RESUMEN

The occurrence of damage on bacterial DNA (mediated by antibiotics, for example) is intimately associated with the activation of the SOS system. This pathway is related to the development of mutations that might result in the acquisition and spread of resistance and virulence factors. The inhibition of the SOS response has been highlighted as an emerging resource, in order to reduce the emergence of drug resistance and tolerance. Herein, we evaluated the ability of betulinic acid (BA), a plant-derived triterpenoid, to reduce the activation of the SOS response and its associated phenotypic alterations, induced by ciprofloxacin in Staphylococcus aureus. BA did not show antimicrobial activity against S. aureus (MIC > 5000 µg/mL), however, it (at 100 and 200 µg/mL) was able to reduce the expression of recA induced by ciprofloxacin. This effect was accompanied by an enhancement of the ciprofloxacin antimicrobial action and reduction of S. aureus cell volume (as seen by flow cytometry and fluorescence microscopy). BA could also increase the hyperpolarization of the S. aureus membrane, related to the ciprofloxacin action. Furthermore, BA inhibited the progress of tolerance and the mutagenesis induced by this drug. Taken together, these findings indicate that the betulinic acid is a promising lead molecule in the development helper drugs. These compounds may be able to reduce the S. aureus mutagenicity associated with antibiotic therapies.

Asunto(s)

Farmacorresistencia Bacteriana/efectos de los fármacos , Rec A Recombinasas/genética , Staphylococcus aureus/genética , Triterpenos/farmacología , Ciprofloxacina/efectos adversos , Ciprofloxacina/farmacología , ADN Bacteriano/efectos de los fármacos , Regulación Bacteriana de la Expresión Génica/efectos de los fármacos , Humanos , Mutagénesis/efectos de los fármacos , Mutagénesis/genética , Triterpenos Pentacíclicos , Respuesta SOS en Genética/efectos de los fármacos , Respuesta SOS en Genética/genética , Staphylococcus aureus/efectos de los fármacos , Staphylococcus aureus/patogenicidad , Factores de Virulencia/genética , Ácido Betulínico

RESUMEN

The oxidative stress response of the highly resistant actinomycete Dietzia cinnamea P4 after treatment with hydrogen peroxide (H2O2) was assessed in order to depict the possible mechanisms underlying its intrinsic high resistance to DNA damaging agents. We used transcriptional profiling to monitor the magnitude and kinetics of changes in the mRNA levels after exposure to different concentrations of H2O2 at 10 min and 1 h following the addition of the stressor. Catalase and superoxide dismutase genes were induced in different ways, according to the condition applied. Moreover, alkyl hydroperoxide reductase ahpCF, thiol peroxidase, thioredoxin and glutathione genes were upregulated in the presence of H2O2. Expression of peroxidase genes was not detected during the experiment. Overall results point to an actinomycete strain endowed with a set of enzymatic defenses against oxidative stress and with the main genes belonging to a functional SOS system (lexA, recA, uvrD), including suppression of lexA repressor, concomitantly to recA and uvrD gene upregulation upon H2O2 challenge.

Asunto(s)

Actinomycetales/efectos de los fármacos , Actinomycetales/metabolismo , Peróxido de Hidrógeno/efectos adversos , Estrés Oxidativo , Respuesta SOS en Genética/fisiología , Actinomycetales/enzimología , Actinomycetales/genética , Proteínas Bacterianas/genética , Catalasa/clasificación , Catalasa/genética , Daño del ADN/efectos de los fármacos , ADN Helicasas/genética , Regulación Bacteriana de la Expresión Génica/efectos de los fármacos , Genes Bacterianos , Glutatión/genética , Cinética , Peroxidasas/genética , Peroxirredoxinas/genética , Filogenia , ARN Mensajero/metabolismo , Rec A Recombinasas/genética , Respuesta SOS en Genética/genética , Análisis de Secuencia , Serina Endopeptidasas/genética , Superóxido Dismutasa/genética , Tiorredoxinas/genética , Factores de Tiempo , Regulación hacia Arriba/efectos de los fármacos

RESUMEN

DNA damage-induced mutagenesis is a process governed by the SOS system that requires the activity of specialized DNA polymerases. These polymerases, which are devoid of proof-reading activity, serve to increase the probability of survival under stressful conditions in exchange for an error-prone DNA synthesis. As an opportunistic pathogen of humans, Pseudomonas aeruginosa employs adaptive responses that originally evolved for survival in many diverse and often stressful environmental conditions, where the action of error-prone DNA polymerases may be crucial. In this study, we have investigated the role of the polymerases ImuB and ImuC in P. aeruginosa DNA-damage induced mutagenesis. UV irradiation of imuB- and imuC-deletion mutants showed that both genes contribute to UV-induced mutagenesis in this bacterium. Furthermore, we confirmed that UV treatment significantly increase the expression levels of the imuB and imuC genes and that they are co-transcribed as a single transcriptional unit under the control of LexA as part of the SOS regulon in P. aeruginosa. Environ. Mol. Mutagen. 2019. © 2019 Wiley Periodicals, Inc.

