WorldWideScience

Sample records for two-component signaling proteins

  1. Features of protein-protein interactions in two-component signaling deduced from genomic libraries.

    Science.gov (United States)

    White, Robert A; Szurmant, Hendrik; Hoch, James A; Hwa, Terence

    2007-01-01

    As more and more sequence data become available, new approaches for extracting information from these data become feasible. This chapter reports on one such method that has been applied to elucidate protein-protein interactions in bacterial two-component signaling pathways. The method identifies residues involved in the interaction through an analysis of over 2500 functionally coupled proteins and a precise determination of the substitutional constraints placed on one protein by its signaling mate. Once identified, a simple log-likelihood scoring procedure is applied to these residues to build a predictive tool for assigning signaling mates. The ability to apply this method is based on a proliferation of related domains within multiple organisms. Paralogous evolution through gene duplication and divergence of two-component systems has commonly resulted in tens of closely related interacting pairs within one organism with a roughly one-to-one correspondence between signal and response. This provides us with roughly an order of magnitude more protein pairs than there are unique, fully sequenced bacterial species. Consequently, this chapter serves as both a detailed exposition of the method that has provided more depth to our knowledge of bacterial signaling and a look ahead to what would be possible on a more widespread scale, that is, to protein-protein interactions that have only one example per genome, as the number of genomes increases by a factor of 10.

  2. Histidine Phosphotransfer Proteins in Fungal Two-Component Signal Transduction Pathways

    OpenAIRE

    2013-01-01

    The histidine phosphotransfer (HPt) protein Ypd1 is an important participant in the Saccharomyces cerevisiae multistep two-component signal transduction pathway and, unlike the expanded histidine kinase gene family, is encoded by a single gene in nearly all model and pathogenic fungi. Ypd1 is essential for viability in both S. cerevisiae and in Cryptococcus neoformans. These and other aspects of Ypd1 biology, combined with the availability of structural and mutational data in S. cerevisiae, s...

  3. Genomic analysis of two-component signal transduction proteins in basidiomycetes.

    Science.gov (United States)

    Lavín, José L; Ramírez, Lucía; Ussery, David W; Pisabarro, Antonio G; Oguiza, José A

    2010-01-01

    Two-component system (TCS) proteins are components of complex signal transduction pathways in fungi, and play essential roles in the regulation of several cellular functions and responses. Species of basidiomycetes have a marked variation in their specific physiological traits, morphological complexity and lifestyles. In this study, we have used the available complete genomes of basidiomycetes to carry out a thorough identification and an extensive comparative analysis of the TCS proteins in this fungal phylum. In comparison with ascomycetes, basidiomycetes exhibit an intermediate number of TCS proteins. Several TCS proteins are highly conserved among all the basidiomycetes, and other TCS proteins appear to be specific to particular species of basidiomycetes. Moreover, some species appear to have developed a unique histidine kinase group with unusual domain architecture, the Dual-histidine kinases. The presence of differential sets of TCS proteins between basidiomycete species might reflect their adaptation to diverse environmental niches.

  4. Histidine phosphotransfer proteins in fungal two-component signal transduction pathways.

    Science.gov (United States)

    Fassler, Jan S; West, Ann H

    2013-08-01

    The histidine phosphotransfer (HPt) protein Ypd1 is an important participant in the Saccharomyces cerevisiae multistep two-component signal transduction pathway and, unlike the expanded histidine kinase gene family, is encoded by a single gene in nearly all model and pathogenic fungi. Ypd1 is essential for viability in both S. cerevisiae and in Cryptococcus neoformans. These and other aspects of Ypd1 biology, combined with the availability of structural and mutational data in S. cerevisiae, suggest that the essential interactions between Ypd1 and response regulator domains would be a good target for antifungal drug development. The goal of this minireview is to summarize the wealth of data on S. cerevisiae Ypd1 and to consider the potential benefits of conducting related studies in pathogenic fungi.

  5. Dissecting the specificity of protein-protein interaction in bacterial two-component signaling: orphans and crosstalks.

