Enterococci have a number of traits that allow them to respond and prevail in the most diverse and varying environmental conditions, both inside and outside a host. Their ability to sense changing environmental stimuli and orchestrating a response accordingly is of the utmost importance in its adaptation to these varying conditions (Hancock and Perego, 2004).
Two-component systems (TCSs)
Bacteria have different mechanisms available for the regulation of stress response. Besides sigma factors and small RNAs, most bacteria monitor and adapt to changing conditions through signal transduction involving two-component signal transduction systems. TCSs generally consist of a sensory histidine kinase (HK) and a cognate response regulator (RR). The HK senses the signal and relays the adaptive response through the transfer of a phosphoryl group to the RR, which can then act as
Chapter 1 a transcriptional regulator to modulate gene expression (Hancock and Perego, 2004). Most bacterial genomes encode several TCSs and enterococcal genomes are not unlike. Taken together, studies on E. faecalis TCSs can emphasize that two-component signal transduction systems govern important biological parameters of this organism ranging from environmental persistence, virulence and biofilm formation, to antibiotic resistance and stress response (Teng, Wang, Singh, Murray, and Weinstock, 2002; Le Breton et al., 2003; Hancock and Perego, 2004). Although all these roles associated to TCSs contribute together to the opportunistic character of E. faecalis, little is known about their target genes.
Quorum sensing (QS)
TCSs allow enterococci to sense environmental stimuli and to subsequently coordinate a suitable response. Part of these environmental stimuli can be autoinducers (AIs), molecules secreted by a bacterial population that accumulate extracellularly and induce genes in that same bacterial population. QS is that specific phenomenon: when a sufficient cell density has been reached, microorganisms communicate, coordinate and switch a set of behaviours among all individuals of the population, by the accumulation of AIs over a given threshold concentration (Hense et al., 2007).
As the three key determinants of AI concentration (cell density, mass-transfer properties and the spatial distribution of cells) can vary independently, cells sensing the AI concentration are unable to distinguish cell density from mass-transfer properties or spatial distribution. They can only assess the combination of these factors. This is why QS and diffusion sensing (DS), where cells use AI sensing to measure the mass-transfer properties of their environment, can be integrated. The recent notion of
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efficiency sensing (ES) helps unify the concepts of what cells sense, why cells sense and the evolutionary hypotheses of the fitness benefits derived from AI sensing (Hense et al., 2007).
Whether bacteria sense their density to allow them to engage in social behaviour that would benefit the group, in QS, or that sensing evolved as an autonomous activity of single cells to detect mass transfer limitations because of a direct fitness benefit for the individual, in DS, does not change the fact that enterococci are able to sense their specific AIs and adequately regulate gene expression.
It was demonstrated that the previously mentioned Fsr system was activated by AI concentration through the TCS Ehk15-Err15 (enterococcal HK and enterococcal RR) (FsrCA) (Nakayama et al., 2001). The fsr locus is comprised of 4 genes, fsrA, fsrB, fsrD, and fsrC, whose products form a system that responds to the extracellular accumulation of the gelatinase biosynthesis-activating pheromone (GBAP) peptide encoded by the fsrD gene that acts as an AI. FsrB acts as a cysteine protease-like processing enzyme involved in the processing of the FsrD peptide. Accumulation of this peptide in the extracellular space is sensed by the FsrC membrane HK, leading to the activation of the RR and transcription factor FsrA. All the FsrABDC proteins are necessary for autoregulation at a promoter located upstream of fsrB and for the expression of two E. faecalis virulence-related proteases, GelE and SprE, from a promoter located upstream of the gelE gene (Del Papa and Perego, 2011). This QS mechanism, that involves the Fsr system and a TCS, also plays a role in efficient biofilm formation, dissemination, and immune system evasion.
Unrelated to the well-known superfamily of TCS, another two component regulatory system, that involves the membrane component CylR1 and the DNA binding component CylR2, is necessary for the QS
Chapter 1 induction of the cytolysin operon, where cytolysin serves as an AI. As previously mentioned, the control of the cytolysin is regulated by a threshold concentration of the posttranslationally modified subunit CylLS.
CylLL has been shown to bind strongly to target cell membranes, allowing
free CylLS to accumulate above a critical induction threshold (Coburn et al.,
2004). This QS system provides a means by which the cytolysin is produced in elevated amounts only when a particular target cell is present (Clewell, 2007), contributing to the virulence of enterococcal species harbouring this trait.
Beyond controlling gene expression on a global scale and allowing communication within species, QS also allows bacteria to communicate between species. That communication is achieved via 4,5-dihydroxy-2,3- pentanedione (DPD), one of the conversion products of S- ribosylhomocysteine (SRH) by the enzyme LuxS, which spontaneously cyclizes into several furanones in chemical equilibrium, collectively referred to as autoinducer-2 (AI-2) (Hense et al., 2007). Specifically, the luxS gene that encodes the LuxS enzyme is present in roughly half of all sequenced bacterial genomes, AI-2 production has been verified in a large number of these species, and AI-2 controls gene expression in a variety of bacteria (Waters et al., 2003), being implicated in the regulation of many bacterial behaviours including biofilm formation, competence, the production of secondary metabolites like antibiotics, and virulence (Pereira et al., 2009). All enterococcal genomes publicly available (http://www.ncbi.nlm.nih.gov/ and http://www.broadinstitute.org/; related websites last visited: 2012.01.05) carry genes annotated as luxS genes, which can allow them to produce AI-2. While in some cases, AI-2 is clearly acting through a canonical QS mechanism, in others no relation to a committed communication between species could be ascribed (Pereira et al., 2009).
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