scholarly journals Novel Role for an HPt Domain in Stabilizing the Phosphorylated State of a Response Regulator Domain

2000 ◽  
Vol 182 (23) ◽  
pp. 6673-6678 ◽  
Author(s):  
Fabiola Janiak-Spens ◽  
David P. Sparling ◽  
Ann H. West
Keyword(s):  
Response Regulator ◽  
Phosphoryl Transfer ◽  
Sensor Kinase ◽  
Signaling System ◽  
Phosphorylated Protein ◽  
Physiological Conditions ◽  
Regulatory Domains ◽  
Regulatory Systems ◽  
Two Component ◽  
Stabilization Effect

ABSTRACT Two-component regulatory systems that utilize a multistep phosphorelay mechanism often involve a histidine-containing phosphotransfer (HPt) domain. These HPt domains serve an essential role as histidine-phosphorylated protein intermediates during phosphoryl transfer from one response regulator domain to another. InSaccharomyces cerevisiae, the YPD1 protein facilitates phosphoryl transfer from a hybrid sensor kinase, SLN1, to two distinct response regulator proteins, SSK1 and SKN7. Because the phosphorylation state largely determines the functional state of response regulator proteins, we have carried out a comparative study of the phosphorylated lifetimes of the three response regulator domains associated with SLN1, SSK1, and SKN7 (R1, R2, and R3, respectively). The isolated regulatory domains exhibited phosphorylated lifetimes within the range previously observed for other response regulator domains (i.e., several minutes to several hours). However, in the presence of YPD1, we found that the half-life of phosphorylated SSK1-R2 was dramatically extended (almost 200-fold longer than in the absence of YPD1). This stabilization effect was specific for SSK1-R2 and was not observed for SLN1-R1 or SKN7-R3. Our findings suggest a mechanism by which SSK1 is maintained in its phosphorylated state under normal physiological conditions and demonstrate an unprecedented regulatory role for an HPt domain in a phosphorelay signaling system.

scholarly journals Feedback Control of a Two-Component Signaling System by an Fe-S-Binding Receiver Domain

mBio ◽  
2020 ◽  
Vol 11 (2) ◽  
Author(s):  
Benjamin J. Stein ◽  
Aretha Fiebig ◽  
Sean Crosson
Keyword(s):  
Histidine Kinase ◽  
Oxygen Sensor ◽  
Cell Physiology ◽  
Phosphoryl Transfer ◽  
Environmental Cues ◽  
Sensor Kinase ◽  
Signaling System ◽  
Receiver Domain ◽  
Two Component ◽  
Feedback Inhibitor

ABSTRACT Two-component signaling systems (TCSs) function to detect environmental cues and transduce this information into a change in transcription. In its simplest form, TCS-dependent regulation of transcription entails phosphoryl-transfer from a sensory histidine kinase to its cognate DNA-binding receiver protein. However, in certain cases, auxiliary proteins may modulate TCSs in response to secondary environmental cues. Caulobacter crescentus FixT is one such auxiliary regulator. FixT is composed of a single receiver domain and functions as a feedback inhibitor of the FixL-FixJ (FixLJ) TCS, which regulates the transcription of genes involved in adaptation to microaerobiosis. We sought to define the impact of fixT on Caulobacter cell physiology and to understand the molecular mechanism by which FixT represses FixLJ signaling. fixT deletion results in excess production of porphyrins and premature entry into stationary phase, demonstrating the importance of feedback inhibition of the FixLJ signaling system. Although FixT is a receiver domain, it does not affect dephosphorylation of the oxygen sensor kinase FixL or phosphoryl-transfer from FixL to its cognate receiver FixJ. Rather, FixT represses FixLJ signaling by inhibiting the FixL autophosphorylation reaction. We have further identified a 4-cysteine motif in Caulobacter FixT that binds an Fe-S cluster and protects the protein from degradation by the Lon protease. Our data support a model in which the oxidation of this Fe-S cluster promotes the degradation of FixT in vivo. This proteolytic mechanism facilitates clearance of the FixT feedback inhibitor from the cell under normoxia and resets the FixLJ system for a future microaerobic signaling event. IMPORTANCE Two-component signal transduction systems (TCSs) are broadly conserved in the bacterial kingdom and generally contain two molecular components, a sensor histidine kinase and a receiver protein. Sensor histidine kinases alter their phosphorylation state in direct response to a physical or chemical cue, whereas receiver proteins “receive” the phosphoryl group from the kinase to regulate a change in cell physiology. We have discovered that a single-domain receiver protein, FixT, binds an Fe-S cluster and controls Caulobacter heme homeostasis though its function as a negative-feedback regulator of the oxygen sensor kinase FixL. We provide evidence that the Fe-S cluster protects FixT from Lon-dependent proteolysis in the cell and endows FixT with the ability to function as a second, autonomous oxygen/redox sensor in the FixL-FixJ signaling pathway. This study introduces a novel mechanism of regulated TCS feedback control by an Fe-S-binding receiver domain.