Asunto(s)

ADN Bacteriano/genética , Mutagénesis/genética , Pseudomonas aeruginosa/genética , Respuesta SOS en Genética/genética , Rayos Ultravioleta/efectos adversos , Daño del ADN/genética , ADN Polimerasa Dirigida por ADN/genética , Regulón/genética

RESUMEN

Translesion DNA polymerases (Pol) function in the bypass of template lesions to relieve stalled replication forks but also display potentially deleterious mutagenic phenotypes that contribute to antibiotic resistance in bacteria and lead to human disease. Effective activity of these enzymes requires association with ring-shaped processivity factors, which dictate their access to sites of DNA synthesis. Here, we show for the first time that the mismatch repair protein MutS plays a role in regulating access of the conserved Y-family Pol IV to replication sites. Our biochemical data reveals that MutS inhibits the interaction of Pol IV with the ß clamp processivity factor by competing for binding to the ring. Moreover, the MutS-ß clamp association is critical for controlling Pol IV mutagenic replication under normal growth conditions. Thus, our findings reveal important insights into a non-canonical function of MutS in the regulation of a replication activity.

Asunto(s)

ADN Polimerasa beta/metabolismo , Replicación del ADN , Proteína MutS de Unión a los Apareamientos Incorrectos del ADN/metabolismo , Pseudomonas aeruginosa/metabolismo , Biocatálisis , ADN/biosíntesis , ADN/química , ADN Polimerasa III/metabolismo , Etilnitrosourea , Mutagénesis/genética , Péptidos/metabolismo , Unión Proteica , Pseudomonas aeruginosa/crecimiento & desarrollo , Respuesta SOS en Genética/genética , Especificidad por Sustrato

RESUMEN

The SOS response is a universal bacterial regulon involved in the cellular response to DNA damage and other forms of stress. In Caulobacter crescentus, previous work has identified a plethora of genes that are part of the SOS regulon, but the biological roles of several of them remain to be determined. In this study, we report that two genes, hereafter named mmcA and mmcB, are involved in the defense against DNA damage caused by mitomycin C (MMC), but not against lesions induced by other common DNA damaging agents, such as UVC light, methyl methanesulfonate (MMS) and hydrogen peroxide. mmcA is a conserved gene that encodes a member of the glyoxalases/dioxygenases protein family, and acts independently of known DNA repair pathways. On the other hand, epistasis analysis showed that mmcB acts in the same pathway as imuC (dnaE2), and is required specifically for MMC-induced mutagenesis, but not for that induced by UV light, suggesting a role for MmcB in translesion synthesis-dependent repair of MMC damage. We show that the lack of MMC-induced mutability in the mmcB strain is not caused by lack of proper SOS induction of the imuABC operon, involved in translesion synthesis (TLS) in C. crescentus. Based on this data and on structural analysis of a close homolog, we propose that MmcB is an endonuclease which creates substrates for ImuABC-mediated TLS patches.

Asunto(s)

Proteínas Bacterianas/genética , Caulobacter crescentus/genética , Genes Bacterianos , Mitomicina/farmacología , Respuesta SOS en Genética/genética , Secuencia de Aminoácidos , Proteínas Bacterianas/química , Proteínas Bacterianas/metabolismo , Secuencia de Bases , Dominio Catalítico , Caulobacter crescentus/efectos de los fármacos , Caulobacter crescentus/crecimiento & desarrollo , Caulobacter crescentus/efectos de la radiación , Secuencia Conservada , Daño del ADN , Farmacorresistencia Bacteriana/efectos de los fármacos , Farmacorresistencia Bacteriana/genética , Epistasis Genética/efectos de los fármacos , Epistasis Genética/efectos de la radiación , Eliminación de Gen , Viabilidad Microbiana/efectos de los fármacos , Viabilidad Microbiana/efectos de la radiación , Modelos Moleculares , Datos de Secuencia Molecular , Mutagénesis/efectos de la radiación , Mutación/genética , Tasa de Mutación , Fenotipo , Regiones Promotoras Genéticas/genética , Respuesta SOS en Genética/efectos de los fármacos , Respuesta SOS en Genética/efectos de la radiación , Rayos Ultravioleta

