scholarly journals Evolutionary history expands the range of signaling interactions in hybrid multikinase networks

2021 ◽  
Vol 11 (1) ◽  
Author(s):  
Philippe Ortet ◽  
Sylvain Fochesato ◽  
Anne-Florence Bitbol ◽  
David E. Whitworth ◽  
David Lalaouna ◽  
...  
Keyword(s):  
Signaling Pathways ◽  
Response Regulator ◽  
Intramolecular Interactions ◽  
Domain Swap ◽  
Receiver Domain ◽  
Component Systems ◽  
Single Protein ◽  
Two Component Systems ◽  
Signaling Domains ◽  
Signaling Interactions

AbstractTwo-component systems (TCSs) are ubiquitous signaling pathways, typically comprising a sensory histidine kinase (HK) and a response regulator, which communicate via intermolecular kinase-to-receiver domain phosphotransfer. Hybrid HKs constitute non-canonical TCS signaling pathways, with transmitter and receiver domains within a single protein communicating via intramolecular phosphotransfer. Here, we report how evolutionary relationships between hybrid HKs can be used as predictors of potential intermolecular and intramolecular interactions (‘phylogenetic promiscuity’). We used domain-swap genes chimeras to investigate the specificity of phosphotransfer within hybrid HKs of the GacS–GacA multikinase network of Pseudomonas brassicacearum. The receiver domain of GacS was replaced with those from nine donor hybrid HKs. Three chimeras with receivers from other hybrid HKs demonstrated correct functioning through complementation of a gacS mutant, which was dependent on strains having a functional gacA. Formation of functional chimeras was predictable on the basis of evolutionary heritage, and raises the possibility that HKs sharing a common ancestor with GacS might remain components of the contemporary GacS network. The results also demonstrate that understanding the evolutionary heritage of signaling domains in sophisticated networks allows their rational rewiring by simple domain transplantation, with implications for the creation of designer networks and inference of functional interactions.

scholarly journals Crystallographic snapshots of DesK and DesR: two component systems on the move

2014 ◽  
Vol 70 (a1) ◽  
pp. C1050-C1050
Author(s):  
Felipe Trajtenberg ◽  
Daniela Albanesi ◽  
Horacio Botti ◽  
Ariel Mechaly ◽  
Nicole Larrieux ◽  
...  
Keyword(s):  
Response Regulator ◽  
Full Length ◽  
Activation Mechanism ◽  
Integral Model ◽  
Allosteric Control ◽  
Receiver Domain ◽  
Activated State ◽  
Component Systems ◽  
Two Component ◽  
Two Component Systems

Two-component systems (TCSs) are key players in bacterial signaling, to better understand signal-transmission with molecular detail. The TCS DesK/DesR controls fatty acid desaturation in Bacillus subtilis in response to cold shock and other membrane-altering effectors. We had previously put forward a model of signal-dependent allosteric control of the sensor kinase catalytic activity [1,2]. We have now turned our attention to the response regulator DesR. A canonical activation pathway has been widely accepted to explain phosphorylation-mediated control of response regulator function, allosterically coupling the phosphorylation site to the α4β5α5 surface. However, the structural evidence supporting the main hypotheses is still highly fragmentary. We are now reporting the crystal structure of full-length DesR, in complex with a phosphoryl-mimetic, showing the activated state [3]. Several crystal forms of the receiver domain were determined in the active and inactive configurations, revealing molecular details of the activation switch. Comparative small angle X ray scattering of full-length constructs, structure-guided point mutagenesis, as well as in vitro and in vivo functional analyses, allow us to propose an integral model of DesR activation. The phosphorylation of the receiver domain is allosterically coupled not to one, but two exposed surfaces, independently controlling its dimerization and tetramerization. Notably, a novel surface is shown to be essential for a non-canonical dimerization and activation mechanism. Direct coupling analysis highlights this interface as a shared feature of all NarL/LuxR regulators. This surface is further involved in cognate histidine kinase binding, disclosing a novel view of response regulator allosteric control. With the data we are now reporting, the DesK/DesR signaling pathway becomes, to the best of our knowledge, one of the most thoroughly studied examples of a thermosensor TCS at the molecular and biological levels.

