Regulation of Competence Development in Haemophilus influenzae

Macfadyen, Leah P.

doi:10.1006/jtbi.2000.2179

Cited by 31 publications

(42 citation statements)

References 37 publications

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“…However, both CRE sites are located between divergently transcribed operons and could effect transformation in either direction. Additionally, MacFadyen identified a site upstream of HI0937 (suhB) in strain Rd KW20 that was categorized as a binding site for the cAMP-CRP complex (35), but this latter site displays an 85% match with the consensus CRE. Our BLAST analyses of the ORFs surrounding each of these sites indicated that the original assignments might be incorrect.…”

Section: Resultsmentioning

confidence: 99%

“…Most of the operons containing genes known to be directly involved in DNA uptake and transformation in H. influenzae have a conserved 22-bp palindromic element called the competence regulatory element (CRE) directly upstream of the transcriptional start sites (32,35). The product of HI0601 tfoX (sxy) is required for transcription of operons containing the CRE, and insertions into tfoX completely abolish transformation development (32,61,63).…”

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confidence: 99%

“…Competence development is also eliminated by mutations in crp, the gene encoding the cAMP-receptor protein (CRP), but cannot be restored by treatment with exogenous cAMP (9). It has been suggested that CRE sites share significant similar-ities to CRP binding sites and that the cAMP-CRP complex might activate transcription of transformation genes by binding to the CRE in association with a competence-specific factor such as TfoX (35).…”

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confidence: 99%

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Characterization of Three New Competence-Regulated Operons in Haemophilus influenzae

VanWagoner

Whitby²,

Morton³

et al. 2004

J Bacteriol

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Haemophilus influenzae is one of a growing number of bacteria in which the natural ability to uptake exogenous DNA for potential genomic transformation has been recognized. To date, several operons involved in transformation in this organism have been described. These operons are characterized by a conserved 22-bp regulatory element upstream of the first gene and are induced coincident with transfer from rich to nutrientdepleted media. The previously identified operons comprised genes encoding proteins that include members of the type II secretion system and type IV pili, shown to be essential for transformation in other bacteria, and other proteins previously identified as required for transformation in H. influenzae. In the present study, three novel competence operons were identified by comparative genomics and transcriptional analysis. These operons have been further characterized by construction of null mutants and examination of the resulting transformation phenotypes. The putative protein encoded by the HI0366 gene was shown to be essential for DNA uptake, but not binding, and is homologous to a protein shown to be required for pilus biogenesis and twitching motility in Pseudomonas aeruginosa. An insertion in HI0939 abolished both DNA binding and uptake. The predicted product of this gene shares characteristics with PulJ, a pseudopilin involved in pullulanase export in Klebsiella oxytoca.

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Section: Resultsmentioning

confidence: 99%

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confidence: 99%

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confidence: 99%

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Characterization of Three New Competence-Regulated Operons in Haemophilus influenzae

VanWagoner

Whitby²,

Morton³

et al. 2004

J Bacteriol

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“…Transcription of some H. influenzae competence genes depends on the cAMP-cAMP receptor protein complex (5,19), and addition of cAMP to cells in exponential growth phase increases transformation frequencies from the threshold of detectability (ϳ10 Ϫ8 ) to about 10…”

Section: Identification Of Naturally Transformable a Actinomycetemcomentioning

confidence: 99%

Natural Transformation and DNA Uptake Signal Sequences in Actinobacillus actinomycetemcomitans

