclear Complex

AbstractFanconi anemia (FA) is a genomic instability disorder, clinically characterized by congenital abnormalities, progressive bone marrow failure, and predisposition to malignancy. Cells derived from patients with FA display a marked sensitivity to DNA cross-linking agents, such as mitomycin C (MMC). This observation has led to the hypothesis that the proteins defective in FA are involved in the sensing or repair of interstrand cross-link lesions of the DNA. A nuclear complex consisting of a majority of the FA proteins plays a crucial role in this process and is required for the monoubiquitination of a downstream target, FANCD2. Two new FA genes, FANCB and FANCL, have recently been identified, and their discovery has allowed a more detailed study into the molecular architecture of the FA pathway. We demonstrate a direct interaction between FANCB and FANCL and that a complex of these proteins binds FANCA. The interaction between FANCA and FANCL is dependent on FANCB, FANCG, and FANCM, but independent of FANCC, FANCE, and FANCF. These findings provide a framework for the protein interactions that occur “upstream” in the FA pathway and suggest that besides the FA core complex different subcomplexes exist that may have specific functions other than the monoubiquitination of FANCD2.

Differential Phosphorylation of FANCA and FANCG Determines Response of Fanconi Anemia Core Complex to DNA Damage and S Phase.

Blood ◽  
2004 ◽  
Vol 104 (11) ◽  
pp. 726-726
Author(s):  
Jun Mi ◽  
Andrei Tomashevski ◽  
Gary M. Kupfer
Keyword(s):  
Dna Damage ◽  
Fanconi Anemia ◽  
Bone Marrow Failure ◽  
Cellular Responses ◽  
S Phase ◽  
Congenital Defects ◽  
Core Complex ◽  
Downstream Target ◽  
Chk1 Kinase ◽  
Differential Phosphorylation

Abstract Fanconi anemia (FA) is a genetic disease marked by bone marrow failure, congenital defects, and cancer. In spite of the identification of at least 8 genes, the biochemistry of the disease and its normal pathway in the cell remains elusive. The FA core complex is composed of at least 5 proteins, 2 of which, FANCA and FANCG, we have shown to be phosphorylated. In these studies, we show that both FANCA and FANCG are phosphorylated in response to DNA damage. In the case of FANCG, we have mapped the site of this phosphorylation to serine 7, using a phosphoserine 7 FANCG antiserum. Because of the link of FA function and the FA core complex-dependent monoubiquitination that occurs both as a result of DNA damage as well as at S phase, we also examined if phosphorylation occurred at S phase as well. While FANCG serine 7 phosphorylation occurs both at S phase and after DNA damage (similar to FANCD2 monoubiquitination), FANCA phosphorylation occurs only after DNA damage. Recent data have implicated the kinase ATR as important in the pathway. In order to assess whether a downstream target of ATR is differentially phosphorylated in FA cells, we tested the phosphorylation status of chk1 in FA-A mutant and corrected cells. Chk1 kinase is phosphorylated at serine 318 in response to DNA damage only in corrected cells but not mutant FA cells, while signaling through chk2 kinase is unaffected. These data suggest the importance of phosphorylation in the FA pathway in the regulation of both cellular responses to DNA damage as well as engagement of the cell cycle.

Fanconi anemia protein complex: mapping protein interactions in the yeast 2- and 3-hybrid systems

Blood ◽  
2003 ◽  
Vol 102 (1) ◽  
pp. 136-141 ◽  
Author(s):  
Susan M. Gordon ◽  
Manuel Buchwald
Keyword(s):  
Hybrid System ◽  
Fanconi Anemia ◽  
Protein Interactions ◽  
Protein Complex ◽  
Bone Marrow Failure ◽  
Direct Interaction ◽  
Terminal Region ◽  
Amino Terminal ◽  
Contact Points ◽  
Complementation Groups