    Directory of Open Access Journals (Sweden)

    Andrea Procaccini

    Full Text Available Predictive understanding of the myriads of signal transduction pathways in a cell is an outstanding challenge of systems biology. Such pathways are primarily mediated by specific but transient protein-protein interactions, which are difficult to study experimentally. In this study, we dissect the specificity of protein-protein interactions governing two-component signaling (TCS systems ubiquitously used in bacteria. Exploiting the large number of sequenced bacterial genomes and an operon structure which packages many pairs of interacting TCS proteins together, we developed a computational approach to extract a molecular interaction code capturing the preferences of a small but critical number of directly interacting residue pairs. This code is found to reflect physical interaction mechanisms, with the strongest signal coming from charged amino acids. It is used to predict the specificity of TCS interaction: Our results compare favorably to most available experimental results, including the prediction of 7 (out of 8 known interaction partners of orphan signaling proteins in Caulobacter crescentus. Surveying among the available bacterial genomes, our results suggest 15∼25% of the TCS proteins could participate in out-of-operon "crosstalks". Additionally, we predict clusters of crosstalking candidates, expanding from the anecdotally known examples in model organisms. The tools and results presented here can be used to guide experimental studies towards a system-level understanding of two-component signaling.

  6. Genomic Analysis of Two-Component Signal Transduction Proteins in Basidiomycetes

    DEFF Research Database (Denmark)

    Ussery, David; Lavín, JL; Binnewies, Tim Terence;

    2010-01-01

    proteins. Several TCS proteins are highly conserved among all the basidiomycetes, and other TCS proteins appear to be specific to particular species of basidiomycetes. Moreover, some species appear to have developed a unique histidine kinase group with unusual domain architecture, the Dual-histidine...

  7. Differences in two-component signal transduction proteins among the genus Brucella: implications for host preference and pathogenesis

    DEFF Research Database (Denmark)

    Binnewies, Tim Terence; Ussery, David; Lavín, JL

    2010-01-01

    Two-component systems (TCSs) are the predominant bacterial signal transduction mechanisms. Species of the genus Brucella are genetically highly related and differ mainly in mammalian host adaptation and pathogenesis. In this study, TCS proteins encoded in the available genome sequences of Brucella...... species have been identified using bioinformatic methods. All the Brucella species share an identical set of TCS proteins, and the number of TCS proteins in the closely related opportunistic human pathogen Ochrobactrum anthropi was higher than in Brucella species as expected from its lifestyle. O....... anthropi lacks orthologs of the Brucella TCSs NodVW, TceSR and TcfSR, suggesting that these TCS proteins could be necessary for the adaptation of Brucella as an intracellular pathogen. This genomic analysis revealed the presence of a differential distribution of TCS pseudogenes among Brucella species...

  8. Analysis of protein expression regulated by the Helicobacter pylori ArsRS two-component signal transduction system.

    Science.gov (United States)

    Loh, John T; Gupta, Shobhana S; Friedman, David B; Krezel, Andrzej M; Cover, Timothy L

    2010-04-01

    Previous studies have shown that the Helicobacter pylori ArsRS two-component signal transduction system contributes to acid-responsive gene expression. To identify additional members of the ArsRS regulon and further investigate the regulatory role of the ArsRS system, we analyzed protein expression in wild-type and arsS null mutant strains. Numerous proteins were differentially expressed in an arsS mutant strain compared to a wild-type strain when the bacteria were cultured at pH 5.0 and also when they were cultured at pH 7.0. Genes encoding 14 of these proteins were directly regulated by the ArsRS system, based on observed binding of ArsR to the relevant promoter regions. The ArsRS-regulated proteins identified in this study contribute to acid resistance (urease and amidase), acetone metabolism (acetone carboxylase), resistance to oxidative stress (thioredoxin reductase), quorum sensing (Pfs), and several other functions. These results provide further definition of the ArsRS regulon and underscore the importance of the ArsRS system in regulating expression of H. pylori proteins during bacterial growth at both neutral pH and acidic pH.

