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1.
Nutrients ; 15(7)2023 Mar 31.
Artículo en Inglés | MEDLINE | ID: mdl-37049562

RESUMEN

Adipose tissue (AT) dysregulation is a key process in the pathophysiology of obesity and its cardiometabolic complications, but even if a growing body of evidence has been collected over recent decades, the underlying molecular basis of adiposopathy remains to be fully understood. In this context, mitochondria, the intracellular organelles that orchestrate energy production and undergo highly dynamic adaptive changes in response to changing environments, have emerged as crucial regulators of both white (WAT) and brown adipose tissue (BAT) metabolism and function. Given that the gut microbiota and its metabolites are able to regulate host metabolism, adipogenesis, WAT inflammation, and thermogenesis, we hypothesize that their frequently observed dysregulation in obesity could affect AT metabolism by exerting direct and indirect effects on AT mitochondria. By collecting and revising the current evidence on the connections between gut microbiota and AT mitochondria in obesity, we gained insights into the molecular biology of their hitherto largely unexplored crosstalk, tracing how gut microbiota may regulate AT mitochondrial function.


Asunto(s)
Microbioma Gastrointestinal , Humanos , Microbioma Gastrointestinal/fisiología , Obesidad/metabolismo , Tejido Adiposo Blanco/metabolismo , Tejido Adiposo Pardo/metabolismo , Mitocondrias/metabolismo , Termogénesis/fisiología , Metabolismo Energético
2.
Sci Rep ; 12(1): 18877, 2022 11 07.
Artículo en Inglés | MEDLINE | ID: mdl-36344536

RESUMEN

Bariatric surgery (BS) is an effective intervention for severe obesity and associated comorbidities. Although several studies have addressed the clinical and metabolic effects of BS, an integrative analysis of the complex body response to surgery is still lacking. We conducted a longitudinal data study with 36 patients with severe obesity who were tested before, 6 and 12 months after restrictive BS for more than one hundred blood biomarkers, including clinical, oxidative stress and metabolic markers, peptide mediators and red blood cell membrane lipids. By using a synthetic data-driven modeling based on principal component and correlation analyses, we provided evidence that, besides the early, well-known glucose metabolism- and weight loss-associated beneficial effects of BS, a tardive, weight-independent increase of the hepatic cholesterol metabolism occurs that is associated with potentially detrimental inflammatory and metabolic effects. Canonical correlation analysis indicated that oxidative stress is the most predictive feature of the BS-induced changes of both glucose and lipids metabolism. Our results show the power of multi-level correlation analysis to uncover the network of biological pathways affected by BS. This approach highlighted potential health risks of restrictive BS that are disregarded with the current practice to use weight loss as surrogate of BS success.


Asunto(s)
Cirugía Bariátrica , Obesidad Mórbida , Humanos , Cirugía Bariátrica/métodos , Pérdida de Peso/fisiología , Aumento de Peso , Medición de Riesgo
3.
Oncotarget ; 8(49): 84827-84840, 2017 Oct 17.
Artículo en Inglés | MEDLINE | ID: mdl-29156686

RESUMEN

DNA repair gene expression in a set of gastric cancers suggested an inverse association between the expression of the mismatch repair (MMR) gene MLH1 and that of the base excision repair (BER) gene DNA polymerase ß (Polß). To gain insight into possible crosstalk of these two repair pathways in cancer, we analysed human gastric adenocarcinoma AGS cells over-expressing Polß or Polß active site mutants, alone or in combination with MLH1 silencing. Next, we investigated the cellular response to the alkylating agent methyl methanesulfonate (MMS) and the purine analogue 6-thioguanine (6-TG), agents that induce lesions that are substrates for BER and/or MMR. AGS cells over-expressing Polß were resistant to 6-TG to a similar extent as when MLH1 was inactivated while inhibition of O6-methylguanine-DNA methyltransferase (MGMT) was required to detect resistance to MMS. Upon either treatment, the association with MLH1 down-regulation further amplified the resistant phenotype. Moreover, AGS cells mutated in Polß were hypersensitive to both 6-TG and MMS killing and their sensitivity was partially rescued by MLH1 silencing. We provide evidence that the critical lethal lesions in this new pathway are double strand breaks that are exacerbated when Polß is defective and relieved when MLH1 is silenced. In conclusion, we provide evidence of crosstalk between MLH1 and Polß that modulates the response to alkylation damage. These studies suggest that the Polß/MLH1 status should be taken into consideration when designing chemotherapeutic approaches for gastric cancer.

