Your browser doesn't support javascript.
Mostrar: 20 | 50 | 100
Resultados 1 - 20 de 8.045
Filtrar
1.
Nat Cell Biol ; 21(9): 1152-1163, 2019 09.
Artigo em Inglês | MEDLINE | ID: mdl-31481791

RESUMO

Ca2+/calmodulin-dependent kinase II (CaMKII) is a multifunctional serine/threonine kinase family, and its δ isoform is predominant in the heart. Excessive CaMKII activation plays a pivotal role in the pathogenesis of severe heart conditions, including myocardial infarction, cardiomyopathy and heart failure. However, the identity of CaMKII splice variants and the mechanism(s) underlying CaMKII-mediated cardiac pathology remain elusive. Here, we show that CaMKII-δ9, the most abundant CaMKII-δ splice variant in human heart, potently promotes cardiomyocyte death, cardiomyopathy and heart failure by disrupting cardiomyocyte genome stability. Mechanistically, CaMKII-δ9, but not the previously well-studied CaMKII-δ2 and CaMKII-δ3, targets the ubiquitin-conjugating enzyme E2T (UBE2T) for phosphorylation and degradation, disrupting UBE2T-dependent DNA repair and leading to the accumulation of DNA damage and genome instability. These findings not only reveal a crucial role of CaMKII in the regulation of DNA repair, but also mark the CaMKII-δ9-UBE2T-DNA damage pathway as an important therapeutic target for cardiomyopathy and heart failure.


Assuntos
Proteína Quinase Tipo 2 Dependente de Cálcio-Calmodulina/metabolismo , Reparo do DNA/genética , Enzimas de Conjugação de Ubiquitina/metabolismo , Animais , Cálcio/metabolismo , Cardiomiopatias/metabolismo , Insuficiência Cardíaca/metabolismo , Humanos , Camundongos , Miócitos Cardíacos/metabolismo , Fosforilação , Isoformas de Proteínas/metabolismo , Enzimas de Conjugação de Ubiquitina/genética
2.
Medicine (Baltimore) ; 98(32): e16541, 2019 Aug.
Artigo em Inglês | MEDLINE | ID: mdl-31393355

RESUMO

BACKGROUNDS: Previous investigations yielded inconsistent results for the associations between pancreatic cancer (PC) risk and genetic polymorphisms. The study aimed to perform a systematic review and meta-analysis of studies exploring association of some genetic polymorphisms and PC risk. METHODS: We systematically searched on PubMed and Web of Science for association of genetic polymorphisms and PC risk published from 1969 to January 2019. We computed the multivariate odd ratio (OR) and 95% confidence intervals (CI), comparing different genetic types. RESULTS: The present meta-analysis showed significant associations between deoxyribonucleic acid (DNA) repair gene (X-ray repair cross-complementing group 1 (XRCC1) Arg399GIn and Arg194Trp, excision repair cross complementation 1 (ERCC1) rs11615 and rs3212986, ERCC2 rs13181) polymorphisms and PC risk. CONCLUSIONS: Because of the limited sample size and ethnicity enrolled in the present meta-analysis, further larger scaled studies should be performed to demonstrate the association.


Assuntos
Neoplasias Pancreáticas/genética , DNA Glicosilases/genética , Reparo do DNA/genética , Proteínas de Ligação a DNA/genética , Endonucleases/genética , Predisposição Genética para Doença , Humanos , Polimorfismo de Nucleotídeo Único , Proteína 1 Complementadora Cruzada de Reparo de Raio-X/genética , Proteína Grupo D do Xeroderma Pigmentoso/genética
3.
Cancer Radiother ; 23(6-7): 666-673, 2019 Oct.
Artigo em Francês | MEDLINE | ID: mdl-31451357

RESUMO

The impact of curative radiotherapy depends mainly on the total dose delivered homogenously in the target volume. Tumor sensitivity to radiotherapy may be particularly inconstant depending on location, histology, somatic genetic parameters and the capacity of the immune system to infiltrate the tumor. In addition, the dose delivered to the surrounding healthy tissues may reduce the therapeutic ratio of many radiation treatments. In a same population treated in one center with the same technique, it appears that individual radiosensitivity clearly exists, namely in terms of late side effects that are in principle non-reversible. This review details the different radiobiological approaches that have been developed to better predict the tumor response but also the radiation-induced late effects.


