RESUMO
An extragenic multicopy suppressor of the cell division inhibition caused by a MalE-MinE fusion protein in Escherichia coli has been mapped and identified as yaeO, one of the two short open reading frames (ORFs) of an operon located at 4.6 min. Overexpressed yaeO also suppressed some temperature-sensitive mutations in division genes ftsA and ftsQ, in chaperone gene groEL and in co-chaperone gene grpE. Gene yaeO, whose expression is regulated by growth rate, codes for a 9 kDa acidic protein with no obvious resemblance to other proteins. Transcription termination protein Rho co-purified with a histidine-tagged derivative of YaeO protein on Ni2+-NTA agarose columns in a manner that suggested direct YaeO-Rho interaction. In vivo, yaeO expression reduced termination at rho-dependent bacteriophage terminator tL1 and at the terminator of autogenously regulated gene rho. The suppression of temperature-sensitive phenotypes was a consequence of anti-termination, as it could be mimicked by a Prho::Tn10 mutation that reduces the expression and activity of gene rho. Our data indicate that the suppression is not caused by overexpression of the mutated genes, but presumably by indirect stabilization of the mutated proteins.
Assuntos
Transportadores de Cassetes de Ligação de ATP , Proteínas do Citoesqueleto , Proteínas de Escherichia coli , Genes Bacterianos , Proteínas de Transporte de Monossacarídeos , Mutação , Proteínas Periplásmicas de Ligação , Fator Rho , Transcrição Gênica , Proteínas de Bactérias/genética , Proteínas de Transporte/genética , Proteínas de Ciclo Celular , Mapeamento Cromossômico , Escherichia coli/genética , Regulação Bacteriana da Expressão Gênica , Proteínas Ligantes de Maltose , Óperon , TemperaturaAssuntos
Escherichia coli/genética , Regulação da Expressão Gênica , Chaperonas Moleculares/genética , Plasmídeos/genética , Dobramento de Proteína , Clonagem Molecular , Eletroforese em Gel de Poliacrilamida , Chaperonas Moleculares/metabolismo , Regiões Promotoras Genéticas , Proteínas Recombinantes/biossíntese , Proteínas Recombinantes/químicaRESUMO
The recently identified recA gene of the naturally transformable bacterium Streptococcus pneumoniae has been further characterized by constructing a recA null mutation and by investigating its regulation. The recA mutation has been shown to confer both DNA repair (as judged from sensitivity to u.v. and methyl methane sulphonate) and recombination deficiencies. Plasmid transformation into the recA mutant was also drastically reduced. Western blotting established that recA gene expression is increased several fold at the onset of competence for genetic transformation. Increased expression was associated with the appearance of a recA-specific transcript, approximately 5.7 kb long. This transcript indicated that recA is part of a competence-inducible (cin) operon. The major (about 4.3 kb) transcript detected from non-competent cells did not include cinA, the first gene in the operon, suggesting that this gene could be specifically required at some stage in the transformation process. Detection of small amounts of the 5.7 kb polycistronic mRNA in cells treated with mitomycin C suggested that the operon could also be damage inducible. In addition, mitomycin C treatment of a recA- lysogenic strain did not lead to prophage induction and cell lysis. This is unlike the situation of a recA+ lysogen. Together these results demonstrate that RecA controls lysogenic induction and suggest the existence of a SOS repair system in S. pneumoniae.