scholarly journals The Two-Component Regulatory System TCS08 Is Involved in Cellobiose Metabolism of Streptococcus pneumoniae R6

2006 ◽  
Vol 189 (4) ◽  
pp. 1342-1350 ◽  
Author(s):  
Stuart J. McKessar ◽  
Regine Hakenbeck
Keyword(s):  
Streptococcus Pneumoniae ◽  
Response Regulator ◽  
Regulatory System ◽  
Phosphotransferase System ◽  
Two Component System ◽  
Transcription Profiles ◽  
Regulatory Systems ◽  
Two Component ◽  
Cognate Response Regulator ◽  
Gram Positive Cocci

ABSTRACT The two-component system TCS08 is one of the regulatory systems that is important for virulence of Streptococcus pneumoniae. In order to investigate the TCS08 regulon, we have analyzed transcription profiles of mutants derived from S. pneumoniae R6 by microarray analysis. Since deletion mutants are often without a significant phenotype, we constructed a mutation in the histidine kinase HK08, T133P, in analogy to the phosphatase mutation T230P in the H box of the S. pneumoniae CiaH kinase described recently (D. Zähner, K. Kaminski, M. van der Linden, T. Mascher, M. Merai, and R. Hakenbeck, J. Mol. Microbiol. Biotechnol. 4:211-216, 2002). In addition, a deletion mutation was constructed in rr08, encoding the cognate response regulator. The most heavily suppressed genes in the hk08 mutant were spr0276 to spr0282, encoding a putative cellobiose phosphoenolpyruvate sugar phosphotransferase system (PTS). Whereas the R6 Smr parent strain and the Δrr08 mutant readily grew on cellobiose, the hk08 mutant and selected mutants with deletions in the PTS cluster did not, strongly suggesting that TCS08 is involved in the catabolism of cellobiose. Homologues of the TCS08 system were found in closely related streptococci and other gram-positive cocci. However, the genes spr0276 to spr0282, encoding the putative cellobiose PTS, represent a genomic island in S. pneumoniae and homologues were found in Streptococcus gordonii only, suggesting that this system might contribute to the pathogenicity potential of the pneumococcus.

scholarly journals The ArcB Sensor Kinase of Escherichia coli Autophosphorylates by an Intramolecular Reaction

2010 ◽  
Vol 192 (6) ◽  
pp. 1735-1739 ◽  
Author(s):  
Gabriela R. Peña-Sandoval ◽  
Dimitris Georgellis
Keyword(s):  
Response Regulator ◽  
Bond Formation ◽  
Sensor Kinase ◽  
Two Component System ◽  
Histidine Kinases ◽  
Intramolecular Reaction ◽  
Redox Active ◽  
Intermolecular Disulfide Bond ◽  
Two Component ◽  
Respiratory Conditions

ABSTRACT The Arc two-component system, comprising the ArcB sensor kinase and the ArcA response regulator, modulates the expression of numerous genes in response to the respiratory conditions of growth. ArcB is a tripartite histidine kinase whose activity is regulated by the oxidation of two cytosol-located redox-active cysteine residues that participate in intermolecular disulfide bond formation. Here we show that ArcB autophosphorylates through an intramolecular reaction which diverges from the usually envisaged intermolecular autophosphorylation of homodimeric histidine kinases.

scholarly journals Progress Overview of Bacterial Two-Component Regulatory Systems as Potential Targets for Antimicrobial Chemotherapy

Antibiotics ◽  
2020 ◽  
Vol 9 (10) ◽  
pp. 635
Author(s):  
Hidetada Hirakawa ◽  
Jun Kurushima ◽  
Yusuke Hashimoto ◽  
Haruyoshi Tomita
Keyword(s):  
Drug Resistance ◽  
Antimicrobial Chemotherapy ◽  
Response Regulator ◽  
Regulatory System ◽  
Laboratory Strain ◽  
Regulatory Systems ◽  
Two Component ◽  
Bacterial Genes ◽  
Conventional Antibiotics ◽  
External Stimuli

