RESUMEN
BACKGROUND: Monoallelic germline pathogenic variants (GPVs) in five Fanconi anemia (FA) genes (BRCA1/FANCS, BRCA2/FANCD1, PALB2/FANCN, BRIP1/FANCJ, and RAD51C/FANCO) confer an increased risk of breast (BC) and/or ovarian (OC) cancer, but the role of GPVs in 17 other FA genes remains unclear. METHODS: Here, we investigated the association of germline variants in FANCG/XRCC9 with BC and OC risk. RESULTS: The frequency of truncating GPVs in FANCG did not differ between BC (20/10,204; 0.20%) and OC (8/2966; 0.27%) patients compared to controls (6/3250; 0.18%). In addition, only one out of five tumor samples showed loss-of-heterozygosity of the wild-type FANCG allele. Finally, none of the nine functionally tested rare recurrent missense FANCG variants impaired DNA repair activities (FANCD2 monoubiquitination and FANCD2 foci formation) upon DNA damage, in contrast to all tested FANCG truncations. CONCLUSION: Our study suggests that heterozygous germline FANCG variants are unlikely to contribute to the development of BC or OC.
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Neoplasias de la Mama , Proteína del Grupo de Complementación G de la Anemia de Fanconi , Predisposición Genética a la Enfermedad , Mutación de Línea Germinal , Neoplasias Ováricas , Humanos , Femenino , Neoplasias de la Mama/genética , Neoplasias Ováricas/genética , Proteína del Grupo de Complementación G de la Anemia de Fanconi/genética , Persona de Mediana Edad , Adulto , Reparación del ADN/genética , Estudios de Casos y Controles , AncianoRESUMEN
Fanconi anemia (FA) develops due to a mutation in one of the FANC genes that are involved in the repair of interstrand crosslinks (ICLs). FANCG, a member of the FA core complex, is essential for ICL repair. Previous FANCG-deficient mouse models were generated with drug-based selection cassettes in mixed mice backgrounds, leading to a disparity in the interpretation of genotype-related phenotype. We created a Fancg-KO (KO) mouse model using CRISPR/Cas9 to exclude these confounders. The entire Fancg locus was targeted and maintained on the immunological well-characterized C57BL/6J background. The intercrossing of heterozygous mice resulted in sub-Mendelian numbers of homozygous mice, suggesting the loss of FANCG can be embryonically lethal. KO mice displayed infertility and hypogonadism, but no other developmental problems. Bone marrow analysis revealed a defect in various hematopoietic stem and progenitor subsets with a bias towards myelopoiesis. Cell lines derived from Fancg-KO mice were hypersensitive to the crosslinking agents cisplatin and Mitomycin C, and Fancg-KO mouse embryonic fibroblasts (MEFs) displayed increased γ-H2AX upon cisplatin treatment. The reconstitution of these MEFs with Fancg cDNA corrected for the ICL hypersensitivity. This project provides a new, genetically, and immunologically well-defined Fancg-KO mouse model for further in vivo and in vitro studies on FANCG and ICL repair.
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Cisplatino , Anemia de Fanconi , Humanos , Animales , Ratones , Cisplatino/metabolismo , Anemia de Fanconi/genética , Anemia de Fanconi/metabolismo , Ratones Endogámicos C57BL , Sistemas CRISPR-Cas , Proteínas de Unión al ADN/metabolismo , Fibroblastos/metabolismo , Mitomicina , Fenotipo , Proteína del Grupo de Complementación G de la Anemia de Fanconi/genéticaRESUMEN
Bernese mountain dogs (BMDs), have an overall cancer incidence of 50%, half of which is comprised of an otherwise rare tumor, histiocytic sarcoma (HS). While recent studies have identified driver mutations in the MAPK pathway, identification of key predisposing genes has been elusive. Studies have identified several loci to be associated with predisposition to HS in BMDs, including near the MTAP/CDKN2A region, but no causative coding variant has been identified. Here we report the presence of a coding polymorphism in the gene encoding FANCG, near the MTAP/CDKN2A locus. This variant is in a conserved region of the protein and appears to be specific to BMDs. Canine fibroblasts derived from dogs homozygous for this variant are hypersensitive to cisplatin. We show this canine FANCG variant and a previously defined hypomorphic FANCG allele in humans impart similar defects in DNA repair. However, our data also indicate that this variant is neither necessary nor sufficient for the development of HS. Furthermore, BMDs homozygous for this FANCG allele display none of the characteristic phenotypes associated with Fanconi anemia (FA) such as anemia, short stature, infertility, or an earlier age of onset for HS. This is similar to findings in FA deficient mice, which do not develop overt FA without secondary genetic mutations that exacerbate the FA deficit. In sum, our data suggest that dogs with deficits in the FA pathway are, like mice, innately resistant to the development of FA.