  9. A Cyclic di-GMP-binding Adaptor Protein Interacts with Histidine Kinase to Regulate Two-component Signaling.

    Science.gov (United States)

    Xu, Linghui; Venkataramani, Prabhadevi; Ding, Yichen; Liu, Yang; Deng, Yinyue; Yong, Grace Lisi; Xin, Lingyi; Ye, Ruijuan; Zhang, Lianhui; Yang, Liang; Liang, Zhao-Xun

    2016-07-29

    The bacterial messenger cyclic di-GMP (c-di-GMP) binds to a diverse range of effectors to exert its biological effect. Despite the fact that free-standing PilZ proteins are by far the most prevalent c-di-GMP effectors known to date, their physiological function and mechanism of action remain largely unknown. Here we report that the free-standing PilZ protein PA2799 from the opportunistic pathogen Pseudomonas aeruginosa interacts directly with the hybrid histidine kinase SagS. We show that PA2799 (named as HapZ: histidine kinase associated PilZ) binds directly to the phosphoreceiver (REC) domain of SagS, and that the SagS-HapZ interaction is further enhanced at elevated c-di-GMP concentration. We demonstrate that binding of HapZ to SagS inhibits the phosphotransfer between SagS and the downstream protein HptB in a c-di-GMP-dependent manner. In accordance with the role of SagS as a motile-sessile switch and biofilm growth factor, we show that HapZ impacts surface attachment and biofilm formation most likely by regulating the expression of a large number of genes. The observations suggest a previously unknown mechanism whereby c-di-GMP mediates two-component signaling through a PilZ adaptor protein.

  10. Differences in two-component signal transduction proteins among the genus Brucella: implications for host preference and pathogenesis.

    Science.gov (United States)

    Lavín, José Luis; Binnewies, Tim T; Pisabarro, Antonio G; Ussery, David W; García-Lobo, Juan M; Oguiza, José A

    2010-08-26

    Two-component systems (TCSs) are the predominant bacterial signal transduction mechanisms. Species of the genus Brucella are genetically highly related and differ mainly in mammalian host adaptation and pathogenesis. In this study, TCS proteins encoded in the available genome sequences of Brucella species have been identified using bioinformatic methods. All the Brucella species share an identical set of TCS proteins, and the number of TCS proteins in the closely related opportunistic human pathogen Ochrobactrum anthropi was higher than in Brucella species as expected from its lifestyle. O. anthropi lacks orthologs of the Brucella TCSs NodVW, TceSR and TcfSR, suggesting that these TCS proteins could be necessary for the adaptation of Brucella as an intracellular pathogen. This genomic analysis revealed the presence of a differential distribution of TCS pseudogenes among Brucella species. Moreover, there were also differences in TCS pseudogenes between strains belonging to the same Brucella species, and in particular between B. suis biovars 1 and 2. Copyright 2010 Elsevier B.V. All rights reserved.

  11. Inhibitors targeting two-component signal transduction.

    Science.gov (United States)

    Watanabe, Takafumi; Okada, Ario; Gotoh, Yasuhiro; Utsumi, Ryutaro

    2008-01-01

    A two-component signal transduction system (TCS) is an attractive target for antibacterial agents. In this chapter, we review the TCS inhibitors developed during the past decade and introduce novel drug discovery systems to isolate the inhibitors of the YycG/YycF system, an essential TCS for bacterial growth, in an effort to develop a new class of antibacterial agents.

  12. Two Component Signal Transduction in Desulfovibrio Species

    Energy Technology Data Exchange (ETDEWEB)

    Luning, Eric; Rajeev, Lara; Ray, Jayashree; Mukhopadhyay, Aindrila

    2010-05-17

    The environmentally relevant Desulfovibrio species are sulfate-reducing bacteria that are of interest in the bioremediation of heavy metal contaminated water. Among these, the genome of D. vulgaris Hildenborough encodes a large number of two component systems consisting of 72 putative response regulators (RR) and 64 putative histidinekinases (HK), the majority of which are uncharacterized. We classified the D. vulgaris Hildenborough RRs based on their output domains and compared the distribution of RRs in other sequenced Desulfovibrio species. We have successfully purified most RRs and several HKs as His-tagged proteins. We performed phospho-transfer experiments to verify relationships between cognate pairs of HK and RR, and we have also mapped a few non-cognate HK-RR pairs. Presented here are our discoveries from the Desulfovibrio RR categorization and results from the in vitro studies using purified His tagged D. vulgaris HKs and RRs.

