The sensor histidine kinase ArlS is necessary for Staphylococcus aureus to activate ArlR in response to nutrient availability

Staphylococcus aureus is a versatile opportunistic pathogen whose success is driven by its ability to adapt to diverse environments and host-imposed stresses. Two-component signal transduction systems, such as ArlRS, often mediate these adaptations. Loss of ArlRS or the response regulator ArlR alone impairs the ability of S. aureus to respond to host-imposed manganese starvation and glucose limitation. As sensor histidine kinases and response regulators frequently work as pairs, it has been assumed that ArlS senses and activates ArlR in response to these stimuli. However, recent work suggests that the sensor histidine kinase GraS can also activate ArlR, calling the contribution of ArlS in responding to manganese and glucose availability into question. The current studies reveal that ArlS is necessary to activate ArlR in response to manganese sequestration by the host immune effector calprotectin and glucose limitation. Although the loss of ArlS does not completely eliminate ArlR activity, this response regulator is no longer responsive to manganese or glucose availability in the absence of its cognate histidine kinase. Despite the residual activity of ArlR in the absence of ArlS, ArlR phosphorylation by ArlS is required for S. aureus to resist calprotectin-imposed metal starvation. Cumulatively, these findings contribute to the understanding of S. aureus signaling transduction in response to nutritional immunity and support the previous observation that indicates ArlRS is activated by a common signal derived from host-imposed manganese and glucose limitation. IMPORTANCE The ability of pathogens, including Staphylococcus aureus , to sense and adapt to diverse environments partially relies on two-component systems, such as ArlRS. Recent work revealed that the response regulator ArlR can be cross-activated by the sensor histidine kinase GraS, rendering the role of its cognate partner, ArlS, in response to manganese and glucose limitation uncertain. This study reveals that ArlS is necessary for the activation of ArlR in response to calprotectin and glucose limitation. Although a low level of ArlR activity remains in the absence of ArlS, ArlS phosphotransfer to ArlR is required for S. aureus to overcome calprotectin-induced nutritional stress. Collectively, this study provides fundamental information to understand how ArlRS mediates staphylococcal adaptation during infection.

Regulation of Resistance in Vancomycin-Resistant Enterococci: The VanRS Two-Component System

Vancomycin-resistant enterococci (VRE) are a serious threat to human health, with few treatment options being available. New therapeutics are urgently needed to relieve the health and economic burdens presented by VRE. A potential target for new therapeutics is the VanRS two-component system, which regulates the expression of vancomycin resistance in VRE. VanS is a sensor histidine kinase that detects vancomycin and in turn activates VanR; VanR is a response regulator that, when activated, directs expression of vancomycin-resistance genes. This review of VanRS examines how the expression of vancomycin resistance is regulated, and provides an update on one of the field’s most pressing questions: How does VanS sense vancomycin?

Dimer Asymmetry and Light Activation Mechanism in Brucella Blue-Light Sensor Histidine Kinase

ABSTRACT The ability to sense and respond to environmental cues is essential for adaptation and survival in living organisms. In bacteria, this process is accomplished by multidomain sensor histidine kinases that undergo autophosphorylation in response to specific stimuli, thereby triggering downstream signaling cascades. However, the molecular mechanism of allosteric activation is not fully understood in these important sensor proteins. Here, we report the full-length crystal structure of a blue light photoreceptor LOV histidine kinase (LOV-HK) involved in light-dependent virulence modulation in the pathogenic bacterium Brucella abortus. Joint analyses of dark and light structures determined in different signaling states have shown that LOV-HK transitions from a symmetric dark structure to a highly asymmetric light state. The initial local and subtle structural signal originated in the chromophore-binding LOV domain alters the dimer asymmetry via a coiled-coil rotary switch and helical bending in the helical spine. These amplified structural changes result in enhanced conformational flexibility and large-scale rearrangements that facilitate the phosphoryl transfer reaction in the HK domain. IMPORTANCE Bacteria employ two-component systems (TCSs) to sense and respond to changes in their surroundings. At the core of the TCS signaling pathway is the multidomain sensor histidine kinase, where the enzymatic activity of its output domain is allosterically controlled by the input signal perceived by the sensor domain. Here, we examine the structures and dynamics of a naturally occurring light-sensitive histidine kinase from the pathogen Brucella abortus in both its full-length and its truncated constructs. Direct comparisons between the structures captured in different signaling states have revealed concerted protein motions in an asymmetric dimer framework in response to light. Findings of this work provide mechanistic insights into modular sensory proteins that share a similar modular architecture.

