Review TRENDS in Microbiology Vol.11 No.8 August 2003 359 Making informed decisions: regulatory interactions between two-component systems Jetta J.E. Bijlsma and Eduardo A. Groisman Department of Molecular Microbiology, Howard Hughes Medical Institute, Washington University School of Medicine, Campus Box 8230, 660 S. Euclid Avenue, St Louis, MO 63110, USA Bacteria experience a multitude of stresses in the com- phosphorylate from small molecular weight phosphoryl plex niches they inhabit. These stresses are denoted by donors, such as acetyl phosphate and carbamoyl phos- specific signals that typically alter the activity of individ- phate [3,4]. In addition, the vast majority of sensor kinases ual two-component regulatory systems. Processing of exhibit phosphatase activity towards their cognate multiple signals by different two-component systems response regulators. Although most two-component sys- occurs when multiple sensors interact with a single tems entail a single His-to-Asp phosphotransfer event response regulator, by phosphatases interrupting phos- between a histidine kinase and a response regulator, there phoryl transfer in phosphorelays and through transcrip- is a subset that consists of a three-step His-Asp-His-Asp tional and post-transcriptional mechanisms. Gaining phosphorelay (Fig. 1b) [1,2]. The extra phosphorylatable insight into these mechanisms provides an understand- domains in a phosphorelay can be present in separate ing at the molecular level of bacterial adaptation to ever proteins or be part of multi-domain (i.e. hybrid) sensor changing multifaceted environments. proteins, which, in either case, will ultimately donate the A crucial question in signal transduction is: how does an (a) organism integrate multiple inputs into the most appro- Sensor Response regulator priate action? Most of the present knowledge on bacterial Sensing Kinase + P Receiver Regulatory gene regulation is based on studies conducted in labora- domain domain domain domain tory settings, where conditions are carefully controlled and, typically, only one stress is applied at a time. Although this reductionist approach has been successful H D in elucidating several regulatory mechanisms, it does not take into consideration the multitude of simultaneous ATP ADP stresses that bacteria experience in the niches they occupy. We are now beginning to uncover how bacteria (b) adjust their behavior in response to multiple cues from complex environments. H D Bacteria sense environmental conditions using a + P ATP + P variety of systems, which include secondary metabolites ADP and ions, such as cyclic adenosine monophosphate and + P Fe2þ, and regulatory proteins, such as catabolite regulat- D H ory protein (CRP) and ferric uptake regulator (Fur), whose affinity for DNA changes on ligand binding. However, two- component systems are the predominant form of signal TRENDS in Microbiology transduction used by bacteria to respond to environmental Fig. 1. Two-component systems consist of a sensor protein (histidine kinase) that stresses [1,2]. Two-component systems usually consist of a transfers a high-energy phosphoryl group to the response regulator, which is sensor protein (or histidine kinase) and a cognate response often a transcription factor. (a) The sensor protein autophosphorylates on a con- regulator. Sensor proteins use adenosine triphosphate served histidine residue (H) in the kinases domain (blue), often in response to sig- nal sensing in the sensing domain (purple). The high-energy phosphoryl group (P) (ATP) to autophosphorylate at a conserved histidine is then transferred to a conserved aspartate (D) residue in the receiver domain residue, often in response to a specific signal. The high- (green) of the response regulator, often changing the ability of its regulatory energy phosphoryl group is then transferred to a con- domain (grey) to bind DNA. (b) The His-Asp-His-Asp three-step phosphorelay con- sists of the transfer of the phosphoryl group from the conserved histidine in the served aspartate residue in the response regulator, often sensor to additional phosphorylatable aspartate and histidine residues in auxiliary altering its ability to bind target DNA sequences (Fig. 1a) domains before transfer to the aspartate in the receiver domain of the response regulator. Many different arrangements of these extra phosphorylatable [1,2]. Certain response regulators have the ability to domains exist within proteins and pathways. They are either present in separate proteins, a representative example of which is depicted here, or part of multi- Corresponding author: Eduardo A. Groisman ([email protected]). domain (hybrid) sensors. http://timi.trends.com 0966-842X/03/$ - see front matter q 2003 Elsevier Ltd. All rights