4.
Free Radic Biol Med ; 107: 278-291, 2017 06.
Artículo en Inglés | MEDLINE | ID: mdl-27932076

RESUMEN

Oxidative stress is associated with a growing number of diseases that span from cancer to neurodegeneration. Most oxidatively induced DNA base lesions are repaired by the base excision repair (BER) pathway which involves the action of various DNA glycosylases. There are numerous genome wide studies attempting to associate single-nucleotide polymorphisms (SNPs) with predispositions to various types of disease; often, these common variants do not have significant alterations in their biochemical function and do not exhibit a convincing phenotype. Nevertheless several lines of evidence indicate that SNPs in DNA repair genes may modulate DNA repair capacity and contribute to risk of disease. This overview provides a convincing picture that SNPs of DNA glycosylases that remove oxidatively generated DNA lesions are susceptibility factors for a wide disease spectrum that includes besides cancer (particularly lung, breast and gastrointestinal tract), cochlear/ocular disorders, myocardial infarction and neurodegenerative disorders which can be all grouped under the umbrella of oxidative stress-related pathologies.


Asunto(s)
Enfermedades Cocleares/genética , ADN Glicosilasas/genética , Reparación del ADN , Oftalmopatías/genética , Infarto del Miocardio/genética , Neoplasias/genética , Enfermedades Neurodegenerativas/genética , Daño del ADN , Predisposición Genética a la Enfermedad , Genotipo , Humanos , Estrés Oxidativo , Fenotipo , Polimorfismo de Nucleótido Simple
5.
Free Radic Biol Med ; 97: 236-243, 2016 08.
Artículo en Inglés | MEDLINE | ID: mdl-27264237

RESUMEN

Inefficient response to oxidative stress has been associated with ageing and health risk. Metals are known to inhibit DNA repair and may modify the antioxidant response. How genetic variability and lifestyle factors modulate the response to oxidative stress is poorly explored. Our study aims to disentangle the contribution of genetics and environmental exposures to oxidative stress response using data from twin pairs. The non-enzymatic antioxidant capacity (NEAC), the repair capacity of 8-oxo-7,8-dihydroguanine (OGG activity) and the levels of 12 metals were measured in blood of 64 monozygotic and 31 dizygotic twin pairs. The contributions of genetic and environmental effects were assessed using standard univariate twin modelling. NEAC and OGG activity significantly decreased with age. Gender-, age- and body mass index-associated differences were identified for some metals. Principal Component Analysis identified two groups of metals whose levels in blood were highly correlated: As, Hg, Pb, Se, Zn and Al, Co, Cr, Mn, Ni. The environmental influence was predominant on OGG activity and NEAC variance whereas for most metals the best-fitting model incorporated additive genetic and unique environmental sources of variance. NEAC and OGG activity were both inversely correlated with blood levels of various metals. The inhibition of OGG activity by Cd was largely explained by smoking. Our data show a substantial role of environmental factors in NEAC and OGG activity variance that is not explained by twins' age. Exogenous environmental factors such as metals contribute to oxidative stress by decreasing NEAC and inhibiting repair of oxidatively-induced DNA damage.