  13. Rewiring the specificity of two-component signal transduction systems.

    Science.gov (United States)

    Skerker, Jeffrey M; Perchuk, Barrett S; Siryaporn, Albert; Lubin, Emma A; Ashenberg, Orr; Goulian, Mark; Laub, Michael T

    2008-06-13

    Two-component signal transduction systems are the predominant means by which bacteria sense and respond to environmental stimuli. Bacteria often employ tens or hundreds of these paralogous signaling systems, comprised of histidine kinases (HKs) and their cognate response regulators (RRs). Faithful transmission of information through these signaling pathways and avoidance of detrimental crosstalk demand exquisite specificity of HK-RR interactions. To identify the determinants of two-component signaling specificity, we examined patterns of amino acid coevolution in large, multiple sequence alignments of cognate kinase-regulator pairs. Guided by these results, we demonstrate that a subset of the coevolving residues is sufficient, when mutated, to completely switch the substrate specificity of the kinase EnvZ. Our results shed light on the basis of molecular discrimination in two-component signaling pathways, provide a general approach for the rational rewiring of these pathways, and suggest that analyses of coevolution may facilitate the reprogramming of other signaling systems and protein-protein interactions.

  14. Screening for compounds that affect the interaction between bacterial two-component signal transduction response regulator protein and cognate promoter DNA.

    Science.gov (United States)

    Erickson, Matthew G; Ulijasz, Andrew T; Weisblum, Bernard

    2008-01-01

    Bacterial signal transduction systems can be used as drug targets. The signal transduction targets fall into two groups--sensor kinases and response regulators. Previously reported studies describe hits that were thought to inactivate sensor kinases but on closer examination were found to act elsewhere instead; a possible reason for this is that full-length sensor kinases are integral membrane proteins whose activity might reflect interaction with the cell membrane or with membrane components. We describe a model system that instead is based on the interaction between a test compound and a response regulator in a homogeneous phase reaction. In this system, response regulator-DNA complex formation and its inhibition by a test compound are measured by fluorescence polarization. The model system should be readily adaptable to drug discovery based on other bacterial two-component s transduction systems.

  15. Evolution of two-component signal transduction systems.

    Science.gov (United States)

    Capra, Emily J; Laub, Michael T

    2012-01-01

    To exist in a wide range of environmental niches, bacteria must sense and respond to a variety of external signals. A primary means by which this occurs is through two-component signal transduction pathways, typically composed of a sensor histidine kinase that receives the input stimuli and then phosphorylates a response regulator that effects an appropriate change in cellular physiology. Histidine kinases and response regulators have an intrinsic modularity that separates signal input, phosphotransfer, and output response; this modularity has allowed bacteria to dramatically expand and diversify their signaling capabilities. Recent work has begun to reveal the molecular basis by which two-component proteins evolve. How and why do orthologous signaling proteins diverge? How do cells gain new pathways and recognize new signals? What changes are needed to insulate a new pathway from existing pathways? What constraints are there on gene duplication and lateral gene transfer? Here, we review progress made in answering these questions, highlighting how the integration of genome sequence data with experimental studies is providing major new insights.

  16. The Evolution of Two-Component Signal Transduction Systems

    Science.gov (United States)

    Capra, Emily J.; Laub, Michael T.