RESUMEN

Cisplatin is currently used in tumor chemotherapy to induce the death of malignant cells through blockage of DNA replication. It is a commonly used chemotherapeutic agent binding mono- or bifunctionally to guanines in DNA. Escherichia coli K12 mutant strains deficient in nucleotide excision repair (NER) were submitted to increasing concentrations of cisplatin, and the results revealed that uvrA and uvrB mutants are sensitive to this agent, while uvrC and cho mutants remain as the wild type strain. The time required for both gene expression turn-off and return to normal weight DNA in wild-type E. coli was not accomplished even after 4 h post-treatment with cisplatin, while the same process takes place within 1.5 h after ultraviolet radiation (UV). Besides, a heavily damaging action of cisplatin can be seen not only by persistent nicks on genomic DNA, but also by NER gene expression exceeding manifold that seen after equivalent lethal doses of UV. Moreover, cisplatin caused an increase in uvrB gene expression from its putative upstream promoter P3 in an SOS-independent manner.

Asunto(s)

Antineoplásicos/toxicidad , Cisplatino/toxicidad , Roturas del ADN de Cadena Simple , Escherichia coli/genética , Respuesta SOS en Genética/genética , Adenosina Trifosfatasas/genética , Adenosina Trifosfatasas/metabolismo , ADN Helicasas/genética , ADN Helicasas/metabolismo , ADN Bacteriano/metabolismo , Proteínas de Unión al ADN/genética , Proteínas de Unión al ADN/metabolismo , Endodesoxirribonucleasas/genética , Endodesoxirribonucleasas/metabolismo , Escherichia coli/metabolismo , Proteínas de Escherichia coli/genética , Proteínas de Escherichia coli/metabolismo , Regulación Bacteriana de la Expresión Génica , Mutación , Regiones Promotoras Genéticas , Transcripción Genética , Rayos Ultravioleta , Regulación hacia Arriba

RESUMEN

Although the use of medicinal plants or natural products has increased in recent decades all over the world, little information is available on their potential risk to health. Annona crassiflora Mart., a plant commonly known as araticum in Brazil, has been widely used in folk medicine for a long time since its seeds and leaves are often utilised in the treatment of cancer, snake bites, and venereal diseases, its fruits are consumed as tonic and astringent, and its bark powder has anti-fungal and anti-rheumatic properties. To evaluate the genotoxic and mutagenic properties induced by the ethanolic extract of araticum leaves, we performed the prophage λ induction (Inductest) and bacterial mutagenicity assays. We used Escherichia coli WP2s(λ) and RJF013 strains in the lysogenic induction test, whereas the mutagenic studies were carried out using Salmonella typhimurium histidine auxotroph strains TA97a, TA98, TA100, and TA102. Each experiment was performed three times in duplicate and included positive and negative controls. No statistically significant (p > 0.05) positive results were obtained for any of the strains tested, which suggests that the ethanolic extract of araticum leaves did not exhibit direct mechanisms of genotoxicity or mutagenicity that could be detected by the tests used in the present work.

Asunto(s)

Annona/química , Escherichia coli/efectos de los fármacos , Pruebas de Mutagenicidad/métodos , Extractos Vegetales/toxicidad , Salmonella typhimurium/efectos de los fármacos , Daño del ADN/efectos de los fármacos , Daño del ADN/genética , Escherichia coli/genética , Profagos/efectos de los fármacos , Profagos/genética , Respuesta SOS en Genética/efectos de los fármacos , Respuesta SOS en Genética/genética , Salmonella typhimurium/genética

RESUMEN

Although the use of medicinal plants or natural products has increased in recent decades all over the world, little information is available on their potential risk to health. Annona crassiflora Mart., a plant commonly known as araticum in Brazil, has been widely used in folk medicine for a long time since its seeds and leaves are often utilised in the treatment of cancer, snake bites, and venereal diseases, its fruits are consumed as tonic and astringent, and its bark powder has anti-fungal and anti-rheumatic properties. To evaluate the genotoxic and mutagenic properties induced by the ethanolic extract of araticum leaves, we performed the prophage λ induction (Inductest) and bacterial mutagenicity assays. We used Escherichia coli WP2s(λ) and RJF013 strains in the lysogenic induction test, whereas the mutagenic studies were carried out using Salmonella typhimurium histidine auxotroph strains TA97a, TA98, TA100, and TA102. Each experiment was performed three times in duplicate and included positive and negative controls. No statistically significant (p > 0.05) positive results were obtained for any of the strains tested, which suggests that the ethanolic extract of araticum leaves did not exhibit direct mechanisms of genotoxicity or mutagenicity that could be detected by the tests used in the present work.