Assuntos
Neoplasias/radioterapia , Órgãos em Risco/efeitos da radiação , Tolerância a Radiação , Biomarcadores Tumorais , Células Sanguíneas/efeitos da radiação , Reparo do DNA/genética , Humanos , Neoplasias/genética , Especificidade de Órgãos , Prognóstico , Proteômica , Lesões por Radiação/etiologia , Dosagem Radioterapêutica , Sensibilidade e Especificidade , Resultado do Tratamento
5.
Genes Dev ; 33(17-18): 1175-1190, 2019 09 01.
Artigo em Inglês | MEDLINE | ID: mdl-31395742

RESUMO

The ribosomal DNA (rDNA) represents a particularly unstable locus undergoing frequent breakage. DNA double-strand breaks (DSBs) within rDNA induce both rDNA transcriptional repression and nucleolar segregation, but the link between the two events remains unclear. Here we found that DSBs induced on rDNA trigger transcriptional repression in a cohesin- and HUSH (human silencing hub) complex-dependent manner throughout the cell cycle. In S/G2 cells, transcriptional repression is further followed by extended resection within the interior of the nucleolus, DSB mobilization at the nucleolar periphery within nucleolar caps, and repair by homologous recombination. We showed that nuclear envelope invaginations frequently connect the nucleolus and that rDNA DSB mobilization, but not transcriptional repression, involves the nuclear envelope-associated LINC complex and the actin pathway. Altogether, our data indicate that rDNA break localization at the nucleolar periphery is not a direct consequence of transcriptional repression but rather is an active process that shares features with the mobilization of persistent DSB in active genes and heterochromatin.


Assuntos
Proteínas de Ciclo Celular/metabolismo , Proteínas Cromossômicas não Histona/metabolismo , Quebras de DNA de Cadeia Dupla , Reparo do DNA/genética , DNA Ribossômico/genética , Regulação da Expressão Gênica/genética , RNA Longo não Codificante/metabolismo , Nucléolo Celular/metabolismo , Histonas/metabolismo , Recombinação Homóloga/genética , Membrana Nuclear/metabolismo
6.
Genes Dev ; 33(19-20): 1397-1415, 2019 10 01.
Artigo em Inglês | MEDLINE | ID: mdl-31467087

RESUMO

DNA repair by homologous recombination (HR) is essential for genomic integrity, tumor suppression, and the formation of gametes. HR uses DNA synthesis to repair lesions such as DNA double-strand breaks and stalled DNA replication forks, but despite having a good understanding of the steps leading to homology search and strand invasion, we know much less of the mechanisms that establish recombination-associated DNA polymerization. Here, we report that C17orf53/HROB is an OB-fold-containing factor involved in HR that acts by recruiting the MCM8-MCM9 helicase to sites of DNA damage to promote DNA synthesis. Mice with targeted mutations in Hrob are infertile due to depletion of germ cells and display phenotypes consistent with a prophase I meiotic arrest. The HROB-MCM8-MCM9 pathway acts redundantly with the HELQ helicase, and cells lacking both HROB and HELQ have severely impaired HR, suggesting that they underpin two major routes for the completion of HR downstream from RAD51. The function of HROB in HR is reminiscent of that of gp59, which acts as the replicative helicase loader during bacteriophage T4 recombination-dependent DNA replication. We therefore propose that the loading of MCM8-MCM9 by HROB may similarly be a key step in the establishment of mammalian recombination-associated DNA synthesis.


Assuntos
Reparo do DNA/genética , Recombinação Homóloga/genética , Proteínas de Manutenção de Minicromossomo/metabolismo , Animais , Linhagem Celular , DNA Helicases/metabolismo , Feminino , Células HCT116 , Células HEK293 , Células HeLa , Humanos , Infertilidade/genética , Masculino , Camundongos Endogâmicos C57BL , Deleção de Sequência , Células Sf9
7.
Int J Occup Environ Med ; 10(3): 124-136, 2019 07.
Artigo em Inglês | MEDLINE | ID: mdl-31325295