Asunto(s)
Contaminantes Ambientales/toxicidad , Estrés Oxidativo , Adulto , Antioxidantes/metabolismo , Biomarcadores/sangre , Daño del ADN , ADN Glicosilasas/sangre , Reparación del ADN , Exposición a Riesgos Ambientales , Femenino , Humanos , Masculino , Metales Pesados/sangre , Gemelos Dicigóticos , Gemelos Monocigóticos
6.
Neural Plast ; 2016: 3619274, 2016.
Artículo en Inglés | MEDLINE | ID: mdl-26942017

RESUMEN

There is a growing body of evidence indicating that the mechanisms that control genome stability are of key importance in the development and function of the nervous system. The major threat for neurons is oxidative DNA damage, which is repaired by the base excision repair (BER) pathway. Functional mutations of enzymes that are involved in the processing of single-strand breaks (SSB) that are generated during BER have been causally associated with syndromes that present important neurological alterations and cognitive decline. In this review, the plasticity of BER during neurogenesis and the importance of an efficient BER for correct brain function will be specifically addressed paying particular attention to the brain region and neuron-selectivity in SSB repair-associated neurological syndromes and age-related neurodegenerative diseases.


Asunto(s)
Encéfalo/metabolismo , Daño del ADN , Reparación del ADN , Enfermedades del Sistema Nervioso/genética , Neuronas/metabolismo , Estrés Oxidativo , Animales , Roturas del ADN de Cadena Simple , Humanos , Neurogénesis/genética
7.
Mutat Res ; 782: 34-43, 2015 Dec.
Artículo en Inglés | MEDLINE | ID: mdl-26546826

RESUMEN

Xeroderma pigmentosum (XP)-A patients are characterized by increased solar skin carcinogenesis and present also neurodegeneration. XPA deficiency is associated with defective nucleotide excision repair (NER) and increased basal levels of oxidatively induced DNA damage. In this study we search for the origin of increased levels of oxidatively generated DNA lesions in XP-A cell genome and then address the question of whether increased oxidative stress might drive genetic instability. We show that XP-A human primary fibroblasts present increased levels and different types of intracellular reactive oxygen species (ROS) as compared to normal fibroblasts, with O2₋• and H2O2 being the major reactive species. Moreover, XP-A cells are characterized by decreased reduced glutathione (GSH)/oxidized glutathione (GSSG) ratios as compared to normal fibroblasts. The significant increase of ROS levels and the alteration of the glutathione redox state following silencing of XPA confirmed the causal relationship between a functional XPA and the control of redox balance. Proton nuclear magnetic resonance (¹H NMR) analysis of the metabolic profile revealed a more glycolytic metabolism and higher ATP levels in XP-A than in normal primary fibroblasts. This perturbation of bioenergetics is associated with different morphology and response of mitochondria to targeted toxicants. In line with cancer susceptibility, XP-A primary fibroblasts showed increased spontaneous micronuclei (MN) frequency, a hallmark of cancer risk. The increased MN frequency was not affected by inhibition of ROS to normal levels by N-acetyl-L-cysteine.


Asunto(s)
Fibroblastos/metabolismo , Micronúcleos con Defecto Cromosómico , Estrés Oxidativo , Especies Reactivas de Oxígeno/metabolismo , Proteína de la Xerodermia Pigmentosa del Grupo A/metabolismo , Xerodermia Pigmentosa/genética , Células Cultivadas , Glutatión/metabolismo , Humanos , Potencial de la Membrana Mitocondrial , Pruebas de Micronúcleos , Mitocondrias/patología , Estrés Oxidativo/genética , Cultivo Primario de Células , Xerodermia Pigmentosa/metabolismo , Xerodermia Pigmentosa/patología , Proteína de la Xerodermia Pigmentosa del Grupo A/genética
8.
Free Radic Biol Med ; 63: 401-9, 2013 Oct.
Artículo en Inglés | MEDLINE | ID: mdl-23726996

RESUMEN

8-Oxoguanine DNA glycosylase (OGG) activity was measured by an in vitro assay in lymphocytes of healthy volunteers genotyped for various OGG1 polymorphisms. Only homozygous carriers of the polymorphic C326 allele showed a significantly lower OGG activity compared to the homozygous S326 genotype. The purified S326C OGG1 showed a decreased ability to complete the repair synthesis step in a base excision repair reaction reconstituted in vitro. The propensity of this variant to dimerize as well as its catalytic impairment were shown to be enhanced under oxidizing conditions. Mass spectrometry revealed that the extra cysteine of the variant protein is involved in disulfide bonds compatible with significant conformational changes and/or dimerization. We propose that the S326C OGG1 catalytic impairment and its susceptibility to dimerization and disulfide bond formation in an oxidizing environment all concur to decrease repair capacity. Consequently, the C326 homozygous carriers may be at increased risk of oxidative pathologies.