Abstract Fanconi anemia (FA) is an autosomal recessive syndrome characterized by progressive bone marrow failure and cancer predisposition. Eight FA complementation groups have been identified. The FANCA, FANCC, FANCE, FANCF, and FANCG proteins form a nuclear complex required for the monoubiquination of the FANCD2 protein. To investigate the architecture of the FA protein complex, the yeast 2-hybrid system was used to map contact points of the FANCA/FANCG, FANCC/FANCE, and FANCF/FANCG interactions. FANCG was shown to interact with both the amino-terminus of FANCA and the carboxyl-terminal region of FANCF. A FANCG mutant truncated at the carboxyl-terminus retained the ability to interact with FANCA. The interaction between FANCG and FANCF was ablated by a Leu71Pro mutant of FANCG. A central region of FANCE was sufficient for FANCC binding. A Leu554Pro mutant of FANCC failed to interact with FANCE. To further examine complex assembly, the yeast 3-hybrid system was used to investigate the ability of FANCG to act as a molecular bridge in mediating interaction between other FA proteins. FANCG was able to mediate interaction between FANCA and FANCF, as well as between monomers of FANCA. Direct interaction between FANCE and FANCD2 was also demonstrated in the yeast 2-hybrid system. This interaction involving an amino-terminal region of FANCD2 may provide a link between the FA protein complex and its downstream targets. (Blood. 2003;102:136-141)

scholarly journals Fanconi anemia signaling and Mus81 cooperate to safeguard development and crosslink repair

2014 ◽  
Vol 42 (15) ◽  
pp. 9807-9820 ◽  
Author(s):  
Meghan Larin ◽  
David Gallo ◽  
Laura Tamblyn ◽  
Jay Yang ◽  
Hudson Liao ◽  
...  
Keyword(s):  
Dna Damage ◽  
Fanconi Anemia ◽  
Congenital Abnormalities ◽  
Bone Marrow Failure ◽  
In Utero ◽  
Critical Threshold ◽  
Growth Defects ◽  
Repair Defect ◽  
In Utero Development ◽  
Crosslink Repair

AbstractIndividuals with Fanconi anemia (FA) are susceptible to bone marrow failure, congenital abnormalities, cancer predisposition and exhibit defective DNA crosslink repair. The relationship of this repair defect to disease traits remains unclear, given that crosslink sensitivity is recapitulated in FA mouse models without most of the other disease-related features. Mice deficient in Mus81 are also defective in crosslink repair, yet MUS81 mutations have not been linked to FA. Using mice deficient in both Mus81 and the FA pathway protein FancC, we show both proteins cooperate in parallel pathways, as concomitant loss of FancC and Mus81 triggered cell-type-specific proliferation arrest, apoptosis and DNA damage accumulation in utero. Mice deficient in both FancC and Mus81 that survived to birth exhibited growth defects and an increased incidence of congenital abnormalities. This cooperativity of FancC and Mus81 in developmental outcome was also mirrored in response to crosslink damage and chromosomal integrity. Thus, our findings reveal that both pathways safeguard against DNA damage from exceeding a critical threshold that triggers proliferation arrest and apoptosis, leading to compromised in utero development.

scholarly journals Gingival Bleeding in a Child with Fanconi Anemia: A Case Report and Literature Review

2020 ◽  
Vol 2020 ◽  
pp. 1-5
Author(s):  
Dorsaf Touil ◽  
Rahma Bouhouch ◽  
Raoua Belkacem Chebil ◽  
Lamia Oualha ◽  
Nabiha Douki
Keyword(s):  
Fanconi Anemia ◽  
Autosomal Recessive ◽  
Congenital Abnormalities ◽  
Bone Marrow Failure ◽  
Autosomal Recessive Disorder ◽  
Oral Manifestations ◽  
Gingival Bleeding ◽  
Oral Conditions ◽  
Multiple Congenital Abnormalities

Fanconi anemia (FA) is a rare autosomal recessive disorder characterized by multiple congenital abnormalities, bone marrow failure, and higher susceptibility to malignancies, especially to head and neck carcinomas. Only few reports about the oral manifestations of FA are available. The main reported oral conditions associated with FA are microdontia and advanced periodontitis. The aim of this paper was to report a case of a 10-year-old patient with FA presenting severe spontaneous gingival bleeding, as well as to discuss the role of the dentist in the management and treatment of this condition.

Coordinated Chromatin-Association of Fanconi Anemia Network Proteins Requires Replication-Coupled DNA Damage Recognition.