RESUMO

BACKGROUND: Coke oven workers are exposed to polycyclic aromatic hydrocarbons (PAHs) with possible genotoxicity and carcinogenicity. Metabolizing enzymes genes and DNA repair genes are suspected to be correlated with the level of DNA damage. They may contribute to variable individual sensitivity to DNA damage induced by PAHs exposure at workplace. OBJECTIVE: To investigate the relationship between biomarkers of PAHs: 1-hydroxypyrene (1-OHP), DNA adducts, and 8-hydroxy-2-deoxyguanosine (8-OHdG) in coke oven workers, and to assess the role of cytochrome P2E1 (CYP2E1) gene expression and DNA repairing gene (XRCC1) polymorphism in detecting workers at risk. METHODS: 85 exposed workers and 85 unexposed controls were enrolled into this study. Urinary 1-OHP, 8-OHdG, and BPDE-DNA adduct were measured. CYP2E1 gene expression and genotyping of XRCC1 399 Arg/Gln were evaluated by real-time PCR. RESULTS: The median urinary 1-OHP levels (6.3 µmol/mol creatinine), urinary 8-OHdG (7.9 ng/mg creatinine), DNA adducts (6.7 ng/µg DNA) in the exposed group were significantly higher than those in the unexposed group. Carriers of the variant allele (Gln) of XRCC1 had the highest levels of 1-OHP, DNA adducts and 8-OHdG, and the lowest level of CYP2E1 gene expression. In exposed workers, significant positive correlations were found between 1-OHP level and each of the work duration, 8-OHdG, and DNA adducts levels. There was a significant negative correlation between 1-OHP level and CYP2E1 gene expression. Work duration and CYP2E1 gene expression were predictors of DNA adducts level; 1-OHP level and work duration were predictors of urinary 8-OHdG level. CONCLUSION: Workers with higher exposure to PAH were more prone to oxidative DNA damage and cancer development. DNA adducts level reflects the balance between their production by CYP2E1 and elimination by XRCC1 gene.


Assuntos
Citocromo P-450 CYP2E1/genética , Adutos de DNA/genética , Desoxiguanosina/análogos & derivados , Monitoramento Ambiental/métodos , Exposição Ocupacional/análise , Pirenos/urina , Proteína 1 Complementadora Cruzada de Reparo de Raio-X/genética , 7,8-Di-Hidro-7,8-Di-Hidroxibenzo(a)pireno 9,10-óxido , Adulto , Biomarcadores/urina , Coque , Citocromo P-450 CYP2E1/biossíntese , Adutos de DNA/urina , Dano ao DNA/efeitos dos fármacos , Reparo do DNA/genética , Desoxiguanosina/urina , Egito , Humanos , Masculino , Pessoa de Meia-Idade , Hidrocarbonetos Policíclicos Aromáticos/urina , Polimorfismo Genético , Medição de Risco , Proteína 1 Complementadora Cruzada de Reparo de Raio-X/biossíntese , Adulto Jovem
8.
Microbes Environ ; 34(3): 316-326, 2019 Sep 25.
Artigo em Inglês | MEDLINE | ID: mdl-31353332

RESUMO

Thermococcus kodakarensis possesses two DNA polymerases, Pol B and Pol D. We generated a T. kodakarensis strain (DPB1) in which polB was completely deleted and a derivative of DPB1 in which polB was overexpressed; neither of the generated strains exhibited any growth delay, indicating that the lack or overexpression of Pol B in T. kodakarensis did not affect cell growth. We also found that DPB1 showed higher sensitivity to four DNA-damaging agents (ultraviolet C irradiation, γ-ray irradiation, methyl methanesulfonate, and mitomycin C) than the parental strain. The sensitivity of DPB1 was restored to the level of the parent strain by the introduction of a plasmid harboring polB, suggesting that the DNA damage-sensitive phenotype of DPB1 was due to the loss of polB. Collectively, these results indicate that Pol B is involved in DNA repair, but not DNA replication, which, in turn, implies that Pol D is the sole replicative DNA polymerase in Thermococcus species.


Assuntos
Reparo do DNA/genética , DNA Arqueal/genética , DNA Polimerase Dirigida por DNA/genética , Thermococcus/enzimologia , Thermococcus/genética , Proteínas de Bactérias/genética , Dano ao DNA/efeitos dos fármacos , Dano ao DNA/genética , Reparo do DNA/efeitos dos fármacos , Replicação do DNA , Deleção de Genes , Expressão Gênica , Inibidores da Síntese de Ácido Nucleico/farmacologia , Thermococcus/efeitos dos fármacos
9.
Cancer Sci ; 110(8): 2620-2628, 2019 Aug.
Artigo em Inglês | MEDLINE | ID: mdl-31152682

