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1

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Identification of competence pheromone responsive genes in Streptococcus pneumoniae by use of DNA microarrays (2004)

Peterson, Scott N. ; Sung, Chang Kyoo ; Cline, Robin ; [et al.]

Oxford, UK : Blackwell Science Ltd

Molecular microbiology 51 (2004), S. 0

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ISSN: 1365-2958

Source: Blackwell Publishing Journal Backfiles 1879-2005

Topics: Biology , Medicine

Notes: Natural genetic transformation in Streptococcus pneumoniae is controlled in part by a quorum-sensing system mediated by a peptide pheromone called competence-stimulating peptide (CSP), which acts to coordinate transient activation of genes required for competence. To characterize the transcriptional response and regulatory events occurring when cells are exposed to competence pheromone, we constructed DNA microarrays and analysed the temporal expression profiles of 1817 among the 2129 unique predicted open reading frames present in the S. pneumoniae TIGR4 genome (84%). After CSP stimulation, responsive genes exhibited four temporally distinct expression profiles: early, late and delayed gene induction, and gene repression. At least eight early genes participate in competence regulation including comX, which encodes an alternative sigma factor. Late genes were dependent on ComX for CSP-induced expression, many playing important roles in transformation. Genes in the delayed class (third temporal wave) appear to be stress related. Genes repressed during the CSP response include ribosomal protein loci and other genes involved in protein synthesis. This study increased the number of identified CSP-responsive genes from approximately 40 to 188. Given the relatively large number of induced genes (6% of the genome), it was of interest to determine which genes provide functions essential to transformation. Many of the induced loci were subjected to gene disruption mutagenesis, allowing us to establish that among 124 CSP-inducible genes, 67 were individually dispensable for transformation, whereas 23 were required for transformation.

Type of Medium: Electronic Resource

URL: http://dx.doi.org/10.1046/j.1365-2958.2003.03907.x

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2

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Identification of ComW as a new component in the regulation of genetic transformation in Streptococcus pneumoniae (2004)

Luo, Ping ; Li, Haiying ; Morrison, Donald A.

Oxford, UK : Blackwell Science Ltd

Molecular microbiology 54 (2004), S. 0

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ISSN: 1365-2958

Source: Blackwell Publishing Journal Backfiles 1879-2005

Topics: Biology , Medicine

Notes: Regulation of competence for genetic transformation in Streptococcus pneumoniae depends on a quorum-sensing system, genes involved in DNA uptake and recombination and a link between these two gene sets. The alternative sigma factor ComX provides this link. ComE, the response regulator of the quorum-sensing system, is required for expression of ComX and other early genes. However, an unknown ComE-dependent regulator is also required for competence when comX is expressed under control of the raffinose-responsive promoter of the aga operon. The gene comW (SP0018) is required for a high level of competence and is regulated by the quorum-sensing system, but its function is unknown. To explore its role further, comW was cloned into the multicopy plasmid pMSP3535, under the control of a nisin-inducible promoter (PN), and transformed into pneumococcal strains containing a raffinose-inducible comX gene (PR::comX). Further introduction of a comE deletion blocked the endogenous CSP signal transduction pathway. In the resulting strain, competence was independent of CSP but depended on treatment with both nisin and raffinose, showing that coexpression of comW and comX complemented the comE deficiency. ComX protein accumulation and expression of a late competence gene in the above strain support the conclusion that ComW is a new positive factor involved in competence regulation.

Type of Medium: Electronic Resource

URL: http://dx.doi.org/10.1111/j.1365-2958.2004.04254.x

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3

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ComX is a unique link between multiple quorum sensing outputs and competence in Streptococcus pneumoniae (2003)

Luo, Ping ; Li, Haiying ; Morrison, Donald A.