Blood ◽  
2004 ◽  
Vol 104 (11) ◽  
pp. 723-723
Author(s):  
Alexandra Sobeck ◽  
Stacie Stone ◽  
Bendert deGraaf ◽  
Vincenzo Costanzo ◽  
Johan deWinter ◽  
...  
Keyword(s):  
Dna Damage ◽  
Dna Replication ◽  
Dna Damage Response ◽  
Fanconi Anemia ◽  
Bone Marrow Failure ◽  
S Phase ◽  
Dependent Manner ◽  
Core Complex ◽  
Damage Response ◽  
Crosslinking Agents

Abstract Fanconi anemia (FA) is a genetic disorder characterized by hypersensitivity to DNA crosslinking agents and diverse clinical symptoms, including developmental anomalies, progressive bone marrow failure, and predisposition to leukemias and other cancers. FA is genetically heterogeneous, resulting from mutations in any of at least eleven different genes. The FA proteins function together in a pathway composed of a mulitprotein core complex that is required to trigger the DNA-damage dependent activation of the downstream FA protein, FANCD2. This activation is thought to be the key step in a DNA damage response that functionally links FA proteins to major breast cancer susceptibility proteins BRCA1 and BRCA2 (BRCA2 is FA gene FANCD1). The essential function of the FA proteins is unknown, but current models suggest that FA proteins function at the interface between cell cycle checkpoints, DNA repair and DNA replication, and are likely to play roles in the DNA damage response during S phase. To provide a platform for dissecting the key functional events during S-phase, we developed cell-free assays for FA proteins based on replicating extracts from Xenopus eggs. We identified the Xenopus homologs of human FANCD2 (xFANCD2) and several of the FA core complex proteins (xCCPs), and biochemically characterized these proteins in replicating cell-free extracts. We found that xCCPs and a modified isoform of xFANCD2 become associated with chromatin during normal and disrupted DNA replication. Blocking initiation of replication with geminin demonstrated that association of xCCPs and xFANCD2 with chromatin occurs in a strictly replication-dependent manner that is enhanced following DNA damage by crosslinking agents or by addition of aphidicolin, an inhibitor of replicative DNA polymerases. In addition, chromatin binding of xFANCD2, but not xBRCA2, is abrogated when xFANCA is quantitatively depleted from replicating extracts suggesting that xFANCA promotes the loading of xFANCD2 on chromatin. The chromatin-association of xFANCD2 and xCCPs is diminished in the presence of caffeine, an inhibitor of checkpoint kinases. Taken together, our data suggest a model in which the ordered loading of FA proteins on chromatin is required for processing a subset of DNA replication-blocking lesions that are resolved during late stages of replication.

scholarly journals Clinical severity in Fanconi anemia correlates with residual function of FANCB missense variants

2019 ◽  
Author(s):  
Moonjung Jung ◽  
Ramanagouda Ramanagoudr-Bhojappa ◽  
Sylvie van Twest ◽  
Rasim Ozgur Rosti ◽  
Vincent Murphy ◽  
...  
Keyword(s):  
Bone Marrow ◽  
Fanconi Anemia ◽  
Congenital Abnormalities ◽  
Bone Marrow Failure ◽  
Complementation Group ◽  
Clinical Severity ◽  
List Type ◽  
Missense Variants ◽  
Pathogenic Variants ◽  
Fancd2 Monoubiquitination

ABSTRACTFanconi anemia (FA) is the most common genetic cause of bone marrow failure, and is caused by inherited pathogenic variants in any of 22 genes. Of these, only FANCB is X-linked. We describe a cohort of 19 children with FANCB variants, from 16 families of the International Fanconi Anemia Registry (IFAR). Those with FANCB deletion or truncation demonstrate earlier than average onset of bone marrow failure, and more severe congenital abnormalities compared to a large series of FA individuals in the published reports. This reflects the indispensable role of FANCB protein in the enzymatic activation of FANCD2 monoubiquitination, an essential step in the repair of DNA interstrand crosslinks. For FANCB missense variants, more variable severity is associated with the extent of residual FANCD2 monoubiquitination activity. We used transcript analysis, genetic complementation, and biochemical reconstitution of FANCD2 monoubiquitination to determine the pathogenicity of each variant. Aberrant splicing and transcript destabilization was associated with two missence variants. Individuals carrying missense variants with drastically reduced FANCD2 monoubiquitination in biochemical and/or cell-based assays showed earlier onset of hematologic disease and shorter survival. Conversely, variants with near-normal FANCD2 monoubiquitination were associated with more favorable outcome. Our study reveals a genotype-phenotype correlation within the FA-B complementation group of FA, where severity is linked to the extent of residual FANCD2 monoubiquitination.KEY POINTSX-linked FANCB pathogenic variants predominantly cause acute, early onset bone marrow failure and severe congenital abnormalitiesBiochemical and cell-based assays with patient variants reveal functional properties of FANCB that associate with clinical severity