Asunto(s)
Daño del ADN/genética , ADN Glicosilasas/genética , Linfocitos/metabolismo , Estrés Oxidativo , Adulto , Alelos , ADN Glicosilasas/metabolismo , Reparación del ADN/genética , Femenino , Estudios de Asociación Genética , Homocigoto , Humanos , Masculino , Persona de Mediana Edad , Oxidación-Reducción , Polimorfismo de Nucleótido Simple , Factores de Riesgo
9.
Mutat Res ; 749(1-2): 73-9, 2013 Sep.
Artículo en Inglés | MEDLINE | ID: mdl-23727398

RESUMEN

Due to the great number of genes involved in DNA repair and the interactions among the pathways responsible for the repair of different types of DNA damage, there is an increasing need for simple and reliable approaches to phenotypically assess DNA repair capacity (DRC). The use of peripheral blood mononuclear cells (PBMCs) in DRC assays is particularly useful for human monitoring studies. However, in such studies it is not always possible to collect and process samples on the same day as the blood is taken. We performed a genotype-phenotype correlation study on DRC on 225 healthy subjects. Due to the large number of blood samples to be processed, PBMCs were either isolated and cryopreserved on the same day of blood collection (day 1) or on the following day after 24h blood storage at room temperature (day 2-RT). Samples processed in different days showed a significant difference in the DRC evaluated as 8-oxoguanine glycosylase activity (OGG assay) in cell extracts (p<0.0001) and as benzo[a]pyrene diol epoxide (BPDE)-induced damage repair by the comet assay (p=0.05). No apparent effect of the blood storage conditions on the outcome of γ-ray induced H2AX phosphorylation assay was reported. These results prompted us to further analyze the effects of blood storage conditions by performing a validation study. Three blood samples were simultaneously taken from ten healthy donors, PBMCs were isolated and cryopreserved as follows: immediately after blood collection (day 1); on the following day, after blood storage at RT (day 2-RT); or after blood storage at 4°C (day 2-4°C). DRC was then evaluated using phenotypic assays. The γ-ray induced H2AX phosphorylation assay has been confirmed as the only assay that showed good reproducibility independently of the blood storage conditions. The measurement of OGG assay was most affected by the blood storage conditions.


Asunto(s)
Conservación de la Sangre/métodos , Reparación del ADN/fisiología , Leucocitos Mononucleares/metabolismo , Adolescente , Adulto , Conservación de la Sangre/efectos adversos , Recolección de Muestras de Sangre/efectos adversos , Recolección de Muestras de Sangre/métodos , Ensayo Cometa , Criopreservación , Femenino , Estudios de Asociación Genética/normas , Humanos , Masculino , Persona de Mediana Edad , Temperatura , Adulto Joven
10.
Mutat Res ; 736(1-2): 104-16, 2012 Aug 01.
Artículo en Inglés | MEDLINE | ID: mdl-22732424

RESUMEN

Oxidative damage to DNA can cause mutations, and mutations can lead to cancer. DNA repair of oxidative damage should therefore play a pivotal role in defending humans against cancer. This is exemplified by the increased risk of colorectal cancer of patients with germ-line mutations of the oxidative damage DNA glycosylase MUTYH. In contrast to germ-line mutations in DNA repair genes, which cause a strong deficiency in DNA repair activity in all cell types, the role of single nucleotide polymorphisms (SNPs) in sporadic cancer is unclear also because deficiencies in DNA repair, if any, are expected to be much milder. Further slowing down progress are the paucity of accurate and reproducible functional assays and poor epidemiological design of many studies. This review will focus on the most common and widely studied SNPs of oxidative DNA damage repair proteins trying to bridge the information available on biochemical and structural features of the repair proteins with the functional effects of these variants and their potential impact on the pathogenesis of disease.

