In Firmicutes bacteria, ATP-binding cassette (ABC) transporters have been recognized as important resistance determinants against antimicrobial peptides. Together with neighboring two-component systems (TCSs), which regulate their expression, they form specific detoxification modules. Both the transport permease and sensor kinase components show unusual domain architecture: the permeases contain a large extracellular domain, while the sensor kinases lack an obvious input domain. One of the best-characterized examples is the bacitracin resistance module BceRS-BceAB of Bacillus subtilis. Strikingly, in this system, the ABC transporter and TCS have an absolute mutual requirement for each other in both sensing of and resistance to bacitracin, suggesting a novel mode of signal transduction in which the transporter constitutes the actual sensor. We identified over 250 such BceAB-like ABC transporters in the current databases. They occurred almost exclusively in Firmicutes bacteria, and 80% of the transporters were associated with a BceRS-like TCS. Phylogenetic analyses of the permease and sensor kinase components revealed a tight evolutionary correlation. Our findings suggest a direct regulatory interaction between the ABC transporters and TCSs, mediating communication between both components. Based on their observed coclustering and conservation of response regulator binding sites, we could identify putative corresponding two-component systems for transporters lacking a regulatory system in their immediate neighborhood. Taken together, our results show that these types of ABC transporters and TCSs have coevolved to form self-sufficient detoxification modules against antimicrobial peptides, widely distributed among Firmicutes bacteria.
Sensing of and responding to environmental changes are of vital importance for microbial cells. Consequently, bacteria have evolved a plethora of signaling systems that usually sense biochemical cues either via direct ligand binding, thereby acting as “concentration sensors,” or by responding to downstream effects on bacterial physiology, such as structural damage to the cell. Here, we describe a novel, alternative signaling mechanism that effectively implements a “flux sensor” to regulate antibiotic resistance. It relies on a sensory complex consisting of a histidine kinase and an ABC transporter, in which the transporter fulfills the dual role of both the sensor of the antibiotic and the mediator of resistance against it. Combining systems biological modeling with in vivo experimentation, we show that these systems in fact respond to changes in activity of individual resistance transporters rather than to changes in the antibiotic concentration. Our model shows that the cell thereby adjusts the rate of de novo transporter synthesis to precisely the level needed for protection. Such a flux-sensing mechanism may serve as a cost-efficient produce-to-demand strategy, controlling a widely conserved class of antibiotic resistance systems.
Background: BceS-like histidine kinases strictly require BceAB-like ABC transporters for sensing of peptide antibiotics. Results: BceAB of Bacillus subtilis interacted with BceS in vivo and in vitro and specifically bound the substrate peptide bacitracin. Conclusion: Complex formation with the ABC transporter affects the activity of the histidine kinase. Significance: Histidine kinase and ABC transporter form a sensory complex for the detection of peptide antibiotics.
In the low-G؉C-content Gram-positive bacteria, resistance to antimicrobial peptides is often mediated by so-called resistance modules. These consist of a two-component system and an ATP-binding cassette transporter and are characterized by an unusual mode of signal transduction where the transporter acts as a sensor of antimicrobial peptides, because the histidine kinase alone cannot detect the substrates directly. Thus, the transporters fulfill a dual function as sensors and detoxification systems to confer resistance, but the mechanistic details of these processes are unknown. The paradigm and best-understood example for this is the BceRS-BceAB module of Bacillus subtilis, which mediates resistance to bacitracin, mersacidin, and actagardine. Using a random mutagenesis approach, we here show that mutations that affect specific functions of the transporter BceAB are primarily found in the C-terminal region of the permease, BceB, particularly in the eighth transmembrane helix. Further, we show that while signaling and resistance are functionally interconnected, several mutations could be identified that strongly affected one activity of the transporter but had only minor effects on the other. Thus, a partial genetic separation of the two properties could be achieved by single amino acid replacements, providing first insights into the signaling mechanism of these unusual modules.
Two-component systems are major signal transduction pathways, which consist of histidine kinases and response regulators that communicate through phosphorylation. Here, we highlight a distinct class of single-domain response regulators containing the PFXFATG[G/Y] motif that are activated by a mechanism distinct from the Y-T coupling described for prototypical receiver domains. We first solved the structures of inactive and active SdrG, a representative of the FAT GUY family, and then biochemically and genetically characterized variants in which residues in this motif were mutated. Our results support a model of activation mainly driven by a conserved lysine and reveal that the rotation of the threonine induces the reorganization of several aromatic residues in and around the PFXFATG[G/Y] motif to generate intermediates resembling those occurring during classical Y-T coupling. Overall, this helps define a new subfamily of response regulators that emerge as important players in physiological adaptation.
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