RESUMO

Tumor mutational burden (TMB) and mutational signatures reflect the process of mutation accumulation in cancer. However, the significance of these emerging characteristics remains unclear. In the present study, we used whole-exome sequencing to analyze the TMB and mutational signature in solid tumors of 4046 Japanese patients. Eight predominant signatures-microsatellite instability, smoking, POLE, APOBEC, UV, mismatch repair, double-strand break repair, and Signature 16-were observed in tumors with TMB higher than 1.0 mutation/Mb, whereas POLE and UV signatures only showed moderate correlation with TMB, suggesting the extensive accumulation of mutations due to defective POLE and UV exposure. The contribution ratio of Signature 16, which is associated with hepatocellular carcinoma in drinkers, was increased in hypopharynx cancer. Tumors with predominant microsatellite instability signature were potential candidates for treatment with immune checkpoint inhibitors such as pembrolizumab and were found in 2.8% of cases. Furthermore, based on microarray analysis, tumors with predominant signatures were classified into 2 subgroups depending on the expression of immune-related genes reflecting differences in the immune context of the tumor microenvironment. Tumor subpopulations differing in the content of infiltrating immune cells might respond differently to immunotherapeutics. An understanding of cancer characteristics based on TMB and mutational signatures could provide new insights into mutation-driven tumorigenesis.


Assuntos
Carcinogênese/genética , Mutação/genética , Neoplasias/genética , Carcinogênese/patologia , Reparo de Erro de Pareamento de DNA/genética , Reparo do DNA/genética , Genoma Humano/genética , Sequenciamento de Nucleotídeos em Larga Escala/métodos , Humanos , Japão , Instabilidade de Microssatélites , Neoplasias/patologia , Carga Tumoral/genética , Microambiente Tumoral/genética , Sequenciamento Completo do Exoma/métodos
10.
Mol Carcinog ; 58(9): 1531-1550, 2019 09.
Artigo em Inglês | MEDLINE | ID: mdl-31168912

RESUMO

Obesity, defined as a state of positive energy balance with a body mass index exceeding 30 kg/m2 in adults and 95th percentile in children, is an increasing global concern. Approximately one-third of the world's population is overweight or obese, and in the United States alone, obesity affects one in six children. Meta-analysis studies suggest that obesity increases the likelihood of developing several types of cancer, and with poorer outcomes, especially in children. The contribution of obesity to cancer risk requires a better understanding of the association between obesity-induced metabolic changes and its impact on genomic instability, which is a major driving force of tumorigenesis. In this review, we discuss how molecular changes during adipose tissue dysregulation can result in oxidative stress and subsequent DNA damage. This represents one of the many critical steps connecting obesity and cancer since oxidative DNA lesions can result in cancer-associated genetic instability. In addition, the by-products of the oxidative degradation of lipids (e.g., malondialdehyde, 4-hydroxynonenal, and acrolein), and gut microbiota-mediated secondary bile acid metabolites (e.g., deoxycholic acid and lithocholic acid), can function as genotoxic agents and tumor promoters. We also discuss how obesity can impact DNA repair efficiency, potentially contributing to cancer initiation and progression. Finally, we outline obesity-related epigenetic changes and identify the gaps in knowledge to be addressed for the development of better therapeutic strategies for the prevention and treatment of obesity-related cancers.


Assuntos
Instabilidade Genômica/genética , Neoplasias/genética , Neoplasias/metabolismo , Obesidade/genética , Obesidade/metabolismo , Tecido Adiposo/metabolismo , Animais , Dano ao DNA/genética , Reparo do DNA/genética , Metabolismo Energético/genética , Epigênese Genética/genética , Humanos , Estresse Oxidativo/genética
11.
PLoS Genet ; 15(6): e1008177, 2019 06.
Artigo em Inglês | MEDLINE | ID: mdl-31170160

RESUMO

During meiotic prophase I, double-strand breaks (DSBs) initiate homologous recombination leading to non-crossovers (NCOs) and crossovers (COs). In mouse, 10% of DSBs are designated to become COs, primarily through a pathway dependent on the MLH1-MLH3 heterodimer (MutLγ). Mlh3 contains an endonuclease domain that is critical for resolving COs in yeast. We generated a mouse (Mlh3DN/DN) harboring a mutation within this conserved domain that is predicted to generate a protein that is catalytically inert. Mlh3DN/DN males, like fully null Mlh3-/- males, have no spermatozoa and are infertile, yet spermatocytes have grossly normal DSBs and synapsis events in early prophase I. Unlike Mlh3-/- males, mutation of the endonuclease domain within MLH3 permits normal loading and frequency of MutLγ in pachynema. However, key DSB repair factors (RAD51) and mediators of CO pathway choice (BLM helicase) persist into pachynema in Mlh3DN/DN males, indicating a temporal delay in repair events and revealing a mechanism by which alternative DSB repair pathways may be selected. While Mlh3DN/DN spermatocytes retain only 22% of wildtype chiasmata counts, this frequency is greater than observed in Mlh3-/- males (10%), suggesting that the allele may permit partial endonuclease activity, or that other pathways can generate COs from these MutLγ-defined repair intermediates in Mlh3DN/DN males. Double mutant mice homozygous for the Mlh3DN/DN and Mus81-/- mutations show losses in chiasmata close to those observed in Mlh3-/- males, indicating that the MUS81-EME1-regulated crossover pathway can only partially account for the increased residual chiasmata in Mlh3DN/DN spermatocytes. Our data demonstrate that mouse spermatocytes bearing the MLH1-MLH3DN/DN complex display the proper loading of factors essential for CO resolution (MutSγ, CDK2, HEI10, MutLγ). Despite these functions, mice bearing the Mlh3DN/DN allele show defects in the repair of meiotic recombination intermediates and a loss of most chiasmata.


