MCP molecules typically have a periplasmic ligand-binding domain for monitoring attractant or repellent levels in the environment and a cytoplasmic signaling domain for communicating with the motor apparatus (1). The MCP signaling domain, highly conserved in structure, forms ternary complexes with two cytoplasmic proteins, CheA, a histidine kinase, and CheW, which couples CheA activity to chemoreceptor control (2, 3). Changes in receptor ligand occupancy trigger conformational changes in the signaling domain that in turn modulate the flux of phosphoryl groups from CheA to effector proteins that elicit the behavioral response (4, 5). MCPs are capable of detecting chemoeffector concentration changes of only a few parts per thousand over more than a five-log concentration range (6-9). The amplification mechanisms responsible for the high-gain signaling characteristics of bacterial chemoreceptors are still poorly understood but may rely on novel signaling principles that will prove to be widely used in biological signal transduction systems.Like many membrane receptors, MCP molecules are not uniformly distributed but rather clustered, typically at the cell poles (10). The CheA and CheW proteins also localize to these receptor clusters and are largely responsible for their integrity (10), suggesting that bacterial chemoreceptors form a two-dimensional lattice that is held together by bridging connections to CheA and CheW. Bray and colleagues (11-13) have theorized that receptors in such an array might, through conformational coupling, communicate their signaling states to neighboring receptors to produce a large gain in detection sensitivity. Experimental work, primarily with Escherichia coli, provides growing support for this notion.E. coli uses five MCP-family receptors to promote chemotactic movements toward different attractant compounds: Tar (aspartate and maltose), Tsr (serine), Tap (dipeptides), Trg (ribose and galactose), and Aer (oxygen and other electron acceptors). Tap, Trg, and, most likely, Aer are present in the cell at roughly 10% the levels of Tsr and Tar (14). Several lines of evidence indicate that high-and low-abundance receptors might signal collaboratively, and that clustering enhances their detection sensitivity. First, the ability of low-abundance receptors to mediate chemotactic responses implies that they are able to exert control over a substantial fraction of the CheA signaling molecules associated with the receptor array. Second, high-abundance receptors assist one another (15) and low-abundance receptors (16-18) in achieving the methylation changes needed to adapt to sensory stimuli. Third, a multivalent galactose ligand that promotes clustering of Trg (19) also served to recruit Tar and Tsr molecules to the cluster and greatly enhanced their detection sensitivity, implying that communication between receptors in a cluster produces signal amplification (20).In vitro studies of receptors and receptor fragments indicate that more than one receptor signaling domain is needed to form a ternar...
SummaryTo test the gearbox model of HAMP signalling in the Escherichia coli serine receptor, Tsr, we generated a series of amino acid replacements at each residue of the AS1 and AS2 helices. The residues most critical for Tsr function defined hydrophobic packing faces consistent with a four-helix bundle. Suppression patterns of helix lesions conformed to the predicted packing layers in the bundle. Although the properties and patterns of most AS1 and AS2 lesions were consistent with both proposed gearbox structures, some mutational features specifically indicate the functional importance of an x-da bundle over an alternative a-d bundle. These genetic data suggest that HAMP signalling could simply involve changes in the stability of its x-da bundle. We propose that Tsr HAMP controls output signals by modulating destabilizing phase clashes between the AS2 helices and the adjoining kinase control helices. Our model further proposes that chemoeffectors regulate HAMP bundle stability through a control cable connection between the transmembrane segments and AS1 helices. Attractant stimuli, which cause inward piston displacements in chemoreceptors, should reduce cable tension, thereby stabilizing the HAMP bundle. This study shows how transmembrane signalling and HAMP input-output control could occur without the helix rotations central to the gearbox model.