    2014-01-01

    To exist in a wide range of environmental niches, bacteria must sense and respond to a myriad of external signals. A primary means by which this occurs is through two-component signal transduction pathways, typically comprised of a histidine kinase that receives the input stimuli and a response regulator that effects an appropriate change in cellular physiology. Histidine kinases and response regulators have an intrinsic modularity that separates signal input, phosphotransfer, and output response; this modularity has allowed bacteria to dramatically expand and diversify their signaling capabilities. Recent work has begun to reveal the molecular basis by which two-component proteins evolve. How and why do orthologous signaling proteins diverge? How do cells gain new pathways and recognize new signals? What changes are needed to insulate a new pathway from existing pathways? What constraints are there on gene duplication and lateral gene transfer? Here, we review progress made in answering these questions, highlighting how the integration of genome sequence data with experimental studies is providing major new insights. PMID:22746333

  17. Determinants of specificity in two-component signal transduction.

    Science.gov (United States)

    Podgornaia, Anna I; Laub, Michael T

    2013-04-01

    Maintaining the faithful flow of information through signal transduction pathways is critical to the survival and proliferation of organisms. This problem is particularly challenging as many signaling proteins are part of large, paralogous families that are highly similar at the sequence and structural levels, increasing the risk of unwanted cross-talk. To detect environmental signals and process information, bacteria rely heavily on two-component signaling systems comprised of sensor histidine kinases and their cognate response regulators. Although most species encode dozens of these signaling pathways, there is relatively little cross-talk, indicating that individual pathways are well insulated and highly specific. Here, we review the molecular mechanisms that enforce this specificity. Further, we highlight recent studies that have revealed how these mechanisms evolve to accommodate the introduction of new pathways by gene duplication. Copyright © 2013 Elsevier Ltd. All rights reserved.

  18. Molecular Mechanisms of Two-Component Signal Transduction.

    Science.gov (United States)

    Zschiedrich, Christopher P; Keidel, Victoria; Szurmant, Hendrik

    2016-09-25

    Two-component systems (TCS) comprising sensor histidine kinases and response regulator proteins are among the most important players in bacterial and archaeal signal transduction and also occur in reduced numbers in some eukaryotic organisms. Given their importance to cellular survival, virulence, and cellular development, these systems are among the most scrutinized bacterial proteins. In the recent years, a flurry of bioinformatics, genetic, biochemical, and structural studies have provided detailed insights into many molecular mechanisms that underlie the detection of signals and the generation of the appropriate response by TCS. Importantly, it has become clear that there is significant diversity in the mechanisms employed by individual systems. This review discusses the current knowledge on common themes and divergences from the paradigm of TCS signaling. An emphasis is on the information gained by a flurry of recent structural and bioinformatics studies.

  19. Receptor domains of two-component signal transduction systems.

    Science.gov (United States)

    Perry, Julie; Koteva, Kalinka; Wright, Gerard

    2011-05-01

    Two-component signal transduction systems are found ubiquitously in prokaryotes, and in archaea, fungi, yeast and some plants, where they regulate physiologic and molecular processes at both transcriptional and post-transcriptional levels. Two-component systems sense changes in environmental conditions when a specific ligand binds to the receptor domain of the histidine kinase sensory component. The structures of many histidine kinase receptors are known, including those which sense extracellular and cytoplasmic signals. In this review, we discuss the basic architecture of two-component signalling circuits, including known system ligands, structure and function of both receptor and signalling domains, the chemistry of phosphotransfer, and cross-talk between different two-component pathways. Given the importance of these systems in regulating cellular responses, many biochemical techniques have been developed for their study and analysis. We therefore also review current methods used to study two-component signalling, including a new affinity-based proteomics approach used to study inducible resistance to the antibiotic vancomycin through the VanSR two-component signal transduction system.

  20. Rewiring two-component signal transduction with small RNAs.

    Science.gov (United States)

    Göpel, Yvonne; Görke, Boris

    2012-04-01

    Bacterial two-component systems (TCSs) and small regulatory RNAs (sRNAs) form densely interconnected networks that integrate and transduce information from the environment into fine-tuned changes of gene expression. Many TCSs control target genes indirectly through regulation of sRNAs, which in turn regulate gene expression by base-pairing with mRNAs or targeting a protein. Conversely, sRNAs may control TCS synthesis, thereby recruiting the TCS regulon to other regulatory networks. Several TCSs control expression of multiple homologous sRNAs providing the regulatory networks with further flexibility. These sRNAs act redundantly, additively or hierarchically on targets. The regulatory speed of sRNAs and their unique features in gene regulation make them ideal players extending the flexibility, dynamic range or timing of TCS signaling. Copyright © 2011 Elsevier Ltd. All rights reserved.