Bacteria adapt to changes in their environment using a mechanism known as the two-component regulatory system (TCS) (also called “two-component signal transduction system” or “two-component system”). It comprises a pair of at least two proteins, namely the sensor kinase and the response regulator. The former senses external stimuli while the latter alters the expression profile of bacterial genes for survival and adaptation. Although the first TCS was discovered and characterized in a non-pathogenic laboratory strain of Escherichia coli, it has been recognized that all bacteria, including pathogens, use this mechanism. Some TCSs are essential for cell growth and fitness, while others are associated with the induction of virulence and drug resistance/tolerance. Therefore, the TCS is proposed as a potential target for antimicrobial chemotherapy. This concept is based on the inhibition of bacterial growth with the substances acting like conventional antibiotics in some cases. Alternatively, TCS targeting may reduce the burden of bacterial virulence and drug resistance/tolerance, without causing cell death. Therefore, this approach may aid in the development of antimicrobial therapeutic strategies for refractory infections caused by multi-drug resistant (MDR) pathogens. Herein, we review the progress of TCS inhibitors based on natural and synthetic compounds.

scholarly journals A Novel Redox-Sensing Histidine Kinase That Controls Carbon Catabolite Repression inAzoarcussp. CIB

mBio ◽  
2019 ◽  
Vol 10 (2) ◽  
Author(s):  
J. Andrés Valderrama ◽  
Helena Gómez-Álvarez ◽  
Zaira Martín-Moldes ◽  
M. Álvaro Berbís ◽  
F. Javier Cañada ◽  
...  
Keyword(s):  
Aromatic Compounds ◽  
Histidine Kinase ◽  
Catabolite Repression ◽  
Carbon Catabolite Repression ◽  
Response Regulator ◽  
Sensor Kinase ◽  
General Role ◽  
Two Component ◽  
Redox Switch ◽  
Redox Sensing

ABSTRACTWe have identified and characterized the AccS multidomain sensor kinase that mediates the activation of the AccR master regulator involved in carbon catabolite repression (CCR) of the anaerobic catabolism of aromatic compounds inAzoarcussp. CIB. A truncated AccS protein that contains only the soluble C-terminal autokinase module (AccS′) accounts for the succinate-dependent CCR control. In vitroassays with purified AccS′ revealed its autophosphorylation, phosphotransfer from AccS′∼P to the Asp60 residue of AccR, and the phosphatase activity toward its phosphorylated response regulator, indicating that the equilibrium between the kinase and phosphatase activities of AccS′ may control the phosphorylation state of the AccR transcriptional regulator. Oxidized quinones, e.g., ubiquinone 0 and menadione, switched the AccS′ autokinase activity off, and three conserved Cys residues, which are not essential for catalysis, are involved in such inhibition. Thiol oxidation by quinones caused a change in the oligomeric state of the AccS′ dimer resulting in the formation of an inactive monomer. This thiol-based redox switch is tuned by the cellular energy state, which can change depending on the carbon source that the cells are using. This work expands the functional diversity of redox-sensitive sensor kinases, showing that they can control new bacterial processes such as CCR of the anaerobic catabolism of aromatic compounds. The AccSR two-component system is conserved in the genomes of some betaproteobacteria, where it might play a more general role in controlling the global metabolic state according to carbon availability.IMPORTANCETwo-component signal transduction systems comprise a sensor histidine kinase and its cognate response regulator, and some have evolved to sense and convert redox signals into regulatory outputs that allow bacteria to adapt to the altered redox environment. The work presented here expands knowledge of the functional diversity of redox-sensing kinases to control carbon catabolite repression (CCR), a phenomenon that allows the selective assimilation of a preferred compound among a mixture of several carbon sources. The newly characterized AccS sensor kinase is responsible for the phosphorylation and activation of the AccR master regulator involved in CCR of the anaerobic degradation of aromatic compounds in the betaproteobacteriumAzoarcussp. CIB. AccS seems to have a thiol-based redox switch that is modulated by the redox state of the quinone pool. The AccSR system is conserved in several betaproteobacteria, where it might play a more general role controlling their global metabolic state.

Phosphorylation and DNA binding of the regulator DcuR of the fumarate-responsive two-component system DcuSR of Escherichia coli

Microbiology ◽  
2004 ◽  
Vol 150 (4) ◽  
pp. 877-883 ◽  
Author(s):  
Ingo G. Janausch ◽  
Inma Garcia-Moreno ◽  
Daniela Lehnen ◽  
Yvonne Zeuner ◽  
Gottfried Unden
Keyword(s):  
Escherichia Coli ◽  
Binding Site ◽  
Response Regulator ◽  
Sensory System ◽  
Phosphoryl Transfer ◽  
Affinity Binding ◽  
High Affinity ◽  
Two Component ◽  
Target Promoters

