Date Published: March 27, 2014
Publisher: Public Library of Science
Author(s): Laura A. Mike, Jacob E. Choby, Paul R. Brinkman, Lorenzo Q. Olive, Brendan F. Dutter, Samuel J. Ivan, Christopher M. Gibbs, Gary A. Sulikowski, Devin L. Stauff, Eric P. Skaar, Vanessa Sperandio.
Two-component signaling systems (TCSs) are one of the mechanisms that bacteria employ to sense and adapt to changes in the environment. A prototypical TCS functions as a phosphorelay from a membrane-bound sensor histidine kinase (HK) to a cytoplasmic response regulator (RR) that controls target gene expression. Despite significant homology in the signaling domains of HKs and RRs, TCSs are thought to typically function as linear systems with little to no cross-talk between non-cognate HK-RR pairs. Here we have identified several cell envelope acting compounds that stimulate a previously uncharacterized Bacillus anthracis TCS. Furthermore, this TCS cross-signals with the heme sensing TCS HssRS; therefore, we have named it HssRS interfacing TCS (HitRS). HssRS reciprocates cross-talk to HitRS, suggesting a link between heme toxicity and cell envelope stress. The signaling between HssRS and HitRS occurs in the parental B. anthracis strain; therefore, we classify HssRS-HitRS interactions as cross-regulation. Cross-talk between HssRS and HitRS occurs at both HK-RR and post-RR signaling junctions. Finally, HitRS also regulates a previously unstudied ABC transporter implicating this transporter in the response to cell envelope stress. This chemical biology approach to probing TCS signaling provides a new model for understanding how bacterial signaling networks are integrated to enable adaptation to complex environments such as those encountered during colonization of the vertebrate host.
One mechanism by which bacteria sense and adapt to their environment is through the use of two-component signaling systems (TCSs). A prototypical TCS consists of a membrane-bound histidine kinase (HK) and cytoplasmic response regulator (RR). In the presence of a specific signal, the HK autophosphorylates at a conserved intracellular histidine residue and then transfers the phosphate to an aspartate on the cognate RR. This phosphorylation event activates the RR, which binds to target promoter regions and subsequently regulates gene expression . In this manner the classic bacterial TCS is thought to function in a linear fashion, in that each HK has a defined input that results in a specific output from the RR.
Here, a small molecule activator (‘205) of S. aureus HssRS was used to identify a previously uncharacterized TCS, HitRS, in B. anthracis and dissect the complex signaling interactions between HitRS and HssRS. The interactions between HssRS and HitRS, which we have documented here, are summarized in Table 1, Table 2 and Figure 1. HitRS recognizes a DR in Phit that differs from the Phrt DR by four nucleotides (Figure 2). These four divergent nucleotides are sufficient to determine the distinct response of the promoter to either heme or ‘205 (Figure 4). The exception to this observation is that the high basal activity of the Phrt promoter requires the Phrt DR, but the Phrt DR is not sufficient to induce high basal activation of the Phit promoter. In total, the high basal activity of Phrt requires the Phrt DR, HssR, and the approximately 200 bp that encompass Phrt. This implicates that other cis- or trans-acting cellular factors influence the high basal level of B. anthracis Phrt activation.