Assuntos
Proteínas de Ligação a DNA/genética , Endonucleases/genética , Prófase Meiótica I/genética , Proteínas MutL/genética , Animais , Pareamento Cromossômico/genética , Troca Genética , Quebras de DNA de Cadeia Dupla , Reparo do DNA/genética , Recombinação Homóloga/genética , Masculino , Meiose/genética , Camundongos , Proteína 1 Homóloga a MutL/genética , Proteínas MutS/genética , Rad51 Recombinase/genética , Espermatócitos/crescimento & desenvolvimento , Espermatócitos/metabolismo
12.
Mol Cell ; 74(5): 866-876, 2019 06 06.
Artigo em Inglês | MEDLINE | ID: mdl-31173722

RESUMO

The replisome quickly and accurately copies billions of DNA bases each cell division cycle. However, it can make errors, especially when the template DNA is damaged. In these cases, replication-coupled repair mechanisms remove the mistake or repair the template lesions to ensure high fidelity and complete copying of the genome. Failures in these genome maintenance activities generate mutations, rearrangements, and chromosome segregation problems that cause many human diseases. In this review, I provide a broad overview of replication-coupled repair pathways, explaining how they fix polymerase mistakes, respond to template damage that acts as obstacles to the replisome, deal with broken forks, and impact human health and disease.


Assuntos
Reparo do DNA/genética , Replicação do DNA/genética , Doenças Genéticas Inatas/genética , Genoma Humano/genética , Ciclo Celular/genética , Segregação de Cromossomos/genética , Dano ao DNA/genética , Instabilidade Genômica/genética , Humanos , Mutação/genética
13.
Nat Commun ; 10(1): 2491, 2019 06 06.
Artigo em Inglês | MEDLINE | ID: mdl-31171785

RESUMO

Genetic factors underlying leukocyte telomere length (LTL) may provide insights into telomere homeostasis, with direct links to disease susceptibility. Genetic evaluation of 23,096 Singaporean Chinese samples identifies 10 genome-wide loci (P < 5 × 10-8). Several of these contain candidate genes (TINF2, PARP1, TERF1, ATM and POT1) with potential roles in telomere biology and DNA repair mechanisms. Meta-analyses with additional 37,505 European individuals reveals six more genome-wide loci, including associations at MPHOSPH6, NKX2-3 and TYMS. We demonstrate that longer LTL associates with protection against respiratory disease mortality [HR = 0.854(0.804-0.906), P = 1.88 × 10-7] in the Singaporean Chinese samples. We further show that the LTL reducing SNP rs7253490 associates with respiratory infections (P = 7.44 × 10-4) although this effect may not be strongly mediated through LTL. Our data expands on the genetic basis of LTL and may indicate on a potential role of LTL in immune competence.


Assuntos
Grupo com Ancestrais do Continente Asiático/genética , Reparo do DNA/genética , Leucócitos/metabolismo , Homeostase do Telômero/genética , Telômero/metabolismo , Adulto , Idoso , Idoso de 80 Anos ou mais , Estudos de Coortes , Grupo com Ancestrais do Continente Europeu/genética , Feminino , Estudo de Associação Genômica Ampla , Humanos , Masculino , Pessoa de Meia-Idade , Estudos Prospectivos , Infecções Respiratórias/genética , Singapura , Adulto Jovem
14.
Biomed Res Int ; 2019: 3842312, 2019.
Artigo em Inglês | MEDLINE | ID: mdl-31058188