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Embora o uso de plantas medicinais ou de produtos naturais venha aumentando nas últimas décadas no mundo todo, existem poucas informações acerca de seu risco potencial para a saúde. Annona crassiflora Mart., uma planta comumente conhecida como araticum no Brasil, tem tido amplo uso em medicina popular há muito tempo, uma vez que suas sementes e folhas são frequentemente empregadas no tratamento de câncer, picadas de cobras e doenças venéreas, seus frutos são consumidos como tônico e adstringente, e o pó da casca de seu tronco apresenta propriedades antifúngicas e antirreumáticas. Para avaliar as propriedades genotóxica e mutagênica induzidas pelo extrato etanólico das folhas de araticum, utilizaram-se os testes de indução do profago λ (Induteste) e de mutagenicidade bacteriana. Foram empregadas as linhagens WP2s(λ) e RJF013 de Escherichia coli no teste de indução lisogênica, enquanto os estudos sobre mutagenicidade foram conduzidos utilizando as linhagens auxotróficas para histidina TA97a, TA98, TA100 e TA102 de Salmonella typhimurium. Cada experimento foi executado três vezes em duplicata, incluindo controles positivo e negativo. Não foram obtidos resultados positivos estatisticamente significativos (p > 0,05) para quaisquer das linhagens testadas, o que sugere que o extrato etanólico das folhas de araticum não apresentou mecanismos diretos de genotoxicidade ou mutagenicidade que pudessem ser detectados pelos testes usados no presente estudo.

Asunto(s)

Annona/química , Escherichia coli/efectos de los fármacos , Pruebas de Mutagenicidad/métodos , Extractos Vegetales/toxicidad , Salmonella typhimurium/efectos de los fármacos , Daño del ADN/efectos de los fármacos , Daño del ADN/genética , Escherichia coli/genética , Profagos/efectos de los fármacos , Profagos/genética , Respuesta SOS en Genética/efectos de los fármacos , Respuesta SOS en Genética/genética , Salmonella typhimurium/genética

RESUMEN

Regardless of official recommendations, the inappropriate use of homemade hair creams has became a popular practice in Brazil and high formaldehyde content in the 'progressive straightening' creams has been reported. In the present work, three of these creams were analyzed by spectrophotometric, chromatographic and genotoxic assays in order to evaluate mutagenic risks associated with the uncontrolled addition of formaldehyde at contents higher than those allowed by regulation. The ultraviolet and Fourier-transformed infrared absorption spectra showed characteristic signals that can be assigned to formaldehyde, although with different relative intensities, revealing distinct compositions. Using high-performance liquid chromatography 1.6-10.5% w/v formaldehyde was quantified. Antibacterial activity was detected in all creams. At 0.10 microg per plate, one of them showed positive mutagenicity induction (P < 0.05) in the Salmonella/microsome assay using the TA100 strain. The measurement of beta-galactosidase induction in the SOS chromotest by this cream, at dosages of 10-100 microg per assay, was positive (P < 0.05) in Escherichia coli PQ37 and OG100 strains. Our data show a more intense genotoxic response than those reported before for formaldehyde, suggesting that this compound may be acting synergistically with any unknown components in the creams or perhaps these unspecified components by themselves might have significant genotoxic potential. We call attention to the popular use of homemade formulations of cosmetics, such as hair straightening creams, because they can contain mutagens that could increase the incidence of neoplasia in those people who use them.

Asunto(s)

Cosméticos , Formaldehído , Preparaciones para el Cabello , Pruebas de Mutagenicidad/métodos , Mutágenos/toxicidad , Cromatografía Líquida de Alta Presión , Cosméticos/química , Cosméticos/toxicidad , Relación Dosis-Respuesta a Droga , Formaldehído/análisis , Formaldehído/toxicidad , Preparaciones para el Cabello/química , Preparaciones para el Cabello/toxicidad , Humanos , Viabilidad Microbiana/efectos de los fármacos , Mutágenos/química , Mutación/genética , Respuesta SOS en Genética/efectos de los fármacos , Respuesta SOS en Genética/genética , Salmonella typhimurium/efectos de los fármacos , Salmonella typhimurium/genética , Salmonella typhimurium/crecimiento & desarrollo

RESUMEN

The SOS regulon is a paradigm of bacterial responses to DNA damage. A wide variety of bacterial species possess homologs of lexA and recA, the central players in the regulation of the SOS circuit. Nevertheless, the genes actually regulated by the SOS have been determined only experimentally in a few bacterial species. In this work, we describe 37 genes regulated in a LexA-dependent manner in the alphaproteobacterium Caulobacter crescentus. In agreement with previous results, we have found that the direct repeat GTTCN7GTTC is the SOS operator of C. crescentus, which was confirmed by site-directed mutagenesis studies of the imuA promoter. Several potential promoter regions containing the SOS operator were identified in the genome, and the expression of the corresponding genes was analyzed for both the wild type and the lexA strain, demonstrating that the vast majority of these genes are indeed SOS regulated. Interestingly, many of these genes encode proteins with unknown functions, revealing the potential of this approach for the discovery of novel genes involved in cellular responses to DNA damage in prokaryotes, and illustrating the diversity of SOS-regulated genes among different bacterial species.