scholarly journals Allosteric Activation of Bacterial Response Regulators: the Role of the Cognate Histidine Kinase Beyond Phosphorylation

mBio ◽  
2014 ◽  
Vol 5 (6) ◽  
Author(s):  
Felipe Trajtenberg ◽  
Daniela Albanesi ◽  
Natalia Ruétalo ◽  
Horacio Botti ◽  
Ariel E. Mechaly ◽  
...  
Keyword(s):  
Signaling Pathways ◽  
Histidine Kinase ◽  
Response Regulator ◽  
Activation Mechanism ◽  
Response Regulators ◽  
Content Type ◽  
Component Systems ◽  
Two Component ◽  
Two Component Systems

ABSTRACT Response regulators are proteins that undergo transient phosphorylation, connecting specific signals to adaptive responses. Remarkably, the molecular mechanism of response regulator activation remains elusive, largely because of the scarcity of structural data on multidomain response regulators and histidine kinase/response regulator complexes. We now address this question by using a combination of crystallographic data and functional analyses in vitro and in vivo, studying DesR and its cognate sensor kinase DesK, a two-component system that controls membrane fluidity in Bacillus subtilis. We establish that phosphorylation of the receiver domain of DesR is allosterically coupled to two distinct exposed surfaces of the protein, controlling noncanonical dimerization/tetramerization, cooperative activation, and DesK binding. One of these surfaces is critical for both homodimerization- and kinase-triggered allosteric activations. Moreover, DesK induces a phosphorylation-independent activation of DesR in vivo, uncovering a novel and stringent level of specificity among kinases and regulators. Our results support a model that helps to explain how response regulators restrict phosphorylation by small-molecule phosphoryl donors, as well as cross talk with noncognate sensors. IMPORTANCE The ability to sense and respond to environmental variations is an essential property for cell survival. Two-component systems mediate key signaling pathways that allow bacteria to integrate extra- or intracellular signals. Here we focus on the DesK/DesR system, which acts as a molecular thermometer in B. subtilis, regulating the cell membrane’s fluidity. Using a combination of complementary approaches, including determination of the crystal structures of active and inactive forms of the response regulator DesR, we unveil novel molecular mechanisms of DesR’s activation switch. In particular, we show that the association of the cognate histidine kinase DesK triggers DesR activation beyond the transfer of the phosphoryl group. On the basis of sequence and structural analyses of other two-component systems, this activation mechanism appears to be used in a wide range of sensory systems, contributing a further level of specificity control among different signaling pathways.

scholarly journals Evolutionary rewiring: a modified prokaryotic gene-regulatory pathway in chloroplasts

2013 ◽  
Vol 368 (1622) ◽  
pp. 20120260 ◽  
Author(s):  
Sujith Puthiyaveetil ◽  
Iskander M. Ibrahim ◽  
John F. Allen
Keyword(s):  
Green Algae ◽  
Gene Transcription ◽  
Response Regulator ◽  
Regulatory System ◽  
Photosynthetic Electron Transport ◽  
Sensor Kinase ◽  
Ps I ◽  
Component Systems ◽  
Two Component Systems ◽  
Gene Regulatory

Photosynthetic electron transport regulates chloroplast gene transcription through the action of a bacterial-type sensor kinase known as chloroplast sensor kinase (CSK). CSK represses photosystem I (PS I) gene transcription in PS I light and thus initiates photosystem stoichiometry adjustment. In cyanobacteria and in non-green algae, CSK homologues co-exist with their response regulator partners in canonical bacterial two-component systems. In green algae and plants, however, no response regulator partner of CSK is found. Yeast two-hybrid analysis has revealed interaction of CSK with sigma factor 1 (SIG1) of chloroplast RNA polymerase. Here we present further evidence for the interaction between CSK and SIG1. We also show that CSK interacts with quinone. Arabidopsis SIG1 becomes phosphorylated in PS I light, which then specifically represses transcription of PS I genes. In view of the identical signalling properties of CSK and SIG1 and of their interactions, we suggest that CSK is a SIG1 kinase. We propose that the selective repression of PS I genes arises from the operation of a gene-regulatory phosphoswitch in SIG1. The CSK-SIG1 system represents a novel, rewired chloroplast-signalling pathway created by evolutionary tinkering. This regulatory system supports a proposal for the selection pressure behind the evolutionary stasis of chloroplast genes.

