Analysis of cepA and other Bacteroides fragilis genes reveals a unique promoter structure

Bayley, Douglas P.

doi:10.1016/s0378-1097(00)00472-9

Cited by 51 publications

(90 citation statements)

References 15 publications

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“…6. There is a variable gap between the 19 bp repeat element and the consensus promoter (Bayley et al, 2000) sequence situated within the invertible region, whereas the fixL repeat consistently overlaps the consensus promoter (Fig. 1).…”

Section: Resultsmentioning

confidence: 97%

“…At either end of these invertible regions are AT-rich sequences containing imperfect dyad symmetry spanning 30-32 bp (inverted repeat regions). The recently proposed consensus B. fragilis promoter regions (Bayley et al, 2000) are conserved and overlap the repeat region by 2 bp.…”

Section: Resultsmentioning

confidence: 99%

“…Primer pairs (Table 1) were designed such that the PCR product would bridge the right end of the invertible region. A product from primer pair A and C would be obtained when the proposed B. fragilis consensus promoter (Bayley et al, 2000) within the invertible region ( Fig. 1) was in the correct orientation with respect to the PS biosynthesis operon ('on' position); a product from primer pair B and C would be expected when in the opposite orientation ('off' position).…”

Section: Resultsmentioning

confidence: 99%

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Multiple inverted DNA repeats of Bacteroides fragilis that control polysaccharide antigenic variation are similar to the hin region inverted repeats of Salmonella typhimurium

et al. 2003

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The important opportunistic pathogen Bacteroides fragilis is a strictly anaerobic Gram-negative bacterium and a member of the normal resident human gastrointestinal microbiota. Our earlier studies indicated that there is considerable within-strain variation in polysaccharide expression, as detected by mAb labelling. Analysis of the genome sequence has revealed multiple invertible DNA regions, designated fragilis invertible (fin) regions, seven of which are upstream of polysaccharide biosynthesis loci and are approximately 226 bp in size. Using orientation-specific PCR primers and sequence analysis with populations enriched for one antigenic type, two of these invertible regions were assigned to heteropolymeric polysaccharides with different sizes of repeating units, as determined by PAGE pattern. The implication of these findings is that inversion of the fin regions switches biosynthesis of these polysaccharides off and on. The invertible regions are bound by inverted repeats of 30 or 32 bp with striking similarity to the Salmonella typhimurium H flagellar antigen inversion cross-over (hix) recombination sites of the invertible hin region. It has been demonstrated that a plasmid-encoded Hin invertase homologue (FinB), present in B. fragilis NCTC 9343, binds specifically to the invertible regions and the recombination sites have been designated as fragilis inversion cross-over (fix) sites.

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Section: Resultsmentioning

confidence: 97%

Section: Resultsmentioning

confidence: 99%

Section: Resultsmentioning

confidence: 99%

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Multiple inverted DNA repeats of Bacteroides fragilis that control polysaccharide antigenic variation are similar to the hin region inverted repeats of Salmonella typhimurium

et al. 2003

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“…The length of the upstream region and putative terminator location is also conserved in P. intermedia 17. Promoter sequences typical for Bacteroides fragilis (Bayley et al, 2000) or P. bryantii (Peterka, 2002) were not found in this region. The aforementioned L31 ribosomal protein shares 90 % amino acid sequence identity with its P. intermedia 17 counterpart.…”

Section: Resultsmentioning

confidence: 90%

Studies on Prevotella nuclease using a system for the controlled expression of cloned genes in P. bryantii TC1-1

Accetto

Avguštin

2007

Microbiology

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Studies on Prevotella nuclease using a system for the controlled expression of cloned genes in P. bryantii TC1-1 Available tools for genetic analysis in the anaerobic rumen bacterium Prevotella bryantii are limited to only two known systems for gene delivery, and no genes, with the exception of plasmid maintenance and selection genes, have been successfully expressed from plasmids in any species of the genus Prevotella until now. It is shown here that nucB, a newly cloned nuclease gene from P. bryantii, can be controllably expressed from shuttle vector pRH3 in P. bryantii strain TC1-1, depending on the tetracycline concentration in the growth medium. nucB expression is also growth-medium dependent and this regulation presumably takes place at the translational level. His-tagged NucB was purified from P. bryantii TC1-1 culture supernatant and was shown to degrade DNA as well as RNA; it is most likely a minor 36 kDa P. bryantii non-specific nuclease. INTRODUCTIONMembers of the bacterial genus Prevotella have been found in the rumen as well as in the oral cavity and large intestine of man and animals (Edwards et al., 2004;Paster et al., 2001;Duncan et al., 2003;Eckburg et al., 2005; Leser et al., 2002). In the rumen they are symbionts contributing to the degradation of plant cell-wall polysaccharides (Miyazaki et al., 2003) and protein metabolism (Walker et al., 2003), whereas oral prevotellas are implicated in dental plaque establishment (Kolenbrander et al., 2002), periodontitis (Paster et al., 2001), advanced caries (Martin et al., 2002), oro-pharyngeal abscesses (Brook, 2004) and noma, a grotesque childhood orofacial gangrene, now confined mostly to sub-Saharan Africa (Enwonwu et al., 2006). The genetic tools that would make possible more thorough studies of prevotellas are still undeveloped despite their ecological and medical importance. This appears to be primarily due to the large phylogenetic distance between prevotellas and other well-characterized micro-organisms, and as a consequence of the rather specific genetic elements, exemplified by promoters, containing distinct 27/233 consensus sequences that are recognized by unusual primary s factor (Vingadassalom et al., 2005). Nevertheless, in two studies (Shoemaker et al., 1991;Accetto et al., 2005) the plasmid replicons and tetQ selectable marker derived from Prevotella and related colonic Bacteroides strains were used successfully for shuttle-vector introduction into the xylan-degrading ruminal species Prevotella bryantii, which forms a distinct phylogenetic lineage apparently somewhat closer to oral prevotellas than the other ruminal Prevotella species (Avguštin et al., 2001). Based on the conjugal transfer protocol developed in the first study, a P. bryantii B 1 4 carboxymethylcellulase gene, fused to a cellulose-binding domain of Thermomonospora fusca cellulase, and driven by a Prevotella ruminicola 23 xylanase promoter, was introduced into P. bryantii B 1 4. The gene was expressed in Bacteroides uniformis 1108, but not in P. bryantii B 1 4, possibly ...