The function of the response regulator DcuR of the DcuSR fumarate two-component sensory system of Escherichia coli was analysed in vitro. Isolated DcuR protein was phosphorylated by the sensory histidine kinase, DcuS, and ATP, or by acetyl phosphate. In gel retardation assays with target promoters (frdA, dcuB, dctA), phosphoryl DcuR (DcuR-P) formed a high-affinity complex, with an apparent K D (app. K D) of 0·2–0·3 μM DcuR-P, and a low-affinity (app. K D 0·8–2 μM) complex. The high-affinity complex was formed only with promoters transcriptionally-regulated by DcuSR, whereas low-affinity binding was seen also with some DcuSR-independent promoters. The binding site of DcuR-P at the dcuB promoter was determined by DNase I footprinting. One binding site of 42–52 nt (position −359 to −400/−410 nt upstream of the transcriptional start) was identified in the presence of low and high concentrations of DcuR-P. Non-phosphorylated DcuR, or DcuR-D56N mutated in the phosphoryl-accepting Asp56 residue, showed low-affinity binding to target promoters. DcuR-D56N was still able to interact with DcuS. DcuR-D56N increased the phosphorylation of DcuS and competitively inhibited phosphoryl transfer to wild-type DcuR.

scholarly journals The DNA-binding domain of the Escherichia coli CpxR two-component response regulator is constitutively active and cannot be fully attenuated by fused adjacent heterologous regulatory domains

Microbiology ◽  
2006 ◽  
Vol 152 (2) ◽  
pp. 431-441 ◽  
Author(s):  
Caroline Tapparel ◽  
Antoinette Monod ◽  
William L. Kelley
Keyword(s):  
Escherichia Coli ◽  
Dna Binding ◽  
Environmental Changes ◽  
Response Regulator ◽  
Effector Domain ◽  
Regulatory Domains ◽  
Rapid Responses ◽  
Two Component ◽  
Two Component Systems ◽  
Constitutively Active

Two-component systems (TCS) based on a sensor histidine kinase and a phosphorylated cognate target regulator allow rapid responses to environmental changes. TCS are highly evolutionarily conserved, though in only a few cases are the inducing signals understood. This study focuses on the Escherichia coli CpxR response regulator that responds to periplasmic and outer-membrane stress. N-terminal deletion mutations have been isolated that render the transcription factor constitutively active, indicating that the N terminus functions, in part, to keep the C-terminal winged-helix DNA-binding effector domain in an inactive state. Analysis of truncations spanning the CpxR interdomain region revealed that mutants retaining the α5 helix significantly augment activation. Hybrid proteins obtained by fusing the CpxR effector domain to structurally similar heterologous N-terminal regulatory domains, or even GFP, failed to restore repression to the C-terminal domain. These findings shed light on the mechanism of CpxR effector domain activation and on the investigation of constitutive mutants obtained by truncation in other TCS.

scholarly journals Identification of Two-Component Regulatory Systems in Bifidobacterium infantis by Functional Complementation and Degenerate PCR Approaches

2003 ◽  
Vol 69 (7) ◽  
pp. 4219-4226 ◽  
Author(s):  
Laura E. MacConaill ◽  
Derek Butler ◽  
Mary O'Connell-Motherway ◽  
Gerald F. Fitzgerald ◽  
Douwe van Sinderen
Keyword(s):  
Response Regulator ◽  
Northern Blotting ◽  
Probiotic Bacterium ◽  
Functional Complementation ◽  
Transcriptional Analysis ◽  
Degenerate Primers ◽  
Bifidobacterium Infantis ◽  
Promoter Sequences ◽  
Regulatory Systems ◽  
Two Component

ABSTRACT Two-component signal transduction systems (2CSs) are widely used by bacteria to sense and adapt to changing environmental conditions. With two separate approaches, three different 2CSs were identified on the chromosome of the probiotic bacterium Bifidobacterium infantis UCC 35624. One locus was identified by means of functional complementation of an Escherichia coli mutant. Another two were identified by PCR with degenerate primers corresponding to conserved regions of one protein component of the 2CS. The complete coding regions for each gene cluster were obtained, which showed that each 2CS-encoding locus specified a histidine protein kinase and an assumed cognate response regulator. Transcriptional analysis of the 2CSs by Northern blotting and primer extension identified a number of putative promoter sequences for this organism while revealing that the expression of each 2CS was growth phase dependent. Analysis of the genetic elements involved revealed significant homology with several distinct regulatory families from other high-G+C-content bacteria. The conservation of the genetic organization of these three 2CSs in other bacteria, including a number of recently published Bifidobacterium genomes, was investigated.

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