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Anemia de Fanconi , Sarcoma Histiocítico , Humanos , Perros , Animales , Ratones , Anemia de Fanconi/genética , Cisplatino , Sarcoma Histiocítico/genética , Mutación , Alelos , Proteína del Grupo de Complementación G de la Anemia de Fanconi/genéticaRESUMEN
Fanconi anemia (FA) is a rare genetic disorder caused by pathogenic variants (PV) in at least 22 genes, which cooperate in the Fanconi anemia/Breast Cancer (FA/BRCA) pathway to maintain genome stability. PV in FANCA, FANCC, and FANCG account for most cases (~90%). This study evaluated the chromosomal, molecular, and physical phenotypic findings of a novel founder FANCG PV, identified in three patients with FA from the Mixe community of Oaxaca, Mexico. All patients presented chromosomal instability and a homozygous PV, FANCG: c.511-3_511-2delCA, identified by next-generation sequencing analysis. Bioinformatic predictions suggest that this deletion disrupts a splice acceptor site promoting the exon 5 skipping. Analysis of Cytoscan 750 K arrays for haplotyping and global ancestry supported the Mexican origin and founder effect of the variant, reaffirming the high frequency of founder PV in FANCG. The degree of bone marrow failure and physical findings (described through the acronyms VACTERL-H and PHENOS) were used to depict the phenotype of the patients. Despite having a similar frequency of chromosomal aberrations and genetic constitution, the phenotype showed a wide spectrum of severity. The identification of a founder PV could help for a systematic and accurate genetic screening of patients with FA suspicion in this population.
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Anemia de Fanconi , Biología Computacional , Anemia de Fanconi/genética , Anemia de Fanconi/metabolismo , Proteína del Grupo de Complementación G de la Anemia de Fanconi/genética , Efecto Fundador , Homocigoto , Humanos , MéxicoRESUMEN
Among the 22 Fanconi anemia (FA) reported genes, 90% of mutational spectra were found in three genes, namely FANCA (64%), FANCC (12%) and FANCG (8%). Therefore, this study aimed to identify the high-risk deleterious variants in three selected genes (FANCA, FANCC, and FANCG) through various computational approaches. The missense variant datasets retrieved from the UCSC genome browser were analyzed for their pathogenicity, stability, and phylogenetic conservancy. A total of 23 alterations, of which 16 in FANCA, 6 in FANCC and one variant in FANCG, were found to be highly deleterious. The native and mutant structures were generated, which demonstrated a profound impact on the respective proteins. Besides, their pathway analysis predicted many other pathways in addition to the Fanconi anemia pathway, homologous recombination, and mismatch repair pathways. Hence, this is the first comprehensive study that can be useful for understanding the genetic signatures in the development of FA.
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Biología Computacional/métodos , Proteína del Grupo de Complementación A de la Anemia de Fanconi/genética , Proteína del Grupo de Complementación C de la Anemia de Fanconi/genética , Proteína del Grupo de Complementación G de la Anemia de Fanconi/genética , Anemia de Fanconi/genética , Mutación Missense , Sitios de Unión , Proteína del Grupo de Complementación A de la Anemia de Fanconi/química , Proteína del Grupo de Complementación C de la Anemia de Fanconi/química , Proteína del Grupo de Complementación G de la Anemia de Fanconi/química , Estudios de Asociación Genética , Predisposición Genética a la Enfermedad , Humanos , Modelos Moleculares , Conformación Proteica , Estabilidad ProteicaRESUMEN
INTRODUCTION: Fanconi anemia (FA) is a rare inherited hematological disease due to a defect in the DNA repair pathway resulting in congenital abnormalities and high susceptibility to develop cancers. The cytogenetic analysis using alkylating agents is still a reference test to establish the diagnosis. Despite the genetic heterogeneity, the identification of the causal mutation is actually performed especially after the development of next generation sequencing (NGS). METHODS: we report here nine Moroccan patients referred to the department of Medical Genetics for suspicion of FA. We realized a genetic consultation to establish a clinical record with biological data before carrying out the genetic analysis. Karyotyping with mitomycin was performed for all the probands before elaborating molecular study. We used massively parallel sequencing to analyse the three most frequent mutated genes FANCA, FANCC, and FANCG, representing 84% of all genes involved in FA. RESULTS: all the patients showed hematological signs associated with at least one extra-hematological congenital anomaly. The chromosomal breaks were significantly higher for the nine patients, compared to the controls. The molecular diagnosis was confirmed in 8 of the 9 families tested (88.8%) with 4 novel mutations. The next generation based sequencing identified 9 variations: 6 in the FANCA gene (66.6%), 3 in the FANCG gene (33.3%) and no FANCC variation was found. Of those, 7 were homozygous and 2 were compounds heterozygous. CONCLUSION: to the best of our knowledge, this is the first molecular report of Moroccan patients with FA suggesting the predominance of two genes without any recurrent mutation. The molecular analysis of FANCA and FANCG genes should be offered first for all patients in Morocco.