Oxford, UK : Blackwell Science Ltd

Molecular microbiology 50 (2003), S. 0

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ISSN: 1365-2958

Source: Blackwell Publishing Journal Backfiles 1879-2005

Topics: Biology , Medicine

Notes: Natural competence for genetic transformation in Streptococcus pneumoniae is achieved directly by specific proteins that are involved in DNA uptake and chromosomal recombination, and is regulated indirectly by a quorum-sensing system encoded by two loci, comAB and comCDE. The alternative sigma factor, ComX, is thought to be the unique link between quorum sensing and competence-specific genes. To test this hypothesis, we replaced the quorum-sensing inducible promoter (PQ) of the comX gene with either a constitutive promoter (PC) or a raffinose-inducible promoter (PR), so that comX transcription would be independent of quorum sensing. Surprisingly, both competence and expression of late genes, such as ssbB, cglA or celB, were found to depend on CSP in these mutants. An unknown, CSP-dependent regulator was needed when comX was expressed from these ectopic promoters, and it appears to act post-transcriptionally. However, when a multicopy nisin-inducible ComX-overexpressing plasmid was introduced, pneumococcal cells developed competence in the presence of nisin even despite deletion of comE. At 1% of the normal protein peak level, ComX protein stimulated competence without the participation of the pheromone response circuit. Thus, ComX is a unique link to competence-specific genes, but depends on multiple outputs of quorum sensing for maximal expression.

Type of Medium: Electronic Resource

URL: http://dx.doi.org/10.1046/j.1365-2958.2003.03714.x

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4

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A functional genomic analysis of type 3 Streptococcus pneumoniae virulence (2001)

Lau, Gee W. ; Haataja, Sauli ; Lonetto, Michael ; [et al.]

Oxford, UK : Blackwell Science Ltd

Molecular microbiology 40 (2001), S. 0

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ISSN: 1365-2958

Source: Blackwell Publishing Journal Backfiles 1879-2005

Topics: Biology , Medicine

Notes: Streptococcus pneumoniae remains a serious cause of morbidity and mortality in humans, but relatively little is known about the molecular basis of its pathogenesis. We used signature-tagged mutagenesis together with an analysis of S. pneumoniae genome sequence to identify and characterize genes required for pathogenesis. A library of signature-tagged mutants was created by insertion–duplication mutagenesis, and 1786 strains were analysed for their inability to survive and replicate in murine models of pneumonia and bacteraemia. One hundred and eighty-six mutant strains were identified as attenuated, and 56 were selected for further genetic characterization based on their ability to excise the integrated plasmid spontaneously. The genomic DNA inserts of the plasmids were cloned in Escherichia coli and sequenced. These sequences were subjected to database searches, including the S. pneumoniae genome sequence, which allowed us to examine the chromosomal regions flanking these genes. Most of the insertions were in probable operons, but no pathogenicity islands were found. Forty-two novel virulence loci were identified. Five strains mutated in genes involved in gene regulation, cation transport or stress tolerance were shown to be highly attenuated when tested individually in a murine respiratory tract infection model. Additional experiments also suggest that induction of competence for genetic transformation has a role in virulence.

Type of Medium: Electronic Resource

URL: http://dx.doi.org/10.1046/j.1365-2958.2001.02335.x

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5

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Identification of the streptococcal competence-pheromone receptor (1996)

Håvarstein, Leiv Sigve ; Gaustad, Peter ; Nes, Ingolf F. ; [et al.]

Oxford BSL : Blackwell Science Ltd

Molecular microbiology 21 (1996), S. 0

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ISSN: 1365-2958

Source: Blackwell Publishing Journal Backfiles 1879-2005

Topics: Biology , Medicine

Notes: Competence for genetic transformation in certain species of streptococci has been known for many years to be induced by a secreted protease-sensitive pheromone, referred to as the competence factor or activator, which acts as a quorum-sensing signal to co-ordinate expression of late competence genes. We recently reported identification of the pheromone of Streptococcus pneumoniae strain Rx as a small unmodified peptide, which was termed competence-stimulating peptide (CSP). By identifying the gene (comC) encoding the Rx CSP we were able to show that it is synthesized as a precursor peptide containing an N-terminal double-glycine type leader. In the present work, we describe two alleles of the corresponding gene from Streptococcus gordonii strains Challis and NCTC 7865, which are strains with distinct competence pheromones and corresponding specific pheromone reactivities. In addition, the nucleic acid sequences of two genes located downstream of comC were determined; interestingly, these genes encode a two-component signal transduction system. We therefore speculated that their products, a histidine kinase (ComD) and its cognate response regulator (ComE), act downstream of the CSP in competence regulation. By tracing the CSP specificity of the competence response in these strains to strain-specific alleles of comD, we obtained evidence demonstrating that the histidine kinase ComD is the competence-pheromone receptor.