Regulation of Chitin-Dependent Growth and Natural Competence in Vibrio parahaemolyticus

Vibrios can degrade chitin surfaces to soluble N-acetyl glucosamine oligosaccharides (GlcNAcn) that can be utilized as a carbon source and also induce a state of natural genetic competence. In this study, we characterized chitin-dependent growth and natural competence in Vibrio parahaemolyticus and its regulation. We found that growth on chitin was regulated through chitin sensors ChiS (sensor histidine kinase) and TfoS (transmembrane transcriptional regulator) by predominantly controlling the expression of chitinase VPA0055 (ChiA2) in a TfoX-dependent manner. The reduced growth of ΔchiA2, ΔchiS and ΔtfoS mutants highlighted the critical role played by ChiA2 in chitin breakdown. This growth defect of ΔchiA2 mutant could be recovered when chitin oligosaccharides GlcNAc2 or GlcNAc6 were supplied instead of chitin. The ΔtfoS mutant was also able to grow on GlcNAc2 but the ΔchiS mutant could not, which indicates that GlcNAc2 catabolic operon is dependent on ChiS and independent of TfoS. However, the ΔtfoS mutant was unable to utilize GlcNAc6 because the periplasmic enzymes required for the breakdown of GlcNAc6 were found to be downregulated at the mRNA level. We also showed that natural competence can be induced only by GlcNAc6, not GlcNAc2, because the expression of competence genes was significantly higher in the presence of GlcNAc6 compared to GlcNAc2. Moreover, this might be an indication that GlcNAc2 and GlcNAc6 were detected by different receptors. Therefore, we speculate that GlcNAc2-dependent activation of ChiS and GlcNAc6-dependent activation of TfoS might be crucial for the induction of natural competence in V. parahaemolyticus through the upregulation of the master competence regulator TfoX.

Structural insights into the histidine-containing phosphotransfer protein and receiver domain of sensor histidine kinase suggest a complex model in the two-component regulatory system in Pseudomonas aeruginosa

In Pseudomonas aeruginosa, an important opportunistic pathogen that causes numerous acute and chronic infections, the hybrid two-component system (TCS) regulates the swarming ability and biofilm formation with a multistep phosphorelay, and consists of hybrid-sensor histidine kinase (HK), histidine-containing phosphotransfer protein (Hpt) and response regulator (RR). In this work, two crystal structures of HptB and the receiver domain of HK PA1611 (PA1611REC) of P. aeruginosa have been determined in order to elucidate their interactions for the transfer of the phosphoryl group. The structure of HptB folds into an elongated four-helix bundle – helices α2, α3, α4 and α5, covered by the short N-terminal helix α1. The imidazole side chain of the conserved active-site histidine residue His57, located near the middle of helix α3, protrudes from the bundle and is exposed to solvent. The structure of PA1611REC possesses a conventional (β/α)5 topology with five-stranded parallel β-sheets folded in the central region, surrounded by five α-helices. The divalent Mg2+ ion is located in the negatively charged active-site cleft and interacts with Asp522, Asp565 and Arg567. The HptB–PA1611REC complex is further modeled to analyze the binding surface and interactions between the two proteins. The model shows a shape complementarity between the convex surface of PA1611REC and the kidney-shaped HptB with fewer residues and a different network involved in interactions compared with other TCS complexes, such as SLN1-R1/YPD1 from Saccharomyces cerevisiae and AHK5RD/AHP1 from Arabidopsis thaliana. These structural results provide a better understanding of the TCS in P. aeruginosa and could potentially lead to the discovery of a new treatment for infection.

Crystal Structure of a Proteolytic Fragment of the Sensor Histidine Kinase NarQ

Two-component signaling systems (TCSs) are a large and important class of sensory systems in bacteria, archaea, and some eukaryotes, yet their mechanism of action is still not fully understood from the structural point of view. Many TCS receptors are elongated flexible proteins with transmembrane (TM) regions, and are difficult to work with. Consequently, truncated fragments of the receptors are often used in structural studies. However, it is not fully clear whether the structures of the fragments correspond well to their native structures in the context of full-length proteins. Recently, we crystallized a fragment of Escherichia coli nitrate/nitrite sensor histidine kinase, NarQ, encompassing the sensor, TM, and HAMP domains. Here we report that a smaller proteolytic fragment consisting of the sensor and TM domains can also be crystallized using the in meso approach. The structure of the fragment is similar to the previously determined one, with minor differences in the vicinity of the truncation site. The results show that the crystallization of such sensor–TM fragments can be accomplished and can provide information on the packing of transmembrane helices, albeit limited, and that the proteolysis may or may not be a problem during crystallization.