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Proteína del Grupo de Complementación A de la Anemia de Fanconi/genética , Proteína del Grupo de Complementación G de la Anemia de Fanconi/genética , Anemia de Fanconi/diagnóstico , Niño , Preescolar , Análisis Citogenético , Anemia de Fanconi/genética , Proteína del Grupo de Complementación C de la Anemia de Fanconi/genética , Femenino , Pruebas Genéticas , Secuenciación de Nucleótidos de Alto Rendimiento , Humanos , Masculino , Marruecos , MutaciónAsunto(s)
Médula Ósea , Rotura Cromosómica , Células Precursoras Eritroides , Proteína del Grupo de Complementación G de la Anemia de Fanconi/genética , Anemia de Fanconi , Mutación , Médula Ósea/metabolismo , Médula Ósea/patología , Preescolar , Células Precursoras Eritroides/metabolismo , Células Precursoras Eritroides/patología , Anemia de Fanconi/genética , Anemia de Fanconi/metabolismo , Anemia de Fanconi/patología , Proteína del Grupo de Complementación G de la Anemia de Fanconi/metabolismo , Humanos , MasculinoRESUMEN
Fanconi anemia (FA) is a rare human genetic disorder characterized by bone marrow failure, predisposition to cancer and developmental defects including hypogonadism. Reproductive defects leading to germ cell aplasia are the most consistent phenotypes seen in FA mouse models. We examined the role of the nuclear FA core complex gene Fancg in the development of primordial germ cells (PGCs), the embryonic precursors of adult gametes, during fetal development. PGC maintenance was severely impaired in Fancg-/- embryos. We observed a defect in the number of PGCs starting at E9.5 and a strong attrition at E11.5 and E13.5. Remarkably, we observed a mosaic pattern reflecting a portion of testicular cords devoid of PGCs in E13.5 fetal gonads. Our in vitro and in vivo data highlight a potential role of Fancg in the proliferation and in the intrinsic cell motility abilities of PGCs. The random migratory process is abnormally activated in Fancg-/- PGCs, altering the migration of cells. Increased cell death and PGC attrition observed in E11.5 Fancg-/- embryos are features consistent with delayed migration of PGCs along the migratory pathway to the genital ridges. Moreover, we show that an inhibitor of RAC1 mitigates the abnormal migratory pattern observed in Fancg-/- PGCs.
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Anemia de Fanconi , Animales , Movimiento Celular/genética , Anemia de Fanconi/genética , Proteína del Grupo de Complementación G de la Anemia de Fanconi/metabolismo , Células Germinativas/metabolismo , Gónadas/metabolismo , Ratones , Transducción de SeñalRESUMEN
Promyelocytic leukemia (PML) protein is the core component of subnuclear structures called PML nuclear bodies that are known to play important roles in cell survival, DNA damage responses, and DNA repair. Fanconi anemia (FA) proteins are required for repairing interstrand DNA crosslinks (ICLs). Here we report a novel role of PML proteins, regulating the ICL repair pathway. We found that depletion of the PML protein led to the significant reduction of damage-induced FANCD2 mono-ubiquitination and FANCD2 foci formation. Consistently, the cells treated with siRNA against PML showed enhanced sensitivity to a crosslinking agent, mitomycin C. Further studies showed that depletion of PML reduced the protein expression of FANCA, FANCG, and FANCD2 via reduced transcriptional activity. Interestingly, we observed that damage-induced CHK1 phosphorylation was severely impaired in cells with depleted PML, and we demonstrated that CHK1 regulates FANCA, FANCG, and FANCD2 transcription. Finally, we showed that inhibition of CHK1 phosphorylation further sensitized cancer cells to mitomycin C. Taken together, these findings suggest that the PML is critical for damage-induced CHK1 phosphorylation, which is important for FA gene expression and for repairing ICLs.
