RESUMO
Topoisomerase II (Top2) is an essential enzyme that decatenates DNA via a transient Top2-DNA covalent intermediate. This intermediate can be stabilized by a class of drugs termed Top2 poisons, resulting in massive DNA damage. Thus, Top2 activity is a double-edged sword that needs to be carefully controlled to maintain genome stability. We show that Uls1, an adenosine triphosphate (ATP)-dependent chromatin remodelling (Snf2) enzyme, can alter Top2 chromatin binding and prevent Top2 poisoning in yeast. Deletion mutants of ULS1 are hypersensitive to the Top2 poison acriflavine (ACF), activating the DNA damage checkpoint. We map Uls1's Top2 interaction domain and show that this, together with its ATPase activity, is essential for Uls1 function. By performing ChIP-seq, we show that ACF leads to a general increase in Top2 binding across the genome. We map Uls1 binding sites and identify tRNA genes as key regions where Uls1 associates after ACF treatment. Importantly, the presence of Uls1 at these sites prevents ACF-dependent Top2 accumulation. Our data reveal the effect of Top2 poisons on the global Top2 binding landscape and highlights the role of Uls1 in antagonizing Top2 function. Remodelling Top2 binding is thus an important new means by which Snf2 enzymes promote genome stability.
Assuntos
DNA Helicases/metabolismo , DNA Topoisomerases Tipo II/metabolismo , Proteínas de Saccharomyces cerevisiae/metabolismo , Acriflavina/toxicidade , DNA Helicases/química , DNA Helicases/genética , DNA Topoisomerases Tipo II/efeitos dos fármacos , DNA Fúngico/metabolismo , Deleção de Genes , Saccharomyces cerevisiae/efeitos dos fármacos , Saccharomyces cerevisiae/genética , Proteínas de Saccharomyces cerevisiae/química , Proteínas de Saccharomyces cerevisiae/genéticaRESUMO
Proflavine derivatives are extremely interesting chemotherapeutic agents, which have shown promising pharmaceutical potential due to their wide range of biological activities. This review summarizes the current state of research into the anticancer, antimicrobial, antimalarial and antileishmanial properties of these attractive compounds. Our attention has focused on new classes of proflavine conjugates, which display significant levels of anticancer activity. Highly promising cytotoxic properties have been identified in proflavine conjugates with imidazolidinones, ureas and thioureas. In particular, proflavine-dialkyldithioureas displayed substantial cytotoxic effect against the human leukemia HL-60 cells with IC50 values from 7.2 to 34.0 µm. As well, palladium complexes with proflavine ligand have important biologic activity. The LC50 values of these complexes were significantly lower than that of cisplatin against the SK-BR-3 cell line.
Assuntos
Acriflavina/farmacologia , Anti-Infecciosos/farmacologia , Antineoplásicos/farmacologia , Proflavina/farmacologia , Acriflavina/análogos & derivados , Acriflavina/toxicidade , Animais , Anti-Infecciosos/toxicidade , Antineoplásicos/toxicidade , Relação Dose-Resposta a Droga , Humanos , Concentração Inibidora 50 , Estrutura Molecular , Proflavina/análogos & derivados , Proflavina/toxicidade , Relação Estrutura-AtividadeRESUMO
Acidophilic bacterium, Acidiphilium symbioticum H8, is resistant to high levels of several heavy metals, hydrophobic agents, and organic solvents. The approximately 9.6 kb plasmid pASH8, was purified, digested with HindIII, and sub-cloned in pUC19 at the respective site. Three different fragment size clones were achieved. The clones were completely sequenced and analyzed. The first clone encodes for a single putative open reading frame (ORF), which showed significant homology to several rusticyaninA1 proteins. The second clone encodes for a 43-kDa protein, which has conserved domain homology with several outer envelop TolC proteins. The clone with pASH8 tolC gene can functionally complement an Escherichia coli tolC mutant strain, making it resistant to several toxic hydrophobic agents, earlier for which it was sensitive. The tolC gene was found to be essential for imparting resistance to the clone toward these toxic hydrophobic agents. The third clone encodes for a putative 318-aa AcrA (acriflavine resistance protein A) protein and the clone was resistance to plasmid curing dye acriflavine. The clone also has a truncated ORF, which showed significant homology to cation-efflux pump AcrB. This study is the first to report a multi-drug efflux system to be encoded on a plasmid of any Acidiphilium strain.