Asunto(s)

Proteínas Bacterianas/genética , Caulobacter crescentus/genética , Regulón/genética , Respuesta SOS en Genética/genética , Proteínas Bacterianas/metabolismo , Secuencia de Bases , Sitios de Unión/genética , Caulobacter crescentus/metabolismo , Daño del ADN , Perfilación de la Expresión Génica , Regulación Bacteriana de la Expresión Génica/efectos de la radiación , Datos de Secuencia Molecular , Mutagénesis Sitio-Dirigida , Técnicas de Amplificación de Ácido Nucleico , Regiones Operadoras Genéticas/genética , Fenotipo , Regiones Promotoras Genéticas/genética , Reacción en Cadena de la Polimerasa de Transcriptasa Inversa , Serina Endopeptidasas/genética , Serina Endopeptidasas/metabolismo , Sitio de Iniciación de la Transcripción , Rayos Ultravioleta

RESUMEN

A range of biologically active secondary metabolites with pharmacological application has been reported to occur in marine sponges. The present study was undertaken to provide a set of data on the safety of a hydro-alcoholic extract (ALE) and an aqueous fraction (AQE) from Aplysina fulva Pallas, 1766 (Aplysinidae, Verongida, Porifera). Salmonella typhimurium strains TA97, TA98, TA100 and TA102, Escherichia coli strains PQ65, OG40, OG100, PQ35 and PQ37 and Balb/c 3T3 mouse fibroblasts were used to detect induction of DNA lesions by ALE and AQE. Assays used for these analyses were a bacterial (reverse) mutation assay (Ames test), the SOS-chromotest and the comet assay. Both extracts presented identical infrared 2-oxazolidone spectra. ALE treatment induced a higher frequency of type-4 comets, indicative of increasing DNA migration, in the alkaline comet assay. ALE also induced a weak genotoxic effect, as expressed by the induction factor (IF) values in the test with E. coli strain PQ35 (IF=1.5) and by cytotoxic effects in strains PQ35, PQ65 and PQ37. Positive SOS induction (IF=1.7) was detected in strain PQ37 treated with diluted AQE. No genotoxic effects were observed in strains PQ35, PQ65, OG40 and OG 100 after treatment with AQE dilutions. Using the bacterial (reverse) mutation test and survival assays with or without S9 mix, after 60min of pre-incubation, we observed for strain TA97 treated with ALE a weak mutagenic response (MI=2.2), while cytotoxic effects were seen for strains TA98, TA100 and TA102. AQE did not show mutagenic activity in any of the strains tested, but a weak cytotoxic effect was noted in strain TA102. Our data suggest that both ALE and AQE from A. fulva induce DNA breaks leading to cytotoxicity and mutagenicity under the conditions used.

Asunto(s)

Mutágenos/toxicidad , Poríferos/química , Respuesta SOS en Genética/efectos de los fármacos , Células 3T3 , Animales , Ensayo Cometa/métodos , Roturas del ADN , ADN Bacteriano/efectos de los fármacos , ADN Bacteriano/genética , Escherichia coli/efectos de los fármacos , Escherichia coli/genética , Ratones , Ratones Endogámicos BALB C , Viabilidad Microbiana/efectos de los fármacos , Viabilidad Microbiana/genética , Pruebas de Mutagenicidad/métodos , Ratas , Ratas Sprague-Dawley , Respuesta SOS en Genética/genética , Salmonella typhimurium/efectos de los fármacos , Salmonella typhimurium/genética

RESUMEN

In the present work, we evaluated (p < 0.05) the participation of base excision repair (BER) and nucleotide excision repair (NER) mechanisms in repairing DNA lesions induced by N-nitrosodiethylamine (NDEA) at 1.5 ng/mL-36.5 microg/mL, through cell survival, in different single and double Escherichia coli DNA repair mutants (uvrA, uvrB, uvrC, fpg, nth, xthA, fpg/nth, uvrA/fpg, fpg/xthA, mutY, and fpg/mutY), using pre-incubation periods of 90 min. Mutant strains BH20 (fpg) and AB1886 (uvrA) showed microsomal enzyme (S9 mix) independent NDEA cytotoxicity. Cytotoxicity was also detected at lowest NDEA concentrations, in the presence of S9 mix, with strains BH980 (mutY) and BH990 (fpg/mutY). NDEA cytotoxicity, without S9 mix, was detected for mutant strains AB1884 (uvrC) and AB1885 (uvrB). Through SOS chromotest with 90 min of pre-incubation for uvrA and nth strains, only NER was shown to be required for repairing NDEA-induced lesions with or without metabolic activation. PQ37 and PQ66 strains, both uvrA mutants, showed different levels of NDEA sensitivity. The findings suggest that, under the used conditions, and at low concentrations, NDEA-induced lesions require both repair pathways.