11.
Mutagenesis ; 27(1): 49-57, 2012 Jan.
Artículo en Inglés | MEDLINE | ID: mdl-21857007

RESUMEN

Recent evidences have highlighted an influence of micronutrients in the maintenance of telomere length (TL). In order to explore whether diet-related telomere shortening had any physiological relevance and was accompanied by significant damage in the genome, in the present study, TL was assessed by terminal restriction fragment (TRF) analysis in peripheral blood lymphocytes of 56 healthy subjects for which detailed information on dietary habits was available and data were compared \with the incidence of nucleoplasmic bridges (NPBs), a marker of chromosomal instability related to telomere dysfunction visualised with the cytokinesis-blocked micronucleus assay. To increase the capability to detect even slight impairment of telomere function, the incidence of NPBs was also evaluated on cells exposed in vitro to ionising radiation. Care was taken to control for potential confounding factors that might influence TL, viz. age, hTERT genotype and smoking status. Data showed that higher consumption of vegetables was related with significantly higher mean TL (P = 0.013); in particular, the analysis of the association between micronutrients and mean TL highlighted a significant role of antioxidant intake, especially beta-carotene, on telomere maintenance (P = 0.004). However, the diet-related telomere shortening did not result in associated increased spontaneous or radiation-induced NPBs. The distribution of TRFs was also analysed and a slight prevalence of radiation-induced NPBs (P = 0.03) was observed in subjects with higher amount of very short TRFs (<2 kb). The relative incidence of very short TRFs was positively associate with ageing (P = 0.008) but unrelated to vegetables consumption and daily intake of micronutrients, suggesting that the degree of telomere erosion related with low dietary intake of antioxidants observed in this study was not so extensive to lead to chromosome instability.


Asunto(s)
Inestabilidad Cromosómica , Dieta , Acortamiento del Telómero , Antioxidantes/administración & dosificación , Biomarcadores/análisis , Femenino , Genotipo , Humanos , Estilo de Vida , Linfocitos/patología , Masculino , Pruebas de Micronúcleos , Micronutrientes/administración & dosificación , Persona de Mediana Edad , Estrés Oxidativo/efectos de los fármacos , Análisis de Secuencia de ADN , Fumar/efectos adversos , Encuestas y Cuestionarios , Telómero/patología , Verduras , beta Caroteno/administración & dosificación
12.
Mutat Res ; 731(1-2): 1-13, 2012 Mar 01.
Artículo en Inglés | MEDLINE | ID: mdl-22155132

RESUMEN

Oxidative damage to DNA can cause mutations, and mutations can lead to cancer. DNA repair of oxidative damage should therefore play a pivotal role in defending humans against cancer. This is exemplified by the increased risk of colorectal cancer of patients with germ-line mutations of the oxidative damage DNA glycosylase MUTYH. In contrast to germ-line mutations in DNA repair genes, which cause a strong deficiency in DNA repair activity in all cell types, the role of single nucleotide polymorphisms (SNPs) in sporadic cancer is unclear also because deficiencies in DNA repair, if any, are expected to be much milder. Further slowing down progress are the paucity of accurate and reproducible functional assays and poor epidemiological design of many studies. This review will focus on the most common and widely studied SNPs of oxidative DNA damage repair proteins trying to bridge the information available on biochemical and structural features of the repair proteins with the functional effects of these variants and their potential impact on the pathogenesis of disease.


Asunto(s)
Daño del ADN , Reparación del ADN/genética , Polimorfismo de Nucleótido Simple , Especies Reactivas de Oxígeno/metabolismo , Enzimas Reparadoras del ADN/genética , Humanos , Mutación , Relación Estructura-Actividad
13.
Mutat Res ; 718(1-2): 62-7, 2011 Jan 10.
Artículo en Inglés | MEDLINE | ID: mdl-20971211