Assuntos
Acidiphilium/efeitos dos fármacos , Acidiphilium/genética , Farmacorresistência Bacteriana , Proteínas de Membrana Transportadoras/genética , Proteínas de Membrana Transportadoras/metabolismo , Metais Pesados/metabolismo , Acriflavina/metabolismo , Acriflavina/toxicidade , Proteínas da Membrana Bacteriana Externa , Clonagem Molecular , DNA Bacteriano/química , DNA Bacteriano/genética , Escherichia coli/genética , Proteínas de Escherichia coli , Teste de Complementação Genética , Proteínas de Membrana Transportadoras/deficiência , Metais Pesados/toxicidade , Dados de Sequência Molecular , Fases de Leitura Aberta , Plasmídeos , Análise de Sequência de DNARESUMO
Caffeine and theophylline are mutagenic at high concentration in the B. subtilis multigene sporulation test for mutagens; caffeine is a stronger mutagen than theophylline in this test. An unusually high fraction of the mutant colonies appear to be phenotypically identical, as judged by colonial morphology and microscopic appearance of the vegetative cells. These mutants do not bring about the pH increase normally associated with sporulation of B. subtilis; such behavior is frequently associated with lack of a functional tricarboxylic acid (TCA) cycle, essential for normal sporulation of this species. Similar mutants have not been noted in the course of screening a variety of well-known mutagens, including acriflavine. Caffeine is maximally effective in inducing these mutants about 10 min after germination commences. Adenosine greatly reduces the ability of caffeine to induce these mutants.
Assuntos
Acriflavina/toxicidade , Aminoacridinas/toxicidade , Bacillus subtilis/efeitos dos fármacos , Cafeína/toxicidade , Mutagênicos , Mutação , Teofilina/toxicidade , Bacillus subtilis/genética , Bacillus subtilis/crescimento & desenvolvimento , Testes de Mutagenicidade , Fenótipo , Relação Estrutura-AtividadeRESUMO
Acriflavine hydrochloride (AFN) is a prospective drug worn in the eradication of HIV1 infection. The toxicity and adverse side effects renders the potent drug to limits its usage. However, to overcome the dilemma we have aimed to select carriers with great complexation efficiencies in different cyclodextrins (CDs) of varying cavity size. The interaction of AFN with α, ß and γ-CDs were investigated using absorption and steady state as well as lifetime measurements. From the obtained data it was found that AFN fits in the cavity of α and ß-CDs but unable to form inclusion complex with γ-CD. The effect of quencher molecules during the inclusion phenomena of AFN with CDs was explored via steady state measurements. The nature of binding forces responsible for the inclusion of AFN with CDs was discussed by using thermodynamic parameters. Using Benesi-Hildebrand equation the stoichiometry of AFN with CDs was predominantly found to be 1:1. To get deeper in situ, the in vitro toxicity of AFN and its complexation product were probed by Artemia salina sp. The toxicity of AFN was reduced when complexed with α and ß-CDs.
Assuntos
Acriflavina/toxicidade , Testes de Toxicidade , alfa-Ciclodextrinas/química , beta-Ciclodextrinas/química , Acriflavina/química , Animais , Artemia/efeitos dos fármacos , Concentração de Íons de Hidrogênio , Cinética , Espectrometria de Fluorescência , TermodinâmicaRESUMO
Induction of single strand breaks in DNA was assessed by the conversion of supercoiled closed circular plasmid DNA into the open circular form. Euflavine produced single-strand breaks following irradiation but not in the control maintained in the dark. The single strand breaking activity of photoactivated euflavine was found to be dose-dependent. The effective dose conversion 50% (ED50) of the closed circular DNA to the open circular form was 0.53 microM. A comparison of 8 acridine compounds revealed that the ED50 of diaminoacridines such as euflavine, proflavine and acridine yellow or the 3,6-dimethylamino-derivative (acridine orange) was less than 1 microM while the ED50 values of the other acridines were greater than 80 microM. Euflavine was markedly inhibited by singlet oxygen scavengers such as NaN3, histidine, alpha-tocopherol or beta-carotene and partly inhibited by superoxide dismutase, mannitol or catalase. These results suggest that enflavine induces single strand breaks in DNA mainly by a type II photodynamic mechanism. Photodynamic single strand breaking activities appeared related to their mutagenic activities on yeast. This experimental system described here is useful for the quantitative assessment of the single strand breaking activities of various photosensitizers in vitro and for the determination of active oxygen species involved in those processes.