RESUMO

There are about 1-2 million follicles presented in the ovary at birth, while only around 1000 primordial follicles are left at menopause. The ovarian function also decreases in parallel with aging. Folliculogenesis is vital for ovarian function, no matter the synthesis of female hormones or ovulation, yet the mechanisms for its changing with increasing age are not fully understood. Early follicle growth up to the large preantral stage is independent of gonadotropins in rodents and relies on intraovarian factors. To further understand the age-related molecular changes in the process of folliculogenesis, we performed microarray gene expression profile analysis using total RNA extracted from young (9 weeks old) and old (32 weeks old) mouse ovarian secondary follicles. The results of our current microarray study revealed that there were 371 (≥2-fold, q-value ≤0.05) genes differentially expressed in which 174 genes were upregulated and 197 genes were downregulated in old mouse ovarian secondary follicles compared to young mouse ovarian secondary follicles. The gene ontology and KEGG pathway analysis of differentially expressed genes uncovered critical biological functions such as immune system process, aging, transcription, DNA replication, DNA repair, protein stabilization, and apoptotic process were affected in the process of aging. The considerable changes in gene expression profile may have an adverse influence on follicle quality and folliculogenesis. Our study provided information on the processes that may contribute to age-related decline in ovarian function.


Assuntos
Envelhecimento/genética , Folículo Ovariano/crescimento & desenvolvimento , Ovário/crescimento & desenvolvimento , RNA/genética , Animais , Reparo do DNA/genética , Replicação do DNA/genética , Feminino , Regulação da Expressão Gênica no Desenvolvimento/genética , Humanos , Menopausa/genética , Camundongos , Oócitos/crescimento & desenvolvimento , Oócitos/metabolismo , Folículo Ovariano/metabolismo , Ovário/metabolismo , Ovulação/genética , RNA/biossíntese , Transcriptoma/genética
15.
Mol Carcinog ; 58(9): 1648-1655, 2019 09.
Artigo em Inglês | MEDLINE | ID: mdl-31140647

RESUMO

SMAD4 is a potent tumor suppressor and a central mediator of the TGFß signaling pathway. SMAD4 genetic loss is frequent in squamous cell carcinomas (SCCs). Reports of SMAD4 expression in SCCs vary significantly possibly due to inter-tumor heterogeneity or technical reasons. SMAD4 loss is an initiation event for SCCs. In tumor epithelial cells, SMAD4 loss causes increased proliferation, decreased apoptosis, and "Brca-like" genomic instability associated with DNA repair defects. SMAD4 loss also plays a role in the expansion of cancer stem cells. Epithelial SMAD4 loss causes overexpression of TGFß that is released into the tumor microenvironment and contributes to SCC progression through proinflammatory and immune evasive mechanisms. SMAD4 loss, while not a direct therapeutic target, is associated with multiple targetable pathways that require further therapeutic studies. Altogether, SMAD4 loss is a potential biomarker in SCCs that should be further studied for its values in prognostic and therapeutic predictions. Such information will potentially guide future biomarker-driven clinical trial designs and improve SCC patient outcomes.


Assuntos
Carcinoma de Células Escamosas/genética , Proteína Smad4/genética , Animais , Biomarcadores Tumorais/genética , Reparo do DNA/genética , Células Epiteliais/patologia , Genes Supressores de Tumor/fisiologia , Humanos , Transdução de Sinais/genética , Microambiente Tumoral/genética
16.
Oncol Rep ; 41(6): 3201-3208, 2019 Jun.
Artigo em Inglês | MEDLINE | ID: mdl-31002369

RESUMO

Intratumoral heterogeneity, particularly the potential cancer stemness of single cancer cells, has not yet been fully elucidated in human esophageal cancer. Single­cell transcriptome sequencing of two types of esophageal adenocarcinoma (EAC) and two types of esophageal squamous cell carcinoma (ESCC) tissues was performed, and the intratumoral cancer stemness of the types of esophageal cancer were characterized at the single­cell level in the present study. By comparing the transcriptomic profiles of single cancer cells with high and low stemness in individual patients, it was revealed that the overexpression of cell cycle­associated genes in EAC cells was highly correlated with stemness, whereas overexpression of genes involved in the signaling pathways of DNA replication and DNA damage repair was significantly correlated with stemness in ESCC. High expression of these stemness­associated genes was correlated with poor prognosis of patients. Additionally, poly [ADP­ribose] polymerase(PARP)4 was identified as a novel cancer stemness­associated gene in ESCC and its association with survival was validated in a cohort of 121 patients with ESCC. These findings have profound potential implications for the use of cell cycle inhibitors in EAC and PARP inhibitors in ESCC, which may provide novel mechanistic insights into the plasticity of esophageal cancer.