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Quinasa 1 Reguladora del Ciclo Celular (Checkpoint 1)/metabolismo , Proteína del Grupo de Complementación A de la Anemia de Fanconi/metabolismo , Proteína del Grupo de Complementación D2 de la Anemia de Fanconi/metabolismo , Proteína del Grupo de Complementación G de la Anemia de Fanconi/metabolismo , Anemia de Fanconi/patología , Regulación de la Expresión Génica , Quinasa 1 Reguladora del Ciclo Celular (Checkpoint 1)/genética , Daño del ADN , Reparación del ADN , Anemia de Fanconi/genética , Anemia de Fanconi/metabolismo , Proteína del Grupo de Complementación A de la Anemia de Fanconi/genética , Proteína del Grupo de Complementación D2 de la Anemia de Fanconi/genética , Proteína del Grupo de Complementación G de la Anemia de Fanconi/genética , Células HeLa , Humanos , Fosforilación , UbiquitinaciónRESUMEN
BACKGROUND: Fanconi anemia (FA) is an inherited bone marrow failure syndrome associated with characteristic dysmorphology primarily caused by biallelic pathogenic germline variants in any of 22 different DNA repair genes. There are limited data on the specific molecular causes of FA in different ethnic groups. METHODS: We performed exome sequencing and copy number variant analyses on 19 patients with FA from 17 families undergoing hematopoietic cell transplantation evaluation in Pakistan. The scientific literature was reviewed, and we curated germline variants reported in patients with FA from South Asia and the Middle East. RESULTS: The genetic causes of FA were identified in 14 of the 17 families: seven FANCA, two FANCC, one FANCF, two FANCG, and two FANCL. Homozygous and compound heterozygous variants were present in 12 and two families, respectively. Nine families carried variants previously reported as pathogenic, including two families with the South Asian FANCL founder variant. We also identified five novel likely deleterious variants in FANCA, FANCF, and FANCG in affected patients. CONCLUSIONS: Our study supports the importance of determining the genomic landscape of FA in diverse populations, in order to improve understanding of FA etiology and assist in the counseling of families.
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Anemia de Fanconi/genética , Frecuencia de los Genes , Adolescente , Asia , Niño , Preescolar , Variaciones en el Número de Copia de ADN , Exoma , Anemia de Fanconi/diagnóstico , Proteína del Grupo de Complementación F de la Anemia de Fanconi/genética , Proteína del Grupo de Complementación G de la Anemia de Fanconi/genética , Proteína del Grupo de Complementación L de la Anemia de Fanconi/genética , Femenino , Efecto Fundador , Humanos , Masculino , Medio Oriente , MutaciónRESUMEN
Fanconi Anemia (FA) is a rare genetic disease with the incidence of 1 in 360,000 and is characterised by bone marrow failure, physical abnormalities, pancytopenia, and high frequency of chromosomal breakage and increased risk of evolving into malignancy. Telomere plays an important role in genomic stability, ageing process and cancers. Telomere shortening has been reported in FA. We studied telomere length in FA subjects and compared with complementation groups. Chromosomal breakage analysis from PHA stimulated, MMC induced peripheral blood culture was carried out in 37 clinically diagnosed FA. Molecular study of FANCA, G, and L was done through Sanger sequencing and next generation sequencing. Telomere length was estimated using real time quantitative polymerase chain reaction (qPCR) method. Student t-test was applied to test the significance. A high frequency of chromosomal breakage was observed in all the patients compared to healthy controls. We found significantly shorter telomere length in all the three complementation groups compare to age matched healthy controls. Among all complementation groups, FANCL showed severe telomere shortening (P value 0.0001). A negative correlation was observed between telomere length and chromosomal breakage frequency (R = -0.3116). Telomere shortening is not uncommon in FA subjects. However the telomere length shortening is different in complementation groups as FANCL showed severe telomere shortening in FA subjects. Though BM transplantation is essential for the management of the FA subjects, the telomere length can be considered as biological marker to understand the prognosis of the disease as FA subjects primarily treated with androgens.
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Proteína del Grupo de Complementación A de la Anemia de Fanconi/genética , Proteína del Grupo de Complementación G de la Anemia de Fanconi/genética , Proteína del Grupo de Complementación L de la Anemia de Fanconi/genética , Anemia de Fanconi/genética , Acortamiento del Telómero/genética , Adolescente , Adulto , Niño , Preescolar , Rotura Cromosómica , Proteínas de Unión al ADN/genética , Anemia de Fanconi/patología , Femenino , Regulación de la Expresión Génica/genética , Secuenciación de Nucleótidos de Alto Rendimiento , Humanos , Masculino , Telómero/genética , Adulto JovenRESUMEN
Fanconi anemia (FA) is the most common inherited bone marrow failure syndrome and presents with cytopenias, characteristic physical features, increased chromosomal breaks, and a higher risk of malignancy. Genetic features of this disease vary among different ethnic groups. We aimed to identify the incidence, outcome, overall condition, and genetic features of patients affected with FA in Lebanon to optimize management, identify the most common genes, describe new mutations, and offer prenatal diagnosis and counseling to the affected families. Over a period of 17 years, 40 patients with FA were identified in 2 major diagnostic laboratories in Lebanon. Information was obtained on their clinical course and outcome from their primary physician. DNA was available in 20 patients and was studied for underlying mutations. FANCA seemed to be the most frequent genetic alteration and 2 novel mutations, one each in FANCA and FANCG, were identified. Nine patients developed various malignancies and died. This is the first study looking at clinical and genetic features of FA in Lebanon, and points to the need for establishing a national and regional registry for this condition.