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“…However, atypical aspects of transcription initiation are still being identified in prokaryotes (Bayley et al, 2000;Gaal et al, 2001;Weiner et al, 2000). Novel structural properties of DNA are also being reported, but their effects on gene expression remain unknown (Petersen et al, 2003;Vanet et al, 2000;Vogel et al, 2003).…”

Section: Introductionmentioning

confidence: 99%

Isolation and characterization of putative Pseudobutyrivibrio ruminis promoters

Schoep

Gregg

2007

Microbiology

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Novel plasmids were constructed for the analysis of DNA fragments from the rumen bacterium Pseudobutyrivibrio ruminis. Five previously unidentified promoters were characterized using a novel primer extension method to identify transcription start sites. The genes downstream of these promoters were not identified, and their activity in expression of genomic traits in wild-type P. ruminis remains putative. Comparison with promoters from this and closely related species revealed a consensus sequence resembling the binding motif for the RNA polymerase s 70 -like factor complex. Consensus "35 and "10 sequences within these elements were TTGACA and ATAATATA respectively, interspaced by 15-16 bp. The consensus for the "10 element was extended by one nucleotide upstream and downstream of the standard hexamer (indicated in bold). Promoter strengths were measured by reverse transcription quantitative PCR and bglucuronidase assays. No correlation was found between the composition and context of elements within P. ruminis promoters, and promoter strength. However, a mutation within the "35 element of one promoter revealed that transcriptional strength and choice of transcription start site were sensitive to this single nucleotide change. INTRODUCTIONDNA binding specificity of bacterial RNA polymerase and the rate of transcription are largely dictated by the sequence of the upstream (UP) element (Estrem et al., 1998), and 210 and 235 motifs, which are common to most bacteria. However, atypical aspects of transcription initiation are still being identified in prokaryotes (Bayley et al., 2000;Gaal et al., 2001;Weiner et al., 2000). Novel structural properties of DNA are also being reported, but their effects on gene expression remain unknown (Petersen et al., 2003;Vanet et al., 2000;Vogel et al., 2003).Pseudobutyrivibrio ruminis (previously classified as Butyrivibrio fibrisolvens) is present in the rumen (Kopecny et al., 2003) and gastrointestinal tract of some animals. Butyrivibrio-like organisms may account for as much as 24-30 % of culturable bacteria from the rumen (Forster et al., 1996), although individual species may constitute much lower proportions of total bacterial numbers (Kobayashi et al., 2000). These bacteria are important for projects that aim to alter rumen function using genetically modified bacteria (Brooker et al., 1989;Gregg et al., 1987 Gregg et al., , 1994Gregg et al., , 1998Gregg & Sharpe, 1991;Mackie & White, 1990;Rogers, 1990;Smith & Hespell, 1983;Teather, 1985;Teather & Forster, 1998). Understanding their transcriptional regulation is essential for controlling the expression of foreign genes within the rumen. Molecular mechanisms governing transcription initiation in P. ruminis and closely related species remain poorly understood. To date, only four promoters have been studied using transcriptional analysis in their species of origin: the flaA and flaB promoters in P. ruminis OR77 (Beard et al., 2000), the Pseudobutyrivibrio sp. OB156 thl promoter (Asanuma et al., 2003), and the rep promoter in B. ...

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Analysis of cepA and other Bacteroides fragilis genes reveals a unique promoter structure

Cited by 51 publications

References 15 publications

Multiple inverted DNA repeats of Bacteroides fragilis that control polysaccharide antigenic variation are similar to the hin region inverted repeats of Salmonella typhimurium

Multiple inverted DNA repeats of Bacteroides fragilis that control polysaccharide antigenic variation are similar to the hin region inverted repeats of Salmonella typhimurium

Studies on Prevotella nuclease using a system for the controlled expression of cloned genes in P. bryantii TC1-1

Isolation and characterization of putative Pseudobutyrivibrio ruminis promoters

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