Assuntos
Adenocarcinoma/genética , Biomarcadores Tumorais/genética , Carcinoma de Células Escamosas do Esôfago/genética , Proteínas Nucleares/genética , Adenocarcinoma/classificação , Adenocarcinoma/diagnóstico , Adenocarcinoma/patologia , Pontos de Checagem do Ciclo Celular/genética , Linhagem Celular Tumoral , Reparo do DNA/genética , Intervalo Livre de Doença , Carcinoma de Células Escamosas do Esôfago/classificação , Carcinoma de Células Escamosas do Esôfago/diagnóstico , Carcinoma de Células Escamosas do Esôfago/patologia , Feminino , Regulação Neoplásica da Expressão Gênica/genética , Humanos , Masculino , Células-Tronco Neoplásicas/metabolismo , Células-Tronco Neoplásicas/patologia , Inibidores de Poli(ADP-Ribose) Polimerases/uso terapêutico , Análise de Célula Única , Transcriptoma/genética , Sequenciamento Completo do Exoma
17.
Orphanet J Rare Dis ; 14(1): 82, 2019 04 17.
Artigo em Inglês | MEDLINE | ID: mdl-30995915

RESUMO

BACKGROUND: Telomeres are nucleoprotein structures present at the terminal region of the chromosomes. Mutations in genes coding for proteins involved in telomere maintenance are causative of a number of disorders known as telomeropathies. The genetic origin of these diseases is heterogeneous and has not been determined for a significant proportion of patients. METHODS: This article describes the genetic characterization of a cohort of patients. Telomere length was determined by Southern blot and quantitative PCR. Nucleotide variants were analyzed either by high-resolution melting analysis and Sanger sequencing of selected exons or by massive sequencing of a panel of genes. RESULTS: Forty-seven patients with telomere length below the 10% of normal population, affected with three telomeropathies: dyskeratosis congenita (4), aplastic anemia (22) or pulmonary fibrosis (21) were analyzed. Eighteen of these patients presented known pathogenic or novel possibly pathogenic variants in the telomere-related genes TERT, TERC, RTEL1, CTC1 and ACD. In addition, the analyses of a panel of 188 genes related to haematological disorders indicated that a relevant proportion of the patients (up to 35%) presented rare variants in genes related to DNA repair or in genes coding for proteins involved in the resolution of complex DNA structures, that participate in telomere replication. Mutations in some of these genes are causative of several syndromes previously associated to telomere shortening. CONCLUSION: Novel variants in telomere, DNA repair and replication genes are described that might indicate the contribution of variants in these genes to the development of telomeropathies. Patients carrying variants in telomere-related genes presented worse evolution after diagnosis than the rest of patients analyzed.


Assuntos
Anemia Aplástica/genética , Reparo do DNA/genética , Disceratose Congênita/genética , Fibrose Pulmonar/genética , Encurtamento do Telômero/genética , Telômero/genética , Adolescente , Adulto , Criança , Pré-Escolar , Éxons/genética , Feminino , Humanos , Lactente , Masculino , Linhagem , RNA/genética , Telomerase/genética , Adulto Jovem
18.
Nat Protoc ; 14(5): 1489-1508, 2019 05.
Artigo em Inglês | MEDLINE | ID: mdl-30962605

RESUMO

Non-coding RNA (ncRNA) molecules have been shown to play a variety of cellular roles; however, the contributions of different types of RNA to specific phenomena are often hard to dissect. To study the role of RNA in the assembly of DNA damage response (DDR) foci, we developed the RNase A treatment and reconstitution (RATaR) method, in which cells are mildly permeabilized, incubated with recombinant RNase A and subsequently reconstituted with different RNA species, under conditions of RNase A inactivation and inhibition of endogenous transcription. The block of transcription right after RNase A removal represents a key innovation of RATaR, preventing potential contributions of endogenously neo-synthesized transcripts to the phenotypes studied. A critical aspect of this technique is the balance between sufficient permeabilization of membranes to allow enzyme/RNA access into the cell nucleus and cell viability. Here, we present our protocol for RNA-dependent DDR foci disassembly and reassembly using fluorescent DDR RNAs (DDRNAs) in NIH2/4 cells, an engineered NIH3T3-derived cell line. The use of sequence-specific, fluorescent RNA molecules permits the concomitant determination of their subcellular localization and biological functions. We also outline adaptations of RATaR when implemented in different cell lines exposed to various genotoxic treatments, such as γ-radiation, restriction enzymes and telomere deprotection. In all these cases, the entire procedure can be completed within 2 h without the need for special equipment or uncommon skills. We believe this technique will prove useful for investigating the contribution of RNA to a variety of relevant cellular processes.