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Proteína del Grupo de Complementación A de la Anemia de Fanconi/genética , Proteína del Grupo de Complementación G de la Anemia de Fanconi/genética , Anemia de Fanconi/genética , Adolescente , Adulto , Niño , Preescolar , Anemia de Fanconi/epidemiología , Femenino , Humanos , Líbano/epidemiología , Masculino , Mutación , Adulto JovenRESUMEN
We describe an infant female with a syndromic neurodevelopmental clinical phenotype and increased chromosome instability as cellular phenotype. Genotype characterization revealed heterozygous variants in genes directly or indirectly linked to DNA repair: a de novo X-linked HDAC8 pathogenic variant, a paternally inherited FANCG pathogenic variant and a maternally inherited BRCA2 variant of uncertain significance. The full spectrum of the phenotype cannot be explained by any of the heterozygous variants on their own; thus, a synergic contribution is proposed. Complementation studies showed that the FANCG gene from the Fanconi Anaemia/BRCA (FA/BRCA) DNA repair pathway was impaired, indicating that the variant in FANCG contributes to the cellular phenotype. The patient's chromosome instability represents the first report where heterozygous variant(s) in the FA/BRCA pathway are implicated in the cellular phenotype. We propose that a multigenic contribution of heterozygous variants in HDAC8 and the FA/BRCA pathway might have a role in the phenotype of this neurodevelopmental disorder. The importance of these findings may have repercussion in the clinical management of other cases with a similar synergic contribution of heterozygous variants, allowing the establishment of new genotype-phenotype correlations and motivating the biochemical study of the underlying mechanisms.
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Proteína BRCA1/genética , Proteína BRCA2/genética , Inestabilidad Cromosómica , Proteína del Grupo de Complementación G de la Anemia de Fanconi/genética , Histona Desacetilasas/genética , Trastornos del Neurodesarrollo/patología , Fenotipo , Proteínas Represoras/genética , Daño del ADN , Reparación del ADN , Femenino , Humanos , Recién Nacido , Mutación , Trastornos del Neurodesarrollo/genéticaRESUMEN
Fanconi anemia (FA) is a rare chromosomal instability syndrome with various clinical features and high cancer incidence. Despite being a DNA repair disorder syndrome and a frequently observed clinical hypersensitivity of FA patients towards ionizing radiation, the experimental evidence regarding the efficiency of radiation-induced DNA double-strand break (DSB) repair in FA is very controversial. Here, we performed a thorough analysis of the repair of radiation-induced DSBs in G1 and G2 in FA fibroblasts of complementation groups A, C, D1 (BRCA2), D2, E, F, G and P (SLX4) in comparison to normal human lung and skin fibroblasts. γH2AX, 53BP1, or RPA foci quantification after X-irradiation was combined with cell cycle markers. Cytogenetic analyses were performed on first metaphases after irradiation in G1 and by premature chromosome condensation after exposure in G2. Furthermore, the role of canonical-NHEJ and alternative-NHEJ for the fidelity of the repair of radiation-induced DSBs was examined. In FA fibroblasts, DSB repair was normal in G1 but compromised and more error-prone in the slow repair component of G2 as suggested by higher yields of radiation-induced γH2AX and 53BP1 foci as well as chromatid exchanges. However, RPA foci quantification in G2 indicated proficiency for homology-directed repair of DSBs in FA except for FA D1 (BRCA2). In lung fibroblasts, DSB repair in G1 was conducted with normal kinetics but elevated chromosome exchanges compared to skin fibroblasts. The overall repair of radiation-induced DSBs and the formation of chromosome exchanges in normal and FA fibroblasts in G1 and G2 were governed by canonical-NHEJ with no contribution of alternative-NHEJ. Together, we show impaired repair of radiation-induced DSBs in various FA complementation groups in the slow repair component of G2 that might promote the formation of potentially oncogenic aberrations and clinical radiation hypersensitivity.