RESUMEN

Several lines of evidence suggest an association between oxidative DNA-damage repair capacity and cancer risk. In particular, a DNA-glycosylase assay for removal of 8-oxoguanine (8-oxoG) in peripheral blood mononuclear cells (PBMC) has been successfully applied to identify populations with increased risk for lung cancer and squamous cell carcinomas of head and neck. In order to verify whether EBV-transformed lymphoblastoid cell lines (LCL) are a suitable surrogate for PBMC in specific DNA-repair phenotypic assays, a validation trial was conducted. PBMC from 20 healthy subjects were collected and an aliquot was transformed with EBV to obtain LCL. The ability of cell-free extracts from both cell types to incise a 3'-fluorescently labelled duplex oligonucleotide containing a single 8-oxoG (OGG assay) was evaluated. Since this activity is mediated predominantly by OGG1, the OGG1 gene expression was also measured. 8-oxoG DNA-glycosylase activity and OGG1 expression were significantly higher (p<0.0001) in LCL than in PBMC. However, while this assay was shown to be robust and reproducible when used on PBMC (intra-assay CV=8%), a high intra-culture variability was observed with LCL (intra-culture CV=16.8%). Neither differences on OGG1 gene expression nor the cell-cycle distribution seemed to account for this variability. Inter-individual variability of OGG activity in PBMC and LCL was not associated with OGG1 gene expression. We have therefore established a non-radioactive cleavage assay that can be easily applied to measure OGG activity in human PBMC. The use of LCL for DNA-repair genotype-phenotype correlation studies seems to be inappropriate, at least with cell-free based functional assays.


Asunto(s)
ADN Glicosilasas/genética , ADN Glicosilasas/metabolismo , Reparación del ADN , Guanina/análogos & derivados , Linfocitos/metabolismo , Línea Celular , Transformación Celular Viral , Criopreservación , Expresión Génica , Estudios de Asociación Genética , Guanina/metabolismo , Herpesvirus Humano 4 , Humanos , Técnicas In Vitro
14.
BMC Cancer ; 10: 129, 2010 Apr 07.
Artículo en Inglés | MEDLINE | ID: mdl-20374648

RESUMEN

BACKGROUND: Prostate stem cell antigen (PSCA) is a glycosylphosphatidylinositol (GPI) anchored protein expressed not only in prostate but also in pancreas and bladder cancer as shown by immunohistochemistry and mRNA analysis. It has been targeted by monoclonal antibodies in preclinical animal models and more recently in a clinical trial in prostate cancer patients. The biological role played in tumor growth is presently unknown. In this report we have characterized the contribution of PSCA expression to tumor growth. METHODS: A bladder cell line was engineered to express a doxycycline (dox) regulated shRNA against PSCA. To shed light on the PSCA biological role in tumor growth, microarray analysis was carried out as a function of PSCA expression. Expression of gene set of interest was further analyzed by qPCR RESULTS: Down regulation of the PSCA expression was associated with reduced cell proliferation in vitro and in vivo. Mice bearing subcutaneous tumors showed a reduced tumor growth upon treatment with dox, which effectively induced shRNA against PSCA as revealed by GFP expression. Pathway analysis of deregulated genes suggests a statistical significant association between PSCA downregulation and activation of genes downstream of the IFNalpha/beta receptor. CONCLUSIONS: These experiments established for the first time a correlation between the level of PSCA expression and tumor growth and suggest a role of PSCA in counteracting the natural immune response.


Asunto(s)
Glicoproteínas de Membrana/biosíntesis , Proteínas de Neoplasias/biosíntesis , Neoplasias de la Vejiga Urinaria/patología , Animales , Antígenos de Neoplasias , Procesos de Crecimiento Celular/fisiología , Línea Celular Tumoral , Células Clonales , Regulación hacia Abajo , Doxiciclina/farmacología , Femenino , Proteínas Ligadas a GPI , Humanos , Glicoproteínas de Membrana/genética , Glicoproteínas de Membrana/inmunología , Ratones , Ratones Desnudos , Proteínas de Neoplasias/genética , Proteínas de Neoplasias/inmunología , Reacción en Cadena de la Polimerasa , Interferencia de ARN , Transducción de Señal/inmunología , Transcripción Genética , Neoplasias de la Vejiga Urinaria/genética , Neoplasias de la Vejiga Urinaria/inmunología , Neoplasias de la Vejiga Urinaria/metabolismo
15.
Mutat Res ; 670(1-2): 79-87, 2009 Nov 02.
Artículo en Inglés | MEDLINE | ID: mdl-19635489

