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Developmental Regulation of the Streptomyces lividans ram Genes: Involvement of RamR in Regulation of the ramCSAB Operon. Bart J. F. Keijser, 2002.Streptomycetes are filamentous soil bacteria that produce spores through a complex process of morphological differentiation . The ram cluster plays an important part during the development . The ram genes encode a membrane-bound kinase (RamC), a small protein (RamS), components of an ABC transporter (RamAB), and a response regulator (RamR) . While the introduction of an extra copy of the ram cluster accelerates development in Streptomyces lividans, ramABR disruption mutants are unable to produce aerial hyphae and spores . The developmental regulation of ram gene transcription was analyzed . Transcription of the ram genes occurred only on solid rich media and not on minimal media . The ramR gene is transcribed from a single promoter during all growth stages, with the highest levels during aerial growth . The ramCSAB genes comprise one operon and are transcribed from one principal promoter, P1, directly upstream of ramC . Transcription of ramCSAB was already observed during vegetative growth, but was strongly upregulated upon initiation of formation of aerial hyphae and was decreased during late stages of development . A large inverted repeat located downstream of ramS terminated the majority of transcripts . The introduction of ramR on a multicopy vector in S . lividans strongly induced P1 activity, while disruption of this regulator eliminated all P1 promoter activity . This shows that ramR is a crucial activator of ramCSAB transcription . Importantly, in bldA, bldB, bldD, or bldH mutants, ramR and ramCSAB are not transcribed, while ram gene transcription was observed in the earliest whi mutant, whiG . This indicates that the transcription of the ram genes marks the transition from vegetative to aerial growth . Rex-Centric Mutualism. Roderick A. Slavcev, 2002.We asked whether Rex exclusion encoded by a lambda prophage confers a protective or a cell-killing phenotype . We found that the Rex system can channel lysogenic cells into an arrested growth phase that gives an overall protective ability to the host despite some associated killing . Transcription Activation at Escherichia coli FNR-Dependent Promoters by the Gonococcal FNR Protein: Effects of a Novel S18F Substitution and Comparisons with the Corresponding Substitution in E . coli FNR. Tim Overton, 2003.The Neisseria gonorrhoeae genome encodes a homologue of the Escherichia coli FNR protein (the fumarate and nitrate reductase regulator) . Despite its similarity to E . coli FNR, the gonococcal FNR only partially complemented an E . coli fnr mutation . After error-prone PCR mutagenesis of the gonococcal fnr gene, we identified four mutant fnr derivatives carrying the same S18F substitution, and we showed that the mutant FNR could activate transcription from a range of class I and class II FNR-dependent promoters in E . coli . Prompted by the similarities between gonococcal and E . coli FNR, we made changes in gonococcal fnr that created substitutions that are equivalent to previously characterized substitutions in E . coli FNR . First, our experiments showed that cysteine, C116, in the gonococcal FNR, equivalent to C122 in E . coli FNR, is essential, presumably because, as in E . coli FNR, it binds to an iron-sulfur center . Second, the L22H and D148A substitutions in gonococcal FNR were made . These changes are equivalent to the L28H and D154A changes in E . coli FNR, which had been shown to increase FNR activity in the presence of oxygen . We show that the effects of these substitutions in gonococcal FNR are distinct from those of the S18F substitution . Similarly, substitutions in the putative activating regions of gonococcal FNR were made . We show that the activity of gonococcal FNR in E . coli can be increased by transplanting certain activating regions from E . coli FNR . The effects of these substitutions are additive to those due to S18F . From these data, we conclude that the effects of the S18F substitution in gonococcal FNR are distinct from the effects of the other substitutions . S18 is immediately adjacent to one of three N-terminal cysteine residues that coordinate the iron-sulfur center, and thus the S18F substitution is most likely to stabilize this center . Support for this came from complementary experiments in which we created the S24F substitution in E . coli FNR, which is equivalent to the S18F substitution in gonococcal FNR . Our results show that the S24F substitution changes the activity of E . coli FNR and that the changes are distinct from those due to previously characterized substitutions .
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