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Ciclo Celular , Aberraciones Cromosómicas , Roturas del ADN de Doble Cadena , Reparación del ADN por Unión de Extremidades , Anemia de Fanconi/metabolismo , Mutación , Reparación del ADN por Recombinación , Proteína BRCA2/genética , Células Cultivadas , ADN/metabolismo , ADN/efectos de la radiación , Anemia de Fanconi/genética , Anemia de Fanconi/fisiopatología , Proteína del Grupo de Complementación A de la Anemia de Fanconi/genética , Proteína del Grupo de Complementación C de la Anemia de Fanconi/genética , Proteína del Grupo de Complementación D2 de la Anemia de Fanconi/genética , Proteína del Grupo de Complementación E de la Anemia de Fanconi/genética , Proteína del Grupo de Complementación F de la Anemia de Fanconi/genética , Proteína del Grupo de Complementación G de la Anemia de Fanconi/genética , Fibroblastos/metabolismo , Fibroblastos/fisiología , Fibroblastos/efectos de la radiación , Histonas/metabolismo , Humanos , Cinética , Recombinasas/genética , Proteína 1 de Unión al Supresor Tumoral P53/metabolismo , Rayos XRESUMEN
Fanconi anemia (FA) is a unique DNA damage repair pathway. To date, 22 genes have been identified that are associated with the FA pathway. A defect in any of those genes causes genomic instability, and the patients bearing the mutation become susceptible to cancer. In our earlier work, we identified that Fanconi anemia protein G (FANCG) protects the mitochondria from oxidative stress. In this report, we have identified eight patients having a mutation (C.65G>C), which converts arginine at position 22 to proline (p.Arg22Pro) in the N terminus of FANCG. The mutant protein, hFANCGR22P, is able to repair the DNA and able to retain the monoubiquitination of FANCD2 in the FANCGR22P/FGR22P cell. However, it lost mitochondrial localization and failed to protect mitochondria from oxidative stress. Mitochondrial instability in the FANCGR22P cell causes the transcriptional downregulation of mitochondrial iron-sulfur cluster biogenesis protein frataxin (FXN) and the resulting iron deficiency of FA protein FANCJ, an iron-sulfur-containing helicase involved in DNA repair.
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Proteína del Grupo de Complementación G de la Anemia de Fanconi/genética , Proteínas del Grupo de Complementación de la Anemia de Fanconi/genética , Inestabilidad Genómica/genética , Proteínas de Unión a Hierro/biosíntesis , Mitocondrias/patología , ARN Helicasas/genética , Secuencia de Aminoácidos/genética , Línea Celular Tumoral , Daño del ADN/genética , Reparación del ADN/genética , Regulación hacia Abajo/genética , Anemia de Fanconi/genética , Anemia de Fanconi/patología , Células HEK293 , Células HeLa , Humanos , Proteínas de Unión a Hierro/genética , Proteínas Hierro-Azufre/genética , Mitocondrias/genética , Mitocondrias/metabolismo , Estrés Oxidativo/fisiología , Especies Reactivas de Oxígeno/metabolismo , Transducción de Señal/genética , FrataxinaRESUMEN
Ovarian cancer is the most lethal gynaecological cancer, and resistance of platinum-based chemotherapy is the main reason for treatment failure. The aim of the present study was to identify candidate genes involved in ovarian cancer platinum response by analysing genes from homologous recombination and Fanconi anaemia pathways. Associations between these two functional genes were explored in the study, and we performed a random walk algorithm based on reconstructed gene-gene network, including protein-protein interaction and co-expression relations. Following the random walk, all genes were ranked and GSEA analysis showed that the biological functions focused primarily on autophagy, histone modification and gluconeogenesis. Based on three types of seed nodes, the top two genes were utilized as examples. We selected a total of six candidate genes (FANCA, FANCG, POLD1, KDM1A, BLM and BRCA1) for subsequent verification. The validation results of the six candidate genes have significance in three independent ovarian cancer data sets with platinum-resistant and platinum-sensitive information. To explore the correlation between biomarkers and clinical prognostic factors, we performed differential analysis and multivariate clinical subgroup analysis for six candidate genes at both mRNA and protein levels. And each of the six candidate genes and their neighbouring genes with a mutation rate greater than 10% were also analysed by network construction and functional enrichment analysis. In the meanwhile, the survival analysis for platinum-treated patients was performed in the current study. Finally, the RT-qPCR assay was used to determine the performance of candidate genes in ovarian cancer platinum response. Taken together, this research demonstrated that comprehensive bioinformatics methods could help to understand the molecular mechanism of platinum response and provide new strategies for overcoming platinum resistance in ovarian cancer treatment.