RESUMEN

DNA repair polymerase beta (Pol beta) gene variants are frequently associated with tumor tissues. In this study a search for Pol beta mutants and splice variants was conducted in matched normal and tumor gastric tissues and blood samples from healthy donors. No tumor associated mutations were found while a variety of alternative Pol beta splicing variants were detected with high frequency in all the specimens analysed. Quantitative PCR of the Pol beta variant lacking exon 2 (Ex2Delta) and the isoforms with exon 11 skipping allowed to clarify that these variants are not tumor- neither tissue-specific and their levels vary greatly among different individuals. The most frequent Ex2Delta variant was further characterized. We clearly demonstrated that this variant does not encode protein, as detected by both western blotting and immunofluorescence analysis of human AGS cells expressing HA-tagged Ex2Delta. The lack of translation was confirmed by comparing the DNA gap-filling capacity and alkylation sensitivity of wild type and Pol beta null murine fibroblasts expressing the human Ex2Delta variant. We showed that the Ex2Delta transcript is polyadenylated and its half-life is significantly longer than that of the wild type mRNA as inferred by treating AGS cells with actinomycin D. Moreover, we found that it localizes to polyribosomes suggesting a role as post-transcriptional regulator. This study identifies a new type of DNA repair variants that do not give rise to functional proteins but to non-coding RNAs that could either modulate target mRNAs or represent unproductive splicing events.


Asunto(s)
ADN Polimerasa beta/genética , Exones , Isoformas de Proteínas , ARN no Traducido , Neoplasias Gástricas/genética , Empalme Alternativo , Animales , Estudios de Casos y Controles , Línea Celular , Línea Celular Tumoral , Reparación del ADN , Humanos , Ratones , Isoformas de Proteínas/genética , Empalme del ARN , Estómago , Transfección
16.
Nucleic Acids Res ; 33(14): 4404-11, 2005.
Artículo en Inglés | MEDLINE | ID: mdl-16077026

RESUMEN

Base excision repair (BER) is the main pathway for repair of DNA damage in mammalian cells. This pathway leads to the formation of DNA repair intermediates which, if still unsolved, cause cell lethality and mutagenesis. To characterize mutations induced by BER intermediates in mammalian cells, an SV-40 derived shuttle vector was constructed carrying a site-specific lesion within the recognition sequence of a restriction endonuclease. The mutation spectra of abasic (AP) sites, 5'-deoxyribose-5-phosphate (5'dRp) and 3'-[2,3-didehydro-2,3-dideoxy-ribose] (3'ddR5p) single-strand breaks (ssb) in mammalian cells was analysed by RFLP/PCR and mutation frequency was estimated by quantitative PCR. Point mutations were the predominant events occurring at all BER intermediates. The AP site-induced mutation spectrum supports evidence for the 'A-rule' and is also consistent with the use of the 5' neighbouring base to instruct nucleotide incorporation (5'-rule). Preferential adenine insertion was also observed after in vivo replication of 5'dRp or 3'ddR5p ssb. We provide original evidence that not only the abasic site but also its derivatives 'faceless' BER intermediates are mutagenic, with a similar mutation frequency, in mammalian cells. Our findings support the hypothesis that unattended BER intermediates could be a constant threat for genome integrity as well as a spontaneous source of mutations.


Asunto(s)
Reparación del ADN , Desoxirribosa/análogos & derivados , Desoxirribosa/metabolismo , Mutagénesis , Ribosamonofosfatos/metabolismo , Animales , Células COS , Chlorocebus aethiops , Enzimas Reparadoras del ADN/metabolismo , Enzimas de Restricción del ADN/metabolismo , Vectores Genéticos , Mutación Puntual , Reacción en Cadena de la Polimerasa , Virus 40 de los Simios/genética
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