  1. Auxiliary phosphatases in two-component signal transduction.

    Science.gov (United States)

    Silversmith, Ruth E

    2010-04-01

    Signal termination in two-component systems occurs by loss of the phosphoryl group from the response regulator protein. This review explores our current understanding of the structures, catalytic mechanisms and means of regulation of the known families of phosphatases that catalyze response regulator dephosphorylation. The CheZ and CheC/CheX/FliY families, despite different overall structures, employ identical catalytic strategies using an amide side chain to orient a water molecule for in-line attack of the aspartyl phosphate. Spo0E phosphatases contain sequence and structural features that suggest a strategy similar to the chemotaxis phosphatases but the mechanism used by the Rap phosphatases is not yet elucidated. Identification of features shared by phosphatase families may aid in the identification of currently unrecognized classes of response regulator phosphatases. Copyright 2010 Elsevier Ltd. All rights reserved.

  2. The mechanism of signal transduction by two-component systems.

    Science.gov (United States)

    Casino, Patricia; Rubio, Vicente; Marina, Alberto

    2010-12-01

    Two-component systems, composed of a homodimeric histidine kinase (HK) and a response regulator (RR), are major signal transduction devices in bacteria. Typically the signal triggers HK autophosphorylation at one His residue, followed by phosphoryl transfer from the phospho-His to an Asp residue in the RR. Signal extinction frequently involves phospho-RR dephosphorylation by a phosphatase activity of the HK. Our understanding of these reactions and of the determinants of partner specificity among HK-RR couples has been greatly increased by recent crystal structures and biochemical experiments on HK-RR complexes. Cis-autophosphorylation (one subunit phosphorylates itself) occurs in some HKs while trans-autophosphorylation takes place in others. We review and integrate this new information, discuss the mechanism of the three reactions and propose a model for transmembrane signaling by these systems. Copyright © 2010 Elsevier Ltd. All rights reserved.

  3. Crystallographic characterization of a multidomain histidine protein kinase from an essential two-component regulatory system

    OpenAIRE

    ZHAO, Haiyan; Tang, Liang

    2009-01-01

    The multidomain cytoplasmic portion of the histidine protein kinase from an essential two-component signal transduction system has been crystallized and X-ray data have been collected to 2.8 Å resolution.

  4. Two-component mediated peroxide sensing and signal transduction in fission yeast.

    Science.gov (United States)

    Quinn, Janet; Malakasi, Panagiota; Smith, Deborah A; Cheetham, Jill; Buck, Vicky; Millar, Jonathan B A; Morgan, Brian A

    2011-07-01

    Two-component related proteins play a major role in regulating the oxidative stress response in the fission yeast, Schizosaccharomyces pombe. For example, the peroxide-sensing Mak2 and Mak3 histidine kinases regulate H(2)O(2)-induced activation of the Sty1 stress-activated protein kinase pathway, and the Skn7-related response regulator transcription factor, Prr1, is essential for activation of the core oxidative stress response genes. Here, we investigate the mechanism by which the S. pombe two-component system senses H(2)O(2), and the potential role of two-component signaling in the regulation of Prr1. Significantly, we demonstrate that PAS and GAF domains present in the Mak2 histidine kinase are essential for redox-sensing and activation of Sty1. In addition, we find that Prr1 is required for the transcriptional response to a wide range of H(2)O(2) concentrations and, furthermore, that two-component regulation of Prr1 is specifically required for the response of cells to high levels of H(2)O(2). Significantly, this provides the first demonstration that the conserved two-component phosphorylation site on Skn7-related proteins influences resistance to oxidative stress and oxidative stress-induced gene expression. Collectively, these data provide new insights into the two-component mediated sensing and signaling mechanisms underlying the response of S. pombe to oxidative stress.