Assuntos
Dano ao DNA , Reparo do DNA , RNA não Traduzido , Ribonuclease Pancreático/metabolismo , Animais , Dano ao DNA/genética , Dano ao DNA/fisiologia , Reparo do DNA/genética , Reparo do DNA/fisiologia , Técnicas Genéticas , Células HeLa , Humanos , Camundongos , Células NIH 3T3 , RNA/análise , RNA/genética , RNA/metabolismo , RNA não Traduzido/genética , RNA não Traduzido/fisiologia
19.
Mol Cell ; 74(3): 466-480.e4, 2019 05 02.
Artigo em Inglês | MEDLINE | ID: mdl-30930055

RESUMO

The mTOR pathway integrates both extracellular and intracellular signals and serves as a central regulator of cell metabolism, growth, survival, and stress responses. Neurotropic viruses, such as herpes simplex virus-1 (HSV-1), also rely on cellular AKT-mTORC1 signaling to achieve viral latency. Here, we define a novel genotoxic response whereby spatially separated signals initiated by extracellular neurotrophic factors and nuclear DNA damage are integrated by the AKT-mTORC1 pathway. We demonstrate that endogenous DNA double-strand breaks (DSBs) mediated by Topoisomerase 2ß-DNA cleavage complex (TOP2ßcc) intermediates are required to achieve AKT-mTORC1 signaling and maintain HSV-1 latency in neurons. Suppression of host DNA-repair pathways that remove TOP2ßcc trigger HSV-1 reactivation. Moreover, perturbation of AKT phosphorylation dynamics by downregulating the PHLPP1 phosphatase led to AKT mis-localization and disruption of DSB-induced HSV-1 reactivation. Thus, the cellular genome integrity and environmental inputs are consolidated and co-opted by a latent virus to balance lifelong infection with transmission.


Assuntos
DNA Topoisomerases Tipo II/genética , Herpesvirus Humano 1/genética , Proteínas Nucleares/genética , Proteínas Proto-Oncogênicas c-akt/genética , Latência Viral/genética , Animais , Quebras de DNA de Cadeia Dupla , Dano ao DNA/genética , Reparo do DNA por Junção de Extremidades/genética , Reparo do DNA/genética , Enzimas Reparadoras do DNA/genética , Proteínas de Ligação a DNA/genética , Herpesvirus Humano 1/patogenicidade , Humanos , Proteína Homóloga a MRE11/genética , Alvo Mecanístico do Complexo 1 de Rapamicina/genética , Neurônios/metabolismo , Neurônios/virologia , Fosforilação , Ratos , Transdução de Sinais/genética , Serina-Treonina Quinases TOR/genética
20.
Arch Microbiol ; 201(7): 991-998, 2019 Sep.
Artigo em Inglês | MEDLINE | ID: mdl-31025056

RESUMO

Caffeine is a compound that can exert physiological-beneficial effects in the organism. Nevertheless, there are controversies about its protective-antioxidant and/or its negative genotoxic effect. To abound on the analysis of the possible genotoxic/antioxidant effect of caffeine, we used as research model the yeast Yarrowia lipolytica parental strain, and mutant strains (∆rad52 and ∆ku80), which are deficient in the DNA repair mechanisms. Caffeine (5 mM) showed a cytostatic effect on all strains, but after 72 h of incubation the parental and ∆ku80 strains were able to recover of this inhibitory effect on growth, whereas ∆rad52 was unable to recover. When cells were pre-incubated with caffeine and H2O2 or incubated with a mixture of both agents, a higher inhibitory effect on growth of mutant strains was observed and this effect was noticeably greater for the Δrad52 strain. The toxic effect of caffeine appears to be through a mechanism of DNA damage (genotoxic effect) that involves DSB generation since, in all tested conditions, the growth of Δrad52 strain (cells deficient in HR DNA repair mechanism) was more severely affected.


Assuntos
Cafeína/toxicidade , Reparo do DNA/genética , Yarrowia/efeitos dos fármacos , Quebras de DNA de Cadeia Dupla/efeitos dos fármacos , Reparo do DNA/efeitos dos fármacos , Peróxido de Hidrogênio/farmacologia , Mutação/genética , Yarrowia/genética , Yarrowia/crescimento & desenvolvimento
SELEÇÃO DE REFERÊNCIAS
DETALHE DA PESQUISA