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Resistencia a Antineoplásicos/genética , Anemia de Fanconi/genética , Recombinación Homóloga/genética , Neoplasias Ováricas/tratamiento farmacológico , Adulto , Anciano , Anciano de 80 o más Años , Algoritmos , Línea Celular Tumoral , ADN Polimerasa III/genética , Supervivencia sin Enfermedad , Anemia de Fanconi/patología , Proteína del Grupo de Complementación A de la Anemia de Fanconi/genética , Proteína del Grupo de Complementación G de la Anemia de Fanconi/genética , Femenino , Regulación Neoplásica de la Expresión Génica/efectos de los fármacos , Histona Demetilasas/genética , Humanos , Persona de Mediana Edad , Neoplasias Ováricas/genética , Neoplasias Ováricas/patología , Platino (Metal)/administración & dosificación , Platino (Metal)/efectos adversos , RecQ Helicasas/genética , Factores de RiesgoRESUMEN
BACKGROUND: Fanconi anemia (FA) is phenotypically diverse, hereditary condition associated with bone marrow failure, multiple physical abnormalities, and an increased susceptibility to the development of malignancies. Less recognized manifestations of FA include endocrine abnormalities. International discourse has highlighted that these abnormalities are widespread among children and adults with FA. To date there has been no systematic study that has evaluated the endocrine abnormalities in a cohort of patients with FA, homozygous for a founder mutation (c.637_643del (p.Tyr213Lysfs*6)) in FANCG. The objectives of the study were to evaluate endocrine gland function in patients with FA of a single FA genotype, and to determine the frequency and nature of endocrine abnormalities in this group. METHODS: Cross-sectional, descriptive study of 24 South African patients of African ancestry with FA (homozygous for a FANCG founder mutation). Outcomes measured included growth, pubertal status, growth hormone axis screening, thyroid gland function, glucose and insulin metabolism and bone age (BA). RESULTS: Endocrine dysfunction was present in 70.8% (17 of 24), including abnormal insulin-like growth factor 1 (IGF-1)/insulin-like growth factor-binding protein 3 (IGFBP-3) in 25.0% (6 of 24), insulin resistance in 41.7% (10 of 24), abnormal thyroid function in 16.7% (4 of 24) and short stature in 45.8% (11 of 24). No abnormalities of glucose metabolism were identified. Abnormal pubertal status was seen in three males (12.5%). Abnormal BAs were present in 34.8% (8 of 23). CONCLUSION: Endocrine abnormalities occur at a high frequency in patients with FA, homozygous for a FANCG founder mutation, similar to other FA cohorts. Our data are specific to FA patients with a single genotype, and therefore provide the first genotype-phenotype information on endocrine abnormalities in South African patients, homozygous for a FANCG founder mutation. Recommendations regarding endocrine screening in this patient subgroup are made, including, but not limited to, baseline testing of thyroid function, fasted insulin and glucose, and IGF-1 and IGFBP-3.
Asunto(s)
Proteína del Grupo de Complementación G de la Anemia de Fanconi/genética , Anemia de Fanconi/genética , Proteína 3 de Unión a Factor de Crecimiento Similar a la Insulina/sangre , Factor I del Crecimiento Similar a la Insulina/metabolismo , Insulina/sangre , Mutación , Hormonas Tiroideas/sangre , Adolescente , Población Negra/genética , Glucemia/metabolismo , Niño , Preescolar , Anemia de Fanconi/sangre , Anemia de Fanconi/patología , Femenino , Efecto Fundador , Homocigoto , Humanos , Masculino , Pubertad/genética , SudáfricaRESUMEN
Monoubiquitination of the Fanconi anemia complementation group D2 (FANCD2) protein by the FA core ubiquitin ligase complex is the central event in the FA pathway. FANCA and FANCG play major roles in the nuclear localization of the FA core complex. Mutations of these two genes are the most frequently observed genetic alterations in FA patients, and most point mutations in FANCA are clustered in the C-terminal domain (CTD). To understand the basis of the FA-associated FANCA mutations, we determined the cryo-electron microscopy (EM) structures of Xenopus laevis FANCA alone at 3.35 Å and 3.46 Å resolution and two distinct FANCA-FANCG complexes at 4.59 and 4.84 Å resolution, respectively. The FANCA CTD adopts an arc-shaped solenoid structure that forms a pseudo-symmetric dimer through its outer surface. FA- and cancer-associated point mutations are widely distributed over the CTD. The two different complex structures capture independent interactions of FANCG with either FANCA C-terminal HEAT repeats, or the N-terminal region. We show that mutations that disturb either of these two interactions prevent the nuclear localization of FANCA, thereby leading to an FA pathway defect. The structure provides insights into the function of FANCA CTD, and provides a framework for understanding FA- and cancer-associated mutations.