Asunto(s)

Alquilantes/toxicidad , Reparación del ADN/efectos de los fármacos , ADN Bacteriano/efectos de los fármacos , Dietilnitrosamina/toxicidad , Respuesta SOS en Genética/efectos de los fármacos , Animales , Recuento de Colonia Microbiana , Daño del ADN/genética , Reparación del ADN/genética , Relación Dosis-Respuesta a Droga , Escherichia coli/efectos de los fármacos , Escherichia coli/genética , Microsomas Hepáticos/efectos de los fármacos , Microsomas Hepáticos/enzimología , Mutación , Ratas , Ratas Sprague-Dawley , Respuesta SOS en Genética/genética

RESUMEN

Chromobacterium violaceum is a Gram-negative beta-proteobacterium that inhabits a variety of ecosystems in tropical and subtropical regions, including the water and banks of the Negro River in the Brazilian Amazon. This bacterium has been the subject of extensive study over the last three decades, due to its biotechnological properties, including the characteristic violacein pigment, which has antimicrobial and anti-tumoral activities. C. violaceum promotes the solubilization of gold in a mercury-free process, and has been used in the synthesis of homopolyesters suitable for the production of biodegradable polymers. The complete genome sequence of this organism has been completed by the Brazilian National Genome Project Consortium. The aim of our group was to study the DNA repair genes in this organism, due to their importance in the maintenance of genomic integrity. We identified DNA repair genes involved in different pathways in C. violaceum through a similarity search against known sequences deposited in databases. The phylogenetic analyses were done using programs of the PHILYP package. This analysis revealed various metabolic pathways, including photoreactivation, base excision repair, nucleotide excision repair, mismatch repair, recombinational repair, and the SOS system. The similarity between the C. violaceum sequences and those of Neisserie miningitidis and Ralstonia solanacearum was greater than that between the C. violaceum and Escherichia coli sequences. The peculiarities found in the C. violaceum genome were the absence of LexA, some horizontal transfer events and a large number of repair genes involved with alkyl and oxidative DNA damage.

Asunto(s)

Proteínas Bacterianas/genética , Chromobacterium/genética , Reparación del ADN/genética , Homología de Secuencia , Disparidad de Par Base/genética , Bases de Datos Genéticas , Filogenia , Recombinación Genética , Respuesta SOS en Genética/genética

RESUMEN

Chromobacterium violaceum is a Gram-negative beta-proteobacterium that inhabits a variety of ecosystems in tropical and subtropical regions, including the water and banks of the Negro River in the Brazilian Amazon. This bacterium has been the subject of extensive study over the last three decades, due to its biotechnological properties, including the characteristic violacein pigment, which has antimicrobial and anti-tumoral activities. C. violaceum promotes the solubilization of gold in a mercury-free process, and has been used in the synthesis of homopolyesters suitable for the production of biodegradable polymers. The complete genome sequence of this organism has been completed by the Brazilian National Genome Project Consortium. The aim of our group was to study the DNA repair genes in this organism, due to their importance in the maintenance of genomic integrity. We identified DNA repair genes involved in different pathways in C. violaceum through a similarity search against known sequences deposited in databases. The phylogenetic analyses were done using programs of the PHILYP package. This analysis revealed various metabolic pathways, including photoreactivation, base excision repair, nucleotide excision repair, mismatch repair, recombinational repair, and the SOS system. The similarity between the C. violaceum sequences and those of Neisserie miningitidis and Ralstonia solanacearum was greater than that between the C. violaceum and Escherichia coli sequences. The peculiarities found in the C. violaceum genome were the absence of LexA, some horizontal transfer events and a large number of repair genes involved with alkyl and oxidative DNA damage

Asunto(s)

Chromobacterium/genética , Proteínas Bacterianas/genética , Reparación del ADN/genética , Homología de Secuencia , Bases de Datos Genéticas , Filogenia , Disparidad de Par Base/genética , Recombinación Genética , Respuesta SOS en Genética/genética

RESUMEN

The recA and the recX genes of Herbaspirillum seropedicae were sequenced. The recX is located 359 bp downstream from recA. Sequence analysis indicated the presence of a putative operator site overlapping a probable sigma70-dependent promoter upstream of recA and a transcription terminator downstream from recX, with no apparent promoter sequence in the intergenic region. Transcriptional analysis using lacZ promoter fusions indicated that recA expression increased three- to fourfold in the presence of methyl methanesulfonate (MMS). The roles of recA and recX genes in the SOS response were determined from studies of chromosomal mutants. The recA mutant showed the highest sensitivity to MMS and UV, and the recX mutant had an intermediate sensitivity, compared with the wild type (SMR1), confirming the essential role of the RecA protein in cell viability in the presence of mutagenic agents and also indicating a role for RecX in the SOS response.