scholarly journals Multiple Mechanisms of Action for Inhibitors of Histidine Protein Kinases from Bacterial Two-Component Systems

1999 ◽  
Vol 43 (7) ◽  
pp. 1693-1699 ◽  
Author(s):  
Jamese J. Hilliard ◽  
Raul M. Goldschmidt ◽  
Lisa Licata ◽  
Ellen Z. Baum ◽  
Karen Bush
Keyword(s):  
Pathogenic Bacteria ◽  
Response Regulator ◽  
Membrane Integrity ◽  
Bacterial Killing ◽  
Gram Positive Bacteria ◽  
Component Systems ◽  
Two Component ◽  
Two Component Systems ◽  
Bacterial Cell Membrane ◽  
Cellular Incorporation

ABSTRACT Many pathogenic bacteria utilize two-component systems consisting of a histidine protein kinase (HPK) and a response regulator (RR) for signal transduction. During the search for novel inhibitors, several chemical series, including benzoxazines, benzimidazoles, bis-phenols, cyclohexenes, trityls, and salicylanilides, were identified that inhibited the purified HPK-RR pairs KinA-Spo0F and NRII-NRI, with 50% inhibitory concentrations (IC50s) ranging from 1.9 to >500 μM and MICs ranging from 0.5 to >16 μg/ml for gram-positive bacteria. However, additional observations suggested that mechanisms other than HPK inhibition might contribute to antibacterial activity. In the present work, representative compounds from the six different series of inhibitors were analyzed for their effects on membrane integrity and macromolecular synthesis. At 4× MIC, 17 of 24 compounds compromised the integrity of the bacterial cell membrane within 10 min, as measured by uptake of propidium iodide. In this set, compounds with lower IC50s tended to cause greater membrane disruption. Eleven of 12 compounds inhibited cellular incorporation of radiolabeled thymidine and uridine >97% in 5 min and amino acids >80% in 15 min. The HPK inhibitor that allowed >25% precursor incorporation had no measurable MIC (>16 μg/ml). Fifteen of 24 compounds also caused hemolysis of equine erythrocytes. Thus, the antibacterial HPK inhibitors caused a rapid decrease in cellular incorporation of RNA, DNA, and protein precursors, possibly as a result of the concomitant disruption of the cytoplasmic membrane. Bacterial killing by these HPK inhibitors may therefore be due to multiple mechanisms, independent of HPK inhibition.

scholarly journals A survey of HK, HPt, and RR domains and their organization in two-component systems and phosphorelays proteins of organisms with fully sequenced genomes

2015 ◽  
Author(s):  
Baldiri Salvado ◽  
Ester Vilaprinyo ◽  
Albert Sorribas ◽  
Rui Alves
Keyword(s):  
Response Regulator ◽  
Signal Transmission ◽  
Pr Proteins ◽  
Component Systems ◽  
Two Component ◽  
Domains Of Life ◽  
Two Component Systems ◽  
Operon Organization ◽  
The Individual ◽  
Selection Of