Type of Medium: Electronic Resource

URL: http://dx.doi.org/10.1046/j.1365-2958.1996.521416.x

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6

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Competence pheromone, oligopeptide permease, and induction of competence in Streptococcus pneumoniae (1996)

Alioing, Geneviève ; Granadel, Chantal ; Morrison, Donald A. ; [et al.]

Oxford, UK : Blackwell Publishing Ltd

Molecular microbiology 21 (1996), S. 0

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ISSN: 1365-2958

Source: Blackwell Publishing Journal Backfiles 1879-2005

Topics: Biology , Medicine

Notes: An unmodified heptadecapeptide pheromone capable of eliciting competence for genetic transformation in Streptococcus pneumoniae has recently been identified and characterized. In considering possible signaltransduction mechanisms for the peptide, the previously characterized Ami oligopeptide permease and the three highly homologous oligopeptide-binding lipoproteins, AmiA. AliA, and AliB, appeared to be good candidates for receptors. We therefore compared the spontaneous transformability of Ami, AliA and AliB mutants to that of an isogenic wild-type strain and we investigated the response of the various mutants to treatment with synthetic competence-stimulating peptide (CSP). Our results clearly demonstrate that neither Ami nor any of the three highly homologous oligopeptide-binding lipoproteins identified so far in S. pneumoniae are required for competence induction following treatment with synthetic CSP. Although the existence of a fourth unidentified oligopeptide-binding lipoprotein and/or a second oligopeptide permease operon could not be completely ruled out, we favour the hypothesis that CSP signal transmission rather involves a two-component regulatory system. Although none of the single or double Ami and Ali mutants tested appeared severely affected for competence, an exceptional aliB plasmid-insertion mutation abolished competence completely. In addition, the triple AmiA-AliA-AliB mutant differed from wild type in showing no sharp peak of competence but exhibiting transformability throughout the exponential phase of growth. These and previous observations are discussed and a general hypothesis is proposed to account for the modulation of competence by peptide permease mutants in S. pneumoniae.

Type of Medium: Electronic Resource

URL: http://dx.doi.org/10.1111/j.1365-2958.1996.tb02556.x

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7

Electronic Resource

Induction of transformation in streptococci by synthetic competence stimulating peptides (1998)

Gaustad, Peter ; Morrison, Donald A.

Springer

Methods in cell science 20 (1998), S. 65-70

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ISSN: 1573-0603

Keywords: Competence ; Competence stimulating peptide ; Oral streptococci ; Pheromone ; Pneumococcus ; Transformation

Source: Springer Online Journal Archives 1860-2000

Topics: Biology

Notes: Abstract Competence in genetic transformation is well known in oral streptococci as well as in the pathogenic pneumococcus. But the application of this genetic phenomenon as a tool to study the biology and the virulence of these organisms has been restricted to a limited number of strains, both by the stringency of the conditions required for competence development, and by the fact that wild types or encapsulated pneumococci transform poorly, or not at all. Many of these limitations have been alleviated by the use of synthetic competence stimulating peptides. These peptides are small, stable and inexpensive molecules inducing competence under a wide variety of conditions in strains belonging to the same pherotype. Their use circumvents many of the limitations to the expression of transformability in oral streptococci and the pneumococcus, and expands the opportunities for application of tools of molecular genetics to many strains of the species without prior genetic manipulation.

Type of Medium: Electronic Resource

URL: http://dx.doi.org/10.1023/A:1009882608636

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