et al. 2002

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Actinobacillus actinomycetemcomitans is a member of the family Pasteurellaceae and a major causative agent of periodontitis. While several genera from this family are known to be competent for transformation, A. actinomycetemcomitans has yet to be fully characterized. Here we show that the competence of A. actinomycetemcomitans is remarkably similar to that of Haemophilus influenzae. In addition to having a similar frequency of transformation as H. influenzae, A. actinomycetemcomitans competence could also be induced at least 100-fold by cyclic AMP, suggesting that, as in H. influenzae, at least some competence genes are regulated by catabolite repression. Even more intriguing was the discovery of a putative A. actinomycetemcomitans DNA uptake signal sequence (USS) virtually identical to the USS of H. influenzae. Moreover, we provide evidence that this sequence functions in the same capacity as that from H. influenzae; the sequence appears to be required and sufficient for DNA uptake in a variety of assays. Finally, we have taken advantage of this system to develop a simple, highly efficient competence-based method for generating site-directed mutations in the wild-type fimbriated A. actinomycetemcomitans.Actinobacillus actinomycetemcomitans, a gram-negative, facultative, nonmotile coccobacillus, is a major causative agent of localized juvenile periodontitis and other forms of chronic periodontitis (36). Moreover, A. actinomycetemcomitans is transmissible among family members and may persist for long periods (4). A. actinomycetemcomitans can also spread from the oral cavity, causing a variety of nonoral infections (31, 36). Indeed, the genomic DNA of A. actinomycetemcomitans was identified in atherosclerotic lesions (14), suggesting a possible role of this bacterium in cardiovascular disease.Some A. actinomycetemcomitans strains are known to be naturally competent for transformation (23,30). In many bacteria, transformation occurs when cells enter the state of competence. This enables them to take up DNA and incorporate it into their genomes (8, 18). In the best-studied naturally competent gram-negative bacteria, Haemophilus influenzae, Haemophilus parainfluenzae, Neisseria gonorrhoeae, and Neisseria meningitidis, double-stranded donor DNA fragments are preferentially bound into a DNase-resistant state if they contain a short recognition sequence abundant in their own genomes, the uptake signal sequence (USS) (10, 11). Although uptake has been demonstrated elsewhere for A. actinomycetemcomitans in two preliminary studies (23, 30), a quantitative analysis has yet to be performed. In particular, neither of these studies examined any of the requirements for the induction of competence. Nor was a specific DNA uptake sequence requirement for transformation investigated.Because the function of the USS has been assumed to be exclusion of DNAs from other species, different genera were expected to use different USSs. However, only USSs from the very distantly related genera Haemophilus and Neisseria have been characteriz...

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“…The TfoX protein of H. influenzae itself does not seem to bind DNA, but to interact with CRP (Macfadyen, 2000;Redfield et al, 2005). Plasmids containing the tfoX gene make H. influenzae cells constitutively competent (Williams et al, 1994;Zulty and Barcak, 1995).…”

Section: Introductionmentioning

confidence: 99%

tfoX (sxy)-dependent transformation of Aggregatibacter (Actinobacillus) actinomycetemcomitans

Bhattacharjee

Fine

Figurski

2007

Gene

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AbstracttfoX (sxy) is a regulatory gene needed to turn on competence genes. Aggregatibacter (Actinobacillus) actinomycetemcomitans has a tfoX gene that is important for transformation. We cloned this gene on an IncQ plasmid downstream of the inducible tac promoter. When this plasmid was resident in cells of A. actinomycetemcomitans and tfoX was induced, the cells became competent for transformation. Several strains of A. actinomycetemcomitans, including different serotypes, as well as rough (adherent) and isogenic smooth (nonadherent) forms were tested. Only our two serotype f strains failed to be transformed. With the other strains, we could easily get transformants with extrachromosomal plasmid DNA when closed circular, replicative plasmid carrying an uptake signal sequence (USS) was used. When a replicative plasmid carrying a USS and cloned DNA from the chromosome of A. actinomycetemcomitans was linearized by digestion with a restriction endonuclease or when genomic DNA was used directly, the outcome was allelic exchange. To facilitate allelic exchange, we constructed a suicide plasmid (pMB78) that does not replicate in A. actinomycetemcomitans and carries a region with two inverted copies of a USS. This vector gave allelic exchange in the presence of cloned and induced tfoX easily and without digestion. Using transposon insertions in cloned katA DNA, we found that as little as 78 bp of homology at one of the ends was sufficient for that end to participate in allelic exchange. The cloning and induction of tfoX makes it possible to transform nearly any strain of A. actinomycetemcomitans, and allelic exchange has proven to be important for site-directed mutagenesis.

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Regulation of Competence Development in Haemophilus influenzae

Cited by 31 publications

References 37 publications

Characterization of Three New Competence-Regulated Operons in Haemophilus influenzae

Characterization of Three New Competence-Regulated Operons in Haemophilus influenzae

Natural Transformation and DNA Uptake Signal Sequences in Actinobacillus actinomycetemcomitans

tfoX (sxy)-dependent transformation of Aggregatibacter (Actinobacillus) actinomycetemcomitans

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