Two Component Systems and Phosphorelays (TCS/PR) are environmental signal transduction cascades in prokaryotes and, less frequently, in eukaryotes. The internal domain organization of proteins and the topology of TCS/PR cascades play an important role in shaping the responses of the circuits. It is thus important to maintain updated censuses of TCS/PR proteins in order to identify the various topologies used by nature and enable a systematic study of the dynamics associated with those topologies. To create such a census, we analyzed the proteomes of 7609 organisms from all domains of life with fully sequenced and annotated genomes. To begin, we survey each proteome searching for proteins containing domains that are associated with internal signal transmission within TCS/PR: Histidine Kinase (HK), Response Regulator (RR) and Histidine Phosphotranfer (HPt) domains, and analyze how these domains are arranged in the individual proteins. Then, we find all types of operon organization and calculate how much more likely are proteins that contain TCS/PR domains to be coded by neighboring genes than one would expect from the genome background of each organism. Finally, we analyze if the fusion of domains into single TCS/PR proteins is more frequently observed than one might expect from the background of each proteome. We find 50 alternative ways in which the HK, HPt, and RR domains are observed to organize into single proteins. In prokaryotes, TCS/PR coding genes tend to be clustered in operons. 90% of all proteins identified in this study contain just one of the three domains, while 8% of the remaining proteins combine one copy of an HK, a RR, and/or an HPt domain. In eukaryotes, 25% of all TCS/PR proteins have more than one domain. These results might have implications for how signals are internally transmitted within TCS/PR cascades. These implications could explain the selection of the various designs in alternative circumstances.

scholarly journals The two-component system CopRS maintains femtomolar levels of free copper in the periplasm of Pseudomonas aeruginosa using a phosphatase-based mechanism

2020 ◽  
Author(s):  
Lorena Novoa-Aponte ◽  
Fernando C. Soncini ◽  
José M. Argüello
Keyword(s):  
Pseudomonas Aeruginosa ◽  
Phosphatase Activity ◽  
Response Regulator ◽  
Redox Enzymes ◽  
Two Component System ◽  
Component System ◽  
Component Systems ◽  
Two Component ◽  
Two Component Systems ◽  
Systems Control

ABSTRACTTwo component systems control periplasmic Cu+ homeostasis in Gram-negative bacteria. In characterized systems such as Escherichia coli CusRS, upon Cu+ binding to the periplasmic sensing domain of CusS, a cytoplasmic phosphotransfer domain phosphorylates the response regulator CusR. This drives the expression of efflux transporters, chaperones, and redox enzymes to ameliorate metal toxic effects. Here, we show that the Pseudomonas aeruginosa two component sensor histidine kinase CopS exhibits a Cu-dependent phosphatase activity that maintains a non-phosphorylated CopR when the periplasmic Cu levels are below its activation threshold. Upon Cu+ binding to the sensor, the phosphatase activity is blocked and the phosphorylated CopR activates transcription of the CopRS regulon. Supporting the model, mutagenesis experiments revealed that the ΔcopS strain showed constitutive high expression of the CopRS regulon, lower intracellular Cu+ levels, and larger Cu tolerance when compared to wild type cells. The invariant phospho-acceptor residue His235 of CopS was not required for the phosphatase activity itself, but necessary for its Cu-dependency. To sense the metal, the periplasmic domain of CopS binds two Cu+ ions at its dimeric interface. Homology modeling of CopS based on CusS structure (four Ag+ binding sites) clearly explains the different binding stoichiometries in both systems. Interestingly, CopS binds Cu+/2+ with 30 × 10−15 M affinities, pointing to the absence of free (hydrated) Cu+/2+ in the periplasm.IMPORTANCECopper is a micronutrient required as cofactor in redox enzymes. When free, copper is toxic, mismetallating proteins, and generating damaging free radicals. Consequently, copper overload is a strategy that eukaryotic cells use to combat pathogens. Bacteria have developed copper sensing transcription factors to control copper homeostasis. The cell envelope is the first compartment that has to cope with copper stress. Dedicated two component systems control the periplasmic response to metal overload. This manuscript shows that the copper sensing two component system present in Pseudomonadales exhibits a signal-dependent phosphatase activity controlling the activation of the response regulator, distinct from previously described periplasmic Cu sensors. Importantly, the data show that the sensor is activated by copper levels compatible with the absence of free copper in the cell periplasm. This emphasizes the diversity of molecular mechanisms that have evolved in various bacteria to manage the copper cellular distribution.

scholarly journals Molecular Characterization of Two-Component Systems of Helicobacter pylori

