Pheromone Recognition and Selectivity by ComR Proteins among Streptococcus Species.

Erin Shanker,Gerd Prehna,Michael J Federle,Sylvie Nessler,Donald A Morrison,Antoine Talagas,Gongyi Zhang

doi:10.1371/journal.ppat.1005979

Abstract

Natural transformation, or competence, is an ability inherent to bacteria for the uptake of extracellular DNA. This process is central to bacterial evolution and allows for the rapid acquirement of new traits, such as antibiotic resistance in pathogenic microorganisms. For the Gram-positive bacteria genus Streptococcus, genes required for competence are under the regulation of quorum sensing (QS) mediated by peptide pheromones. One such system, ComRS, consists of a peptide (ComS) that is processed (XIP), secreted, and later imported into the cytoplasm, where it binds and activates the transcription factor ComR. ComR then engages in a positive feedback loop for the expression of ComS and the alternative sigma-factor SigX. Although ComRS are present in the majority of Streptococcus species, the sequence of both ComS/XIP and ComR diverge significantly, suggesting a mechanism for species-specific communication. To study possible cross-talk between streptococcal species in the regulation of competence, and to explore in detail the molecular interaction between ComR and XIP we undertook an interdisciplinary approach. We developed a ‘test-bed’ assay to measure the activity of different ComR proteins in response to cognate and heterologous XIP peptides in vivo, revealing distinct ComR classes of strict, intermediate, and promiscuous specificity among species. We then solved an X-ray crystal structure of ComR from S. suis to further understand the interaction with XIP and to search for structural features in ComR proteins that may explain XIP recognition. Using the structure as a guide, we probed the apo conformation of the XIP-binding pocket by site-directed mutagenesis, both in test-bed cultures and biochemically in vitro. In alignments with ComR proteins from other species, we find that the pocket is lined by a variable and a conserved face, where residues of the conserved face contribute to ligand binding and the variable face discriminate among XIP peptides. Together, our results not only provide a model for XIP recognition and specificity, but also allow for the prediction of novel XIP peptides that induce ComR activity.

Highlights

Competence, or the ability of bacteria to import extracellular DNA, is a trait widely conserved across both Gram-negative and Gram-positive bacteria
Bacteria transmit chemical signals to each other in a process known as quorum sensing
Quorum sensing in many species of Gram-positive bacteria utilizes secreted short peptide signals that are sensed by protein receptors that alter gene expression

Summary

Introduction

Competence, or the ability of bacteria to import extracellular DNA, is a trait widely conserved across both Gram-negative and Gram-positive bacteria. This natural transformation process allows bacteria to acquire new genes that increase genetic diversity and fitness, such as the gain of antibiotic-resistance determinants [1]. Common among bacteria that are able to undergo natural transformation is a minimal set of genes enabling transport and integration of DNA by homologous recombination. Broad conservation of these genes, even among bacteria for which natural transformation has not been demonstrated in laboratory settings [2] provides a mechanistic explanation for the widespread evidence of horizontal gene transfer in bacteria, and strongly suggests their evolutionary importance. Competence in Streptococcus spp, is regulated either by the ComCDE system [9] or by the subject of our study, the ComRS pathway [10]

Methods

Results

Discussion

Conclusion