Abstract
Two small quorum sensing (QS) peptides regulate competence in S. mutans in a cell density dependent manner: XIP (sigX inducing peptide) and CSP (competence stimulating peptide). Depending on the environmental conditions isogenic S. mutans cells can split into a competent and non-competent subpopulation. The origin of this population heterogeneity has not been experimentally determined and it is unknown how the two QS systems are connected. We developed a toolbox of single and dual fluorescent reporter strains and systematically knocked out key genes of the competence signaling cascade in the reporter strain backgrounds. By following signal propagation on the single cell level we discovered that the master regulator of competence, the alternative sigma factor SigX, directly controls expression of the response regulator for bacteriocin synthesis ComE. Consequently, a SigX binding motif (cin-box) was identified in the promoter region of comE. Overexpressing the genetic components involved in competence development demonstrated that ComRS represents the origin of bimodality and determines the modality of the downstream regulators SigX and ComE. Moreover these analysis showed that there is no direct regulatory link between the two QS signaling cascades. Competence is induced through a hierarchical XIP signaling cascade, which has no regulatory input from the CSP cascade. CSP exclusively regulates bacteriocin synthesis. We suggest renaming it mutacin inducing peptide (MIP). Finally, using phosphomimetic comE mutants we show that unimodal bacteriocin production is controlled posttranslationally, thus solving the puzzling observation that in complex media competence is observed in a subpopulation only, while at the same time all cells produce bacteriocins. The control of both bacteriocin synthesis and competence through the alternative sigma-factor SigX suggests that S. mutans increases its genetic repertoire via QS controlled predation on neighboring species in its natural habitat.
Highlights
Horizontal gene transfer in prokaryotes is mediated via three distinct mechanisms comprising conjugation, transduction, and transformation [1,2]
Streptococcus mutans is a bacterium of the human dental plaque that contributes to caries development
It controls two important survival mechanisms via a cell-density dependent communication system: The synthesis of peptide antibiotics, and of a membrane apparatus for genetic competence, i.e. the ability to take up external DNA and integrate it into its own genome
Summary
Horizontal gene transfer in prokaryotes is mediated via three distinct mechanisms comprising conjugation, transduction, and transformation [1,2]. I.e. the uptake of extracellular DNA from the environment via genetic competence, is a powerful process able to expand and modify the gene inventory in both Proteobacteria and Firmicutes [4] It requires a multi-protein complex localized in the cell membrane, many elements of which are highly conserved, and was studied in most detail in Bacillus subtilis and Vibrio cholerae [5,6]. SigX binding to the RNA polymerase activates transcription of a core set of ~ 20 “late” competence effector genes [7] They carry a nine bp cin-box in their promoter region and mediate the synthesis of proteins for DNA uptake and recombination [8]. The SigX regulon has been found in all streptococci sequenced to date, suggesting that genetic competence is ubiquitous in streptococci, until now it could only be demonstrated experimentally in very few [9]
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