reserved. doi:10.1016/S0966-842X(03)00176-8 360 Review TRENDS in Microbiology Vol.11 No.8 August 2003 phosphoryl group to the response regulator controlling chemoreceptors [13,14]. The chemoreceptor clusters form gene transcription (Fig. 1b). a complex with CheA and CheW [14,15], thus enabling How is specificity between cognate sensor/regulator simultaneous processing of multiple signals. Furthermore, pairs maintained in organisms harboring multiple two- ligand binding by one chemoreceptor influences the component systems? Based on conserved residues around surrounding chemoreceptors, leading to increased stimu- the phosphorylatable histidine, sensor proteins have been lation of CheA kinase activity [14,16,17]. divided into two major families and several minor ones [5]. A similar convergence of sensor proteins controls The conserved residues of sensor proteins are proposed to sporulation in the Gram-positive bacterium Bacillus function as ‘anchors’ that mediate the interaction with subtilis. Here, five related histidine kinases – KinA, conserved residues around the phosphorylatable aspar- KinB, KinC, KinD and KinE – phosphorylate the response tate in the response regulator, bringing the catalytic regulator Spo0F (Fig. 2a) [18]. The Kin proteins exhibit residues of both molecules in close proximity [6,7]. sequence similarity in the residues surrounding the Specificity between a cognate sensor/regulator pair is phosphorylatable histidine residue that will donate the then mediated by hypervariable residues around the phosphoryl group to the conserved aspartatyl residue in phosphorylatable aspartate in the response regulators Spo0F but differ in their putative sensing domains, [6,8]. This notion is strengthened by the observation that indicating that they respond to different ligands or mutations in the hypervariable region of the response conditions. Consistent with this notion, the severity of regulator PhoB increase its affinity for the non-cognate the sporulation defect displayed by mutants in KinA, KinB sensor VanS [9]. or KinC depends on the growth conditions [19]. Two-component systems are ideally suited to function in Quorum sensing governs bioluminescence in the mar- the integration of multiple signals because the membrane ine bacterium Vibrio harveyi in response to two different location of the sensor protein allows transduction of autoinducers: AI-1, specifically produced and detected by environmental cues, and the high degree of specificity V. harveyi, and AI-2, a product of the metabolism of a wide of each sensor/regulator pair ensures a physiological variety of microorganisms [20]. These autoinducers are response. Different two-component systems can process sensed by two different hybrid histidine kinases: LuxN, multiple signals into a particular response by interacting which senses AI-1, and LuxQ, which responds to AI-2 via with one another and/or by phosphorylating from small the periplasmic protein LuxP. These sensors converge onto molecular weight phosphodonors [10,11]. This article a histidine-containing phosphotransfer protein termed will discuss: (1) how multiple sensors interact with a LuxU, which will ultimately donate the phosphate to the single response regulator (Fig. 2a); (2) the phosphorylation response regulator LuxO [21]. Phosphorylated LuxO of response regulators by non-cognate sensors (Fig. 2b); activates a repressor that turns off transcription of the (3) the ability of phosphatases to interrupt the phosphoryl bioluminescence genes. The potential for AI-1 and AI-2 to flow in His-Asp-His-Asp phosphorelays (Fig. 2a) and control independently the phosphorylated state of LuxO (4) transcriptional and post-transcriptional control mech- could allow V. harveyi to respond not only to its own cell anisms (Fig. 2c, Fig. 2d). density but also to that of other microbes. However, when Multiple sensors acting on a single response regulator both autoinducers are present, expression of the LuxO- Different signals can be fed into the same regulatory controlled genes changes 10- to 100-fold more than when pathway if the phosphorylated state of a single response only one of them is present, suggesting that the quorum- regulator is modified by several cognate sensors, each sensing circuit of V. harveyi functions as a ‘coincidence responding to a different signal. The best-studied example detector’ that discriminates between the presence of both of signal integration is bacterial chemotaxis, where autoinducers and all other conditions (i.e. the presence of swimming behavior is modified in response to changes in either autoinducer or no autoinducer at all) [21]. the concentration of different substrates. These changes E. coli harbors a pair of two-component systems – are sensed by distinct chemoreceptors that, through the NarQ/NarP