Asunto(s)

Proteínas Bacterianas/fisiología , Betaproteobacteria/genética , Respuesta SOS en Genética/fisiología , Betaproteobacteria/clasificación , Betaproteobacteria/efectos de los fármacos , Betaproteobacteria/efectos de la radiación , Recuento de Colonia Microbiana , Metilmetanosulfonato/farmacología , Modelos Genéticos , Fijación del Nitrógeno , Respuesta SOS en Genética/genética , Rayos Ultravioleta

RESUMEN

Phyllanthus orbicularis HBK is an endemic Cuban plant whose aqueous extract has been proposed as an effective drug for the treatment of viral diseases. In addition, antimutagenic properties of this extract have also been reported. In the present study, the genotoxicity of this plant extract was assessed using different in vitro and in vivo assays. Results from SOS gene induction, gene reversion and conversion, and SMART assays clearly show that P. orbicularis aqueous extract does not induce either primary DNA damage or mutation. Additionally, no statistically significant difference was found in the percentage of chromosomal aberrations in Chinese hamster ovarian (CHO) cells treated with the plant extract. On the contrary, micronuclei and abnormal anaphase were induced by this extract in CHO cells. This genotoxic effect was related to a high cytotoxicity. Single spots were detected in the SMART assay. These results point to a possible aneugenic effect of the P. orbicularis aqueous extract at cytotoxic doses which are much higher than those seen by their antiviral and antimutagenic activities.

Asunto(s)

Mutación/efectos de los fármacos , Phyllanthus/toxicidad , Extractos Vegetales/toxicidad , Anafase/efectos de los fármacos , Animales , Células CHO , Aberraciones Cromosómicas/efectos de los fármacos , Cricetinae , ADN/análisis , Daño del ADN/efectos de los fármacos , Daño del ADN/genética , Relación Dosis-Respuesta a Droga , Drosophila melanogaster/efectos de los fármacos , Drosophila melanogaster/genética , Escherichia coli/efectos de los fármacos , Escherichia coli/genética , Femenino , Pruebas de Mutagenicidad , Mutación/genética , Respuesta SOS en Genética/efectos de los fármacos , Respuesta SOS en Genética/genética , Saccharomyces cerevisiae/efectos de los fármacos , Saccharomyces cerevisiae/genética , Salmonella typhimurium/efectos de los fármacos , Salmonella typhimurium/genética

RESUMEN

Despite 2,9-dimethyl 1,10-phenanthroline (NC) has been extensively used as a potential inhibitor of damage due to oxidative stress in biological systems, the incubation of E. coli cultures with the copper ion chelator NC prior to the challenge with hydrogen peroxide caused a lethal synergistic effect. The SOS response seems to be involved in the repair of the synergistic lesions through the recombination pathway. Furthermore, there is evidence for the UvrABC excinuclease participation in the repair of the synergistic lesions, and the base excision repair may also be required for bacterial survival to the synergistic effect mainly at high concentrations of H2O2, being the action of Fpg protein an important event. Incubation of lexA (Ind-) cultures with iron (II) ion chelator 2,2'-dipyridyl simultaneously with NC prevented the lethal synergistic effect. This result suggests an important role of the Fenton reaction on the phenomenon. NC treatment was able to increase the number of DNA strand breaks (DNAsb) induced by 10 mM of H2O2 in lexA (Ind-) strain and the simultaneous treatment with 2,2'-dipyridyl was able to block this effect.

Asunto(s)

Daño del ADN/genética , Sinergismo Farmacológico , Escherichia coli/efectos de los fármacos , Peróxido de Hidrógeno/metabolismo , 2,2'-Dipiridil/farmacología , Proteínas Bacterianas/genética , Centrifugación por Gradiente de Densidad , Quelantes/metabolismo , ADN/análisis , Reparación del ADN/genética , Escherichia coli/genética , Peróxido de Hidrógeno/farmacología , Peróxido de Hidrógeno/toxicidad , Hierro/farmacología , Mutación/genética , Fenantrolinas/metabolismo , Fenantrolinas/toxicidad , Recombinación Genética/genética , Respuesta SOS en Genética/genética