2000 ◽  
Vol 182 (8) ◽  
pp. 2068-2076 ◽  
Author(s):  
Dagmar Beier ◽  
Rainer Frank
Keyword(s):  
Helicobacter Pylori ◽  
Histidine Kinase ◽  
Response Regulator ◽  
Response Regulators ◽  
Reading Frame ◽  
Component Systems ◽  
Two Component ◽  
Two Component Systems

ABSTRACT Two-component systems are frequently involved in the adaptation of bacteria to changing environmental conditions at the level of transcriptional regulation. Here we report the characterization of members of the two-component systems of the gastric pathogenHelicobacter pylori deduced from the genome sequence of strain 26695. We demonstrate that the response regulators HP166, HP1043, and HP1021 have essential functions, as disruption of the corresponding genes is lethal for the bacteria, irrespective of the fact that HP1043 and HP1021 have nonconserved substitutions in crucial amino acids of their receiver domains. An analysis of the in vitro phosphorylation properties of the two-component proteins demonstrates that HP244-HP703 and HP165-HP166 are cognate histidine kinase-response regulator pairs. Furthermore, we provide evidence that the variability of the histidine kinase HP165 caused by a poly(C) tract of variable length close to the 3′ end of open reading frame 165/164 does not interfere with the kinase activity of the transmitter domain of HP165.

scholarly journals Unique Patterns and Biogeochemical Relevance of Two-Component Sensing in Marine Bacteria

mSystems ◽  
2019 ◽  
Vol 4 (1) ◽  
Author(s):  
Noelle A. Held ◽  
Matthew R. McIlvin ◽  
Dawn M. Moran ◽  
Michael T. Laub ◽  
Mak A. Saito
Keyword(s):  
Marine Environment ◽  
Marine Bacteria ◽  
Response Regulator ◽  
Bacterial Species ◽  
South Pacific ◽  
Response Regulators ◽  
Orphan Genes ◽  
Component Systems ◽  
Two Component ◽  
Two Component Systems

ABSTRACTTwo-component sensory (TCS) systems link microbial physiology to the environment and thus may play key roles in biogeochemical cycles. In this study, we surveyed the TCS systems of 328 diverse marine bacterial species. We identified lifestyle traits such as copiotrophy and diazotrophy that are associated with larger numbers of TCS system genes within the genome. We compared marine bacterial species with 1,152 reference bacterial species from a variety of habitats and found evidence of extra response regulators in marine genomes. Examining the location of TCS genes along the circular bacterial genome, we also found that marine bacteria have a large number of “orphan” genes, as well as many hybrid histidine kinases. The prevalence of “extra” response regulators, orphan genes, and hybrid TCS systems suggests that marine bacteria break with traditional understanding of how TCS systems operate. These trends suggest prevalent regulatory networking, which may allow coordinated physiological responses to multiple environmental signals and may represent a specific adaptation to the marine environment. We examine phylogenetic and lifestyle traits that influence the number and structure of two-component systems in the genome, finding, for example, that a lack of two-component systems is a hallmark of oligotrophy. Finally, in an effort to demonstrate the importance of TCS systems to marine biogeochemistry, we examined the distribution ofProchlorococcus/Synechococcusresponse regulator PMT9312_0717 in metaproteomes of the tropical South Pacific. We found that this protein’s abundance is related to phosphate concentrations, consistent with a putative role in phosphate regulation.IMPORTANCEMarine microbes must manage variation in their chemical, physical, and biological surroundings. Because they directly link bacterial physiology to environmental changes, TCS systems are crucial to the bacterial cell. This study surveyed TCS systems in a large number of marine bacteria and identified key phylogenetic and lifestyle patterns in environmental sensing. We found evidence that, in comparison with bacteria as a whole, marine organisms have irregular TCS system constructs which might represent an adaptation specific to the marine environment. Additionally, we demonstrate the biogeochemical relevance of TCS systems by correlating the presence of the PMT9312_0717 response regulator protein to phosphate concentrations in the South Pacific. We highlight that despite their potential ecological and biogeochemical relevance, TCS systems have been understudied in the marine ecosystem. This report expands our understanding of the breadth of bacterial TCS systems and how marine bacteria have adapted to survive in their unique environment.