Asunto(s)
Proteína del Grupo de Complementación A de la Anemia de Fanconi/ultraestructura , Proteína del Grupo de Complementación D2 de la Anemia de Fanconi/ultraestructura , Proteína del Grupo de Complementación G de la Anemia de Fanconi/ultraestructura , Anemia de Fanconi/genética , Animales , Núcleo Celular/genética , Núcleo Celular/ultraestructura , Microscopía por Crioelectrón , Cristalografía por Rayos X , Proteínas de Unión al ADN/genética , Proteínas de Unión al ADN/ultraestructura , Anemia de Fanconi/patología , Proteína del Grupo de Complementación A de la Anemia de Fanconi/genética , Proteína del Grupo de Complementación D2 de la Anemia de Fanconi/genética , Proteína del Grupo de Complementación G de la Anemia de Fanconi/química , Humanos , Complejos Multiproteicos/química , Complejos Multiproteicos/genética , Mutación , Unión Proteica/genética , Conformación Proteica , Xenopus laevis/genéticaRESUMEN
BACKGROUND/AIM: Radiation mitigator, GS-nitroxide, JP4-039, was evaluated for mitigation of total body irradiation (TBI) in Fanconi anemia (FA) Fancd2-/- (129/Sv), Fancg-/- (B6), and Fanca-/- (129/Sv) mice. MATERIALS AND METHODS: JP4-039 dissolved in 30% 2-hydroxypropyl-ß-cyclodextrin was injected intramuscularly 24 h after total body irradiation (9.25 Gy) into Fanca-/-, Fancd2-/- and Fancg-/- mice. Irradiation survival curves were performed in vitro using bone marrow stromal cell lines derived from Fanca-/-, Fancd2-/- and Fancg-/- mice. RESULTS: FA mice demonstrate genotype specific differences in TBI mitigation by JP4-039. Radiation effects in derived bone marrow stromal cell lines in vitro were mitigated by drugs that block apoptosis, but not necroptosis or ferroptosis. CONCLUSION: FA mouse models are valuable for elucidating DNA repair pathways in cell and tissue responses to TBI, and the role of drugs that target distinct cell death pathways.
Asunto(s)
Anemia de Fanconi/genética , Óxidos de Nitrógeno/farmacología , Protectores contra Radiación/farmacología , Irradiación Corporal Total/efectos adversos , Animales , Apoptosis/efectos de los fármacos , Apoptosis/genética , Apoptosis/efectos de la radiación , Médula Ósea/efectos de los fármacos , Médula Ósea/efectos de la radiación , Línea Celular , Modelos Animales de Enfermedad , Proteína del Grupo de Complementación G de la Anemia de Fanconi/genética , Genotipo , Células Madre Mesenquimatosas/efectos de los fármacos , Células Madre Mesenquimatosas/efectos de la radiación , Ratones , Ratones Endogámicos C57BL , Traumatismos Experimentales por Radiación/tratamiento farmacológico , Traumatismos Experimentales por Radiación/genética , Tolerancia a Radiación/efectos de los fármacos , Tolerancia a Radiación/genéticaRESUMEN
We investigated the potential of next-generation sequencing (NGS) as an alternative method for preimplantation genetic testing of monogenic disease (PGT-M) with human leukocyte antigen (HLA) matching and for noninvasive prenatal diagnosis follow-up. The case involved parents who were carriers of the Fanconi anemia complementation group G (FANCG) 260delG mutation. After clinical PGT using conventional short tandem repeat and mutation analysis, two euploid disease-free embryos were transferred, resulting in a twin pregnancy. Using the original embryo whole genome amplification products from 10 embryos, NGS confirmed the genotypes of the eight nontransferred embryos for both mutation status and HLA combination. NGS also confirmed that the two transferred embryos, which resulted in a twin pregnancy, were euploid, Fanconi disease free, and HLA matched to their sick sibling. At 15 weeks' gestation, noninvasive prenatal diagnosis of the maternal cell-free DNA determined fetal fractions of 14% and 6.6% for twins 1 and 2, respectively. The maternal plasma FANCG 260delG mutation ratio was measured at 46.2%, consistent with the presence of a carrier fetus and a normal fetus. These findings provide proof of concept that NGS has clinical utility as a safe and effective PGT-M method for embryo genotyping as well as more complex direct HLA matching. In addition, NGS can be used to confirm the original PGT-M and HLA matching embryo results in early pregnancy without the need for invasive prenatal diagnosis.