RESUMEN

The effect of sodium diethyldithiocarbamate (DDC) and S-2-aminoethyl-isothiouronicadenosin-5-triphosphate (adeturon) in the induction of Escherichia coli SOS response promoted by gamma-irradiation was studied by measuring the induction of sulA gene and the induction of lambda prophage. Furthermore, as a way of measure the exonuclease activity in gamma-irradiated cells in the presence or absence of both compounds, the DNA degradation was determined. Adeturon did not affected DNA degradation, but inhibited the induction of the SOS functions studied. On the contrary, DDC inhibited DNA degradation as well as the induction of the sulA gene, but enhanced lambda induction in E. coli lysogenic strains. These results indicate that both compounds diminish the DNA damage produced by gamma-irradiation and also suggest that the mechanisms of radioprotection must be different. Thus, radioprotection mediated by DDC should involve free hydroxyl radical scavenging and a minor activity of exonuclease. The enhancement of phage induction in E. coli cells that DDC produces could be attributed to its quelant effect and this would not be not probably directly related to radioprotection. Adeturon, as thiols, may serve also as scavenging agent of free hydroxyl radicals, diminishing indirectly the DNA damage level. In addition, adeturon must interact with DNA in the same form that other aminothiol compounds do it. This interaction, mediated by amino groups of adeturon, may serve to concentrate these compounds near of the DNA damage site, increasing the potential for the thiol portion of the molecule to donate hydrogen, decreasing the damage level on DNA molecule. However, adeturon do not modify the exonuclease activity. Some topic about the possible clinical application of both compounds are discussed.

Asunto(s)

Adenosina Trifosfato/análogos & derivados , Ditiocarba/farmacología , Proteínas de Escherichia coli , Escherichia coli/efectos de la radiación , Rayos gamma/efectos adversos , Protectores contra Radiación/farmacología , beta-Aminoetil Isotiourea/análogos & derivados , Adenosina Trifosfato/farmacología , Proteínas Bacterianas/efectos de los fármacos , Proteínas Bacterianas/genética , Proteínas Bacterianas/efectos de la radiación , Bacteriófago lambda/genética , Bacteriófago lambda/efectos de la radiación , Daño del ADN/efectos de la radiación , ADN Bacteriano/efectos de los fármacos , ADN Bacteriano/metabolismo , ADN Bacteriano/efectos de la radiación , Escherichia coli/efectos de los fármacos , Escherichia coli/genética , Respuesta SOS en Genética/efectos de los fármacos , Respuesta SOS en Genética/genética , Respuesta SOS en Genética/efectos de la radiación , beta-Aminoetil Isotiourea/farmacología

RESUMEN

The presence of the phi 105cts23 mutant prophage in Bacillus subtilis induces a series of pleiotropic effects that could be ascribed to an anti-SOS activity. In order to circumvent the phage function responsible for this phenomenon, the cts23 mutant repressor was cloned and sequenced. The isolated repressor reduced the survival capacity of the host cells after mitomycin C or nalidixic acid treatments and lowered the spontaneous reversion frequency. When SOS induction kinetics were studied, low or null induction of the damage-inducible din22::LacZ fusion was observed. In contrast, the presence of the wild-type prophage amplified the SOS response. Sequencing of the mutant repressor revealed that the cts23 mutation is a T-->C transition affecting the 5' closest codon to one of the two reported DNA binding domains.

Asunto(s)

Fagos de Bacillus/genética , Bacillus subtilis/genética , Bacillus subtilis/virología , Genes Virales , Proteínas Represoras/genética , Respuesta SOS en Genética/genética , Fusión Artificial Génica , Daño del ADN , Reparación del ADN , Regulación Bacteriana de la Expresión Génica , Mutación , Plásmidos , Temperatura , Proteínas Virales/genética , beta-Galactosidasa/metabolismo

RESUMEN

The nucleotide sequence upstream of the Escherichia coli yebG gene presents features similar to those found in SOS system regulatory sites (putative SOS box, -10 and -35 promoter boxes and a ribosome binding site). Operon fusion assays demonstrate now that this region controls transcription in a recA-, lexA-dependent way and that the reporter gene expression is inducible by DNA damage consequent to mitomycin C treatment. Increased expression does not result from an increase in plasmid copy number. These results indicate that yebG is a novel SOS regulon gene. The yebG product is predicted to be a 96 amino acid residue, 10.7 kDa protein whose function is not yet known. Unlike other SOS genes, the construct carrying the yebG regulatory region is not stationary phase inducible.

Asunto(s)

Daño del ADN/genética , Escherichia coli/genética , Genes Bacterianos , Secuencia de Bases , Secuencia de Consenso , ADN Bacteriano/genética , Escherichia coli/crecimiento & desarrollo , Escherichia coli/metabolismo , Datos de Secuencia Molecular , Operón , Regulón , Respuesta SOS en Genética/genética