RESUMO
The SeqA protein is a negative regulator of initiation of DNA replication in the Escherichia coli chromosome. Here, we demonstrate that SeqA stimulates transcription from the bacteriophage lambda pR promoter both in vivo and in vitro. The activity of the lambda pL promoter was found not to be affected by this protein. SeqA-mediated stimulation of pR was dependent on the state of template methylation: transcription was activated on fully methylated and hemimethylated templates but not on an unmethylated template. Using electrophoretic mobility shift assay and electron microscopy, we demonstrated that SeqA interacts specifically with a pR promoter region located on both fully methylated and hemimethylated DNA molecules, but not on unmethylated DNA. The activity of SeqA was found to affect the initiation of lambda plasmid replication positively in vivo, probably via pR-dependent expression of lambda replication genes and transcriptional activation of ori lambda. We conclude that, apart from its function in the control of DNA replication, SeqA is also a specific transcription factor.
Assuntos
Escherichia coli/genética , Fatores de Transcrição/metabolismo , Proteínas da Membrana Bacteriana Externa , Bacteriófago lambda/genética , Proteínas de Ligação a DNA , Eletroforese/métodos , Escherichia coli/metabolismo , Proteínas de Escherichia coli , Dosagem de Genes , Mutação , Regiões Promotoras Genéticas , Fatores de Transcrição/genéticaRESUMO
Anisakis simplex, a nematode from the family Anisakidae, is a parasite of fish and mammals. It is a casual agent of a human disease called anisakiosis. We found that the assay based on PCR amplification of the ITS-1-5.8 S-ITS-2 fragment of rDNA and subsequent restriction fragment length polymorphism, previously described on the basis of A. simplex isolated solely from one geographical region, can be used as a general test for identification of this worm species. The restriction patterns analysed for four restriction enzymes were found to be identical in the case of all A. simplex individuals isolated from as different geographical regions as Baltic Sea, Norwegian Sea, Bering Sea and Sea of Okchotsk. Moreover, our results support the previously proposed hypothesis, based on the studies of isoenzymes, that there is a remarkable genetic homogeneity within A. simplex from different geographical regions.
Assuntos
Anisakis/genética , DNA de Protozoário/análise , Peixes/genética , Reação em Cadeia da Polimerase/métodos , Polimorfismo de Fragmento de Restrição , Animais , Anisakis/química , Anisakis/classificação , Primers do DNA/química , DNA Ribossômico/química , Marcadores Genéticos , Geografia , Japão , Larva/genética , Mapeamento por RestriçãoRESUMO
On infection of its host, Escherichia coli, bacteriophage lambda can follow one of two alternative developmental pathways: lytic or lysogenic. Here we demonstrate that the "lysis-versus-lysogenization" decision is influenced by guanosine tetraphosphate (ppGpp), a nucleotide that is synthesized in E. coli cells in response to amino acid or carbon source starvation. We found that the efficiency of lysogenization is the highest at ppGpp concentrations somewhat higher than the basal level; too low and too high levels of ppGpp result in less efficient lysogenization. Maintenance of the already integrated lambda prophage and phage lytic development were not significantly influenced in the host lacking ppGpp. We found that the level of HflB/FtsH protease, responsible for degradation of the CII protein, an activator of "lysogenic" promoters, depends on ppGpp concentration. The highest levels of HflB/FtsH was found in bacteria lacking ppGpp and in cells bearing increased concentrations of this nucleotide. Using lacZ fusions, we investigated the influence of ppGpp on activities of lambda promoters important at the stage of the lysis-versus-lysogenization decision. We found that each promoter is regulated differentially in response to the abundance of ppGpp. Moreover, our results suggest that the cAMP level may influence ppGpp concentration in cells. The mechanism of the ppGpp-mediated control of lambda development at the stage of the lysis-versus-lysogenization decision may be explained on the basis of differential influence of guanosine tetraphosphate on activities of p(L), p(R), p(E), p(I), and p(aQ) promoters and by dependence of HflB/FtsH protease level on ppGpp concentration.
Assuntos
Bacteriófago lambda/fisiologia , Escherichia coli/metabolismo , Escherichia coli/virologia , Guanosina Tetrafosfato/metabolismo , Lisogenia , Proteases Dependentes de ATP , Proteínas de Bactérias/metabolismo , Bacteriólise , Bacteriófago lambda/genética , AMP Cíclico/metabolismo , Escherichia coli/enzimologia , Escherichia coli/genética , Proteínas de Escherichia coli , Regulação Bacteriana da Expressão Gênica , Regulação Viral da Expressão Gênica , Genes Virais/genética , Guanosina Tetrafosfato/genética , Proteínas de Membrana/metabolismo , Mutação , Fenótipo , Regiões Promotoras Genéticas/genética , Fatores de Transcrição/metabolismo , Transcrição Gênica/genética , Proteínas ViraisRESUMO
The methods of preservation of the material for corneal grafting are developing already for many years. The method of conservation of the corneae is closely connected with the way of the collection of the material: the whole globe or only the cornea with a 2-3 mm strip of the sclera. The whole globe is put into a moisture chamber, instead the cornea with a strip of sclera is preserved in various preservation fluids.
Assuntos
Córnea , Transplante de Córnea , Preservação de Tecido/métodos , Transplante de Córnea/métodos , Humanos , Técnicas de Cultura de Órgãos/métodosRESUMO
The problems of diagnosis of primary hyperaldosteronism are described calling attention to a significant advance in the methods of localization diagnosis owing to the introduction of computed tomography of the adrenals.