Mechanisms of Vertebrate DNA Interstrand Cross-Link Repair

2021 ◽  
Vol 90 (1) ◽  
Author(s):  
Daniel R. Semlow ◽  
Johannes C. Walter
Keyword(s):  
Fanconi Anemia ◽  
Double Helix ◽  
Bone Marrow Failure ◽  
Chemotherapy Resistance ◽  
Chemical Diversity ◽  
Annual Review ◽  
Publication Date ◽  
Cross Link ◽  
Cancer Predisposition Syndrome ◽  
Repair Pathways

DNA interstrand cross-links (ICLs) covalently connect the two strands of the double helix and are extremely cytotoxic. Defective ICL repair causes the bone marrow failure and cancer predisposition syndrome, Fanconi anemia, and upregulation of repair causes chemotherapy resistance in cancer. The central event in ICL repair involves resolving the cross-link (unhooking). In this review, we discuss the chemical diversity of ICLs generated by exogenous and endogenous agents. We then describe how proliferating and nonproliferating vertebrate cells unhook ICLs. We emphasize fundamentally new unhooking strategies, dramatic progress in the structural analysis of the Fanconi anemia pathway, and insights into how cells govern the choice between different ICL repair pathways. Throughout, we highlight the many gaps that remain in our knowledge of these fascinating DNA repair pathways. Expected final online publication date for the Annual Review of Biochemistry, Volume 90 is June 2021. Please see http://www.annualreviews.org/page/journal/pubdates for revised estimates.

Nuclear Localization of Fanconi Anemia Protein FANCA Is Regulated by Hsc70/Hsp90 Chaperone Machinery.

Blood ◽  
2004 ◽  
Vol 104 (11) ◽  
pp. 2835-2835
Author(s):  
Tsukasa Oda ◽  
Hidenobu Miyaso ◽  
Takayuki Yamashita
Keyword(s):  
Nuclear Localization ◽  
Fanconi Anemia ◽  
Subcellular Distribution ◽  
Molecular Mechanisms ◽  
Bone Marrow Failure ◽  
Genetic Disorder ◽  
Specific Inhibitor ◽  
Current Evidence ◽  
Core Complex ◽  
Hsp90 Chaperone

Abstract Fanconi anemia (FA) is a genetic disorder characterized by bone marrow failure, cancer susceptibility and cellular hypersensitivity to DNA crosslinkers such as mitomycin C (MMC). Current evidence indicates that formation of a nuclear multiprotein complex (core complex) including six FA proteins FANCA/C/E/F/G/L is essential for FANCL/PHF9 ubiquitin ligase-mediated activation of FANCD2 into a monoubiquinated form, which participates in BRCA1 and FANCD1/BRCA2-mediated DNA repair (the FA/BRCA pathway). Subcellular distribution of FANCA plays a crucial role in the regulation of the FA/BRCA pathway. However, the underlying molecular mechanisms are not fully understood. To address this issue, we tried to identify FANCA-associated proteins. To this end, Flag-FANCA ectopically expressed in HeLa cells was immunopurified from the cytoplasmic fraction, using anti-Flag antibody-conjugated sepharose beads. Analysis of the immune complex on SDS polyacrylamide gel electrophoresis revealed that several proteins of Mr. 60–70 kD specifically associated with Flag-FANCA. These proteins were identified as FANCG and Hsc (heat shock cognate protein) 70 by LC-MS/MS. Immunoblot analysis showed that FANCA associated with Hsp90 as well as Hsc70. Hsc70 is an ATP-dependent molecular chaperone highly homologous to Hsp70 and often cooperates with Hsp90 to form a chaperone machinery involved in the regulation of diverse protein functions. Patient-derived FANCA mutants failed to bind FANCC but associated with larger amounts of Hsc70 than wt-FANCA, indicating that the interaction between FANCA and Hsc70 is not mediated by FANCC, as suggested by previous observations of the interaction of FANCC with Hsp70. To study the role of Hsc70 and Hsp90 in the regulation of FANCA, we examined effects of a dominant-negative (dn) form of Hsc70 with inactivated ATPase activity, and a specific inhibitor of Hsp90, 17-AAG (a geldanamycin analog). Overexpression of dn-Hsc70 inhibited nuclear localization of FANCA and inhibited its core complex formation, whereas wt-Hsc70 did not. 17-AAG induced cytoplasmic distribution and proteosomal degradation of FANCA and suppressed FANCD2 mono-ubiquitination. Taken together, these results suggest that Hsc70/Hsp90 chaperone machinery interacts with FANCA and regulates its subcellular distribution and stability, thereby controlling activation of the FA/BRCA pathway.