SUMMARY HAMP domains mediate input-output communication in many bacterial signaling proteins. To explore the dynamic bundle model of HAMP signaling (Zhou et al., Mol. Microbiol. 73: 801, 2009), we characterized the signal outputs of 118 HAMP missense mutants of the serine chemoreceptor, Tsr, by flagellar rotation patterns. Receptors with proline or charged amino acid replacements at critical hydrophobic packing residues in the AS1 and AS2 HAMP helices had locked kinase-off outputs, indicating that drastic destabilization of the Tsr-HAMP bundle prevents kinase activation, both in the absence and presence of the sensory adaptation enzymes, CheB and CheR. Attractant-mimic lesions that enhance the structural stability of the HAMP bundle also suppressed kinase activity, demonstrating that Tsr-HAMP has two kinase-off output states at opposite extremes of its stability range. HAMP mutants with locked-on kinase outputs appeared to have intermediate bundle stabilities, implying a biphasic relationship between HAMP stability and kinase activity. Some Tsr-HAMP mutant receptors exhibited reversed output responses to CheB and CheR action that are readily explained by a biphasic control logic. The findings of this study provide strong support for a three-state dynamic bundle model of HAMP signaling in Tsr, and possibly in other bacterial transducers as well.
Tsr, the serine chemoreceptor of Escherichia coli, has two signaling modes. One augments clockwise (CW) flagellar rotation, and the other augments counterclockwise (CCW) rotation. To identify the portion of the Tsr molecule responsible for these activities, we isolated soluble fragments of the Tsr cytoplasmic domain that could alter the flagellar rotation patterns of unstimulated wild-tpe cells. Residues 290 to 470 from wild-type Tsr generated a CW signal, whereas the same fragment with a single amino acid replacement (alanine 413 to valine) produced a CCW signal. The soluble components of the chemotaxis phosphorelay system needed for expression of these Tsr fragment signals were identified by epistasis analysis. Like full-length receptors, the fragments appeared to generate signals through interactions with the CheA autokinase and the CheW coupli factor. CheA was required for both signaling activities, whereas CheW was needed only for CW signaling. Purified Tsr fragments were also examined for effects on CheA autophosphorylation activity in vitro.Consistent with the in vivo findings, the CW fragment stimulated CheA, whereas the CCW f ent inhibited CheA. CheW was required for stimulation but not for inhibition. These findings demonstrate that a 180-residue segment of the Tsr cytoplasmic domain can produce two active signals. The CCW signal involves a direct contact between the receptor and the CheA kinase, whereas the CW signal requires participation of CheW as well. The correlation between the in vitro effects of Tsr signaling fragments on CheA activity and their in vivo behavioral effects lends convincing support to the phosphorelay model of chemotactic signaling.Methyl-accepting chemotaxis proteins (MCPs) mediate chemotactic responses in a variety of motile bacteria, ranging from archaebacteria to gram-positive and gram-negative eubacteria (reviewed in reference 11). These sensory transducers offer tractable models for molecular studies of transmembrane and intracellular signaling. Escherichia coli has four different MCPs that detect the attractants serine (Tsr), aspartate (Tar), dipeptides (Tap), and ribose and galactose (Trg). MCP molecules are about 550 amino acids in length and span the cytoplasmic membrane. They have a periplasmic ligand-binding domain that monitors attractant levels in the environment and a cytoplasmic signaling domain that controls rotation of the flagellar motors (14,21). Occupancy changes in the sensing domain modulate the output activity of the signaling domain to elicit chemotactic movements. For example, attractant increases promote smooth, forward swimming by reducing the probability of the episodes of clockwise (CW) flagellar rotation that cause turns or tumbling movements. Ensuing changes in MCP methylation state cancel excitatory stimulus signals to achieve sensory adaptation, thereby enabling the organism to detect further changes in its chemical environment (31).MCPs in E. coli appear to control swimming behavior by modulating the autophosphorylation activity of CheA, a cyto...
scite is a Brooklyn-based organization that helps researchers better discover and understand research articles through Smart Citations–citations that display the context of the citation and describe whether the article provides supporting or contrasting evidence. scite is used by students and researchers from around the world and is funded in part by the National Science Foundation and the National Institute on Drug Abuse of the National Institutes of Health.
hi@scite.ai
10624 S. Eastern Ave., Ste. A-614
Henderson, NV 89052, USA
Copyright © 2024 scite LLC. All rights reserved.
Made with 💙 for researchers
Part of the Research Solutions Family.