scholarly journals FimY Does Not Interfere with FimZ-FimW Interaction during Type 1 Fimbria Production by Salmonella enterica Serovar Typhimurium

2013 ◽  
Vol 81 (12) ◽  
pp. 4453-4460 ◽  
Author(s):  
Sarah A. Zeiner ◽  
Brett E. Dwyer ◽  
Steven Clegg
Keyword(s):  
Salmonella Enterica ◽  
Salmonella Enterica Serovar Typhimurium ◽  
Response Regulator ◽  
Response Regulators ◽  
Content Type ◽  
Component Systems ◽  
The Family ◽  
Serovar Typhimurium ◽  
Two Component Systems

ABSTRACTThe production of type 1 fimbriae inSalmonella entericaserovar Typhimurium is controlled, in part, by three proteins, FimZ, FimY, and FimW. Amino acid sequence analysis indicates that FimZ belongs to the family of bacterial response regulators of two-component systems. In these studies, we have demonstrated that introducing a mutation mimicking phosphorylation of FimZ is necessary for activation of its target gene,fimA. In addition, the interaction of FimZ with FimW, a repressor offimAexpression, occurs only when FimZ is phosphorylated. Consequently, the negative regulatory effect of FimW is most likely due to downmodulation of the active FimZ protein. FimY does not appear to function as a response regulator, and its activity can be lost by mimicking the phosphorylation of FimY. Overproduction of FimY cannot alleviate the nonfimbriate phenotype in a FimZ mutant, whereas high levels of FimZ can overcome the nonfimbriate phenotype of a FimY mutant. It appears that FimY acts upstream of FimZ to activatefimAexpression.

scholarly journals Stochastic activation of the response regulator PhoB by noncognate histidine kinases

2005 ◽  
Vol 2 (1) ◽  
pp. 10-22 ◽  
Author(s):  
Lu Zhou ◽  
Gérald Grégori ◽  
Jennifer Masella Blackman ◽  
J. Paul Robinson ◽  
Barry L. Wanner
Keyword(s):  
Environmental Changes ◽  
Fluorescent Protein ◽  
Response Regulator ◽  
Component Systems ◽  
Epigenetic Diversity ◽  
Homogeneous Cell ◽  
Two Component Systems ◽  
Gene Regulatory ◽  
Green Fluorescent ◽  
Fluorescent Protein Reporter

Abstract Two-component systems (TCS) are the most prevalent gene regulatory mechanism in bacteria. A typical TCS is comprised of a histidine kinase (HK) and a partner response regulator (RR). Specific environment signals lead to autophosphorylation of different HKs, which in turn act as phosphoryl donors for autophosphorylation of their partner RRs. Nonpartner HKs and RRs also interact, giving rise to cross regulation among TCSs in response to diverse signals.PhoR (HK) and PhoB (RR) constitute the TCS for detection of environmental (extracellular) inorganic phosphate (Pi). The PhoR/PhoB TCS controls the expression of a large number of genes for acquisition of alternative phosphorus sources, including phoA, which encodes the non-specific phosphohydrolase bacterial alkaline phosphatase (Bap). Cross activation of PhoB by the nonpartner HK CreC is now a classic example of cross regulation among TCSs. A systematic search for other cross talking HKs revealed five additional HKs that activate (phosphorylate) PhoB (J. M. B. and B. L. W., unpublished data).Examination of cross activation of PhoB by these non-partner HKs by flow cytometry at the single-cell level revealed a bimodal, “all-or-none,” distribution pattern for expression of a phoAp-gfp (green fluorescent protein) reporter fusion. Although the basis of the observed stochastic behavior is unclear, it seems to reflect an inherent property of TCSs. We propose that cells exploit the stochastic character of TCSs to achieve nongenetic (epigenetic) diversity within genetically homogeneous cell populations in order to facilitate adaptation to environmental changes.

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