scholarly journals HES1 is a novel interactor of the Fanconi anemia core complex

Blood ◽  
2008 ◽  
Vol 112 (5) ◽  
pp. 2062-2070 ◽  
Author(s):  
Cédric S. Tremblay ◽  
Feng F. Huang ◽  
Ouassila Habi ◽  
Caroline C. Huard ◽  
Chantal Godin ◽  
...  
Keyword(s):  
Fanconi Anemia ◽  
Bone Marrow Failure ◽  
Developmental Pathway ◽  
Core Complex ◽  
Interacting Protein ◽  
Developmental Abnormalities ◽  
Protein Protein Interaction ◽  
The Core ◽  
Cross Links ◽  
Enhancer Of Split

Abstract Fanconi anemia (FA) proteins are thought to play a role in chromosome stability and repair of DNA cross-links; however, these functions may not fully explain the developmental abnormalities and bone marrow failure that are characteristic of FA individuals. Here we associate the FA proteins with the Notch1 developmental pathway through a direct protein-protein interaction between the FA core complex and the hairy enhancer of split 1 (HES1). HES1 interaction with FA core complex members is dependent on a functional FA pathway. Cells depleted of HES1 exhibit an FA-like phenotype that includes cellular hypersensitivity to mitomycin C (MMC) and lack of FANCD2 monoubiquitination and foci formation. HES1 is also required for proper nuclear localization or stability of some members of the core complex. Our results suggest that HES1 is a novel interacting protein of the FA core complex.

scholarly journals Chk1-Mediated Phosphorylation of FANCE Is Required for the Fanconi Anemia/BRCA Pathway

2007 ◽  
Vol 27 (8) ◽  
pp. 3098-3108 ◽  
Author(s):  
XiaoZhe Wang ◽  
Richard D. Kennedy ◽  
Kallol Ray ◽  
Patricia Stuckert ◽  
Tom Ellenberger ◽  
...  
Keyword(s):  
Fanconi Anemia ◽  
Mutant Form ◽  
Cross Link ◽  
Core Complex ◽  
Core Enzyme ◽  
Cross Links ◽  
Phase Progression ◽  
Nuclear Foci ◽  
Fancd2 Monoubiquitination

ABSTRACT The eleven Fanconi anemia (FA) proteins cooperate in a novel pathway required for the repair of DNA cross-links. Eight of the FA proteins (A, B, C, E, F, G, L, and M) form a core enzyme complex, required for the monoubiquitination of FANCD2 and the assembly of FANCD2 nuclear foci. Here, we show that, in response to DNA damage, Chk1 directly phosphorylates the FANCE subunit of the FA core complex on two conserved sites (threonine 346 and serine 374). Phosphorylated FANCE assembles in nuclear foci and colocalizes with FANCD2. A nonphosphorylated mutant form of FANCE (FANCE-T346A/S374A), when expressed in a FANCE-deficient cell line, allows FANCD2 monoubiquitination, FANCD2 foci assembly, and normal S-phase progression. However, the mutant FANCE protein fails to complement the mitomycin C hypersensitivity of the transfected cells. Taken together, these results elucidate a novel role of Chk1 in the regulation of the FA/BRCA pathway and in DNA cross-link repair. Chk1-mediated phosphorylation of FANCE is required for a function independent of FANCD2 monoubiquitination.

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