  5. Two-Component Signal Transduction Systems in the Cyanobacterium Synechocystis sp. PCC 6803

    Institute of Scientific and Technical Information of China (English)

    LIU Xingguo; HUANG Wei; WU Qingyu

    2006-01-01

    Two-component systems are signal transduction systems which enable bacteria to regulate cellular functions in response to changing environmental conditions. The unicellular Synechocystis sp. PCC 6803 has become a model organism for a range of biochemical and molecular biology studies aiming at investigating environmental stress response. The publication of the complete genome sequence of the cyanobacterium Synechocystis sp. PCC 6803 provided a tremendous stimulus for research in this field, and at least 80 open reading frames were identified as members of the two-component signal transduction systems in this single species of cyanobacteria. To date, functional roles have been determined for only a limited number of such proteins. This review summarizes our current knowledge about the two-component signal transduction systems in Synechocystis sp. PCC 6803 and describes recent achievements in elucidating the functional roles of these systems.

  6. Two-component systems and their co-option for eukaryotic signal transduction.

    Science.gov (United States)

    Schaller, G Eric; Shiu, Shin-Han; Armitage, Judith P

    2011-05-10

    Two-component signaling pathways involve histidine kinases, response regulators, and sometimes histidine-containing phosphotransfer proteins. Prevalent in prokaryotes, these signaling elements have also been co-opted to meet the needs of signal transduction in eukaryotes such as fungi and plants. Here we consider the evolution of such regulatory systems, with a particular emphasis on the roles they play in signaling by the plant hormones cytokinin and ethylene, in phytochrome-mediated perception of light, and as integral components of the circadian clock. Copyright © 2011 Elsevier Ltd. All rights reserved.

  7. Use of two-component signal transduction systems in the construction of synthetic genetic networks.

    Science.gov (United States)

    Ninfa, Alexander J

    2010-04-01

    Two-component signal transduction systems are a common type of signaling system in prokaryotes; the typical cell has dozens of systems regulating aspects of physiology and controlling responses to environmental conditions. In this review, I consider how these systems may be useful for engineering novel cell functions. Examples of successful incorporation of two-component systems into engineered systems are noted, and features of the systems that favor or hinder potential future use of these signaling systems for synthetic biology applications are discussed. The focus will be on the engineering of novel couplings of sensory functions to signaling outputs. Recent successes in this area are noted, such as the development of light-sensitive transmitter proteins and chemotactic receptors responsive to nitrate. Copyright 2010 Elsevier Ltd. All rights reserved.

  8. Specificity residues determine binding affinity for two-component signal transduction systems.

    Science.gov (United States)

    Willett, Jonathan W; Tiwari, Nitija; Müller, Susanne; Hummels, Katherine R; Houtman, Jon C D; Fuentes, Ernesto J; Kirby, John R

    2013-11-05

    Two-component systems (TCS) comprise histidine kinases and their cognate response regulators and allow bacteria to sense and respond to a wide variety of signals. Histidine kinases (HKs) phosphorylate and dephosphorylate their cognate response regulators (RRs) in response to stimuli. In general, these reactions appear to be highly specific and require an appropriate association between the HK and RR proteins. The Myxococcus xanthus genome encodes one of the largest repertoires of signaling proteins in bacteria (685 open reading frames [ORFs]), including at least 127 HKs and at least 143 RRs. Of these, 27 are bona fide NtrC-family response regulators, 21 of which are encoded adjacent to their predicted cognate kinases. Using system-wide profiling methods, we determined that the HK-NtrC RR pairs display a kinetic preference during both phosphotransfer and phosphatase functions, thereby defining cognate signaling systems in M. xanthus. Isothermal titration calorimetry measurements indicated that cognate HK-RR pairs interact with dissociation constants (Kd) of approximately 1 µM, while noncognate pairs had no measurable binding. Lastly, a chimera generated between the histidine kinase, CrdS, and HK1190 revealed that residues conferring phosphotransfer and phosphatase specificity dictate binding affinity, thereby establishing discrete protein-protein interactions which prevent cross talk. The data indicate that binding affinity is a critical parameter governing system-wide signaling fidelity for bacterial signal transduction proteins. Using in vitro phosphotransfer and phosphatase profiling assays and isothermal titration calorimetry, we have taken a system-wide approach to demonstrate specificity for a family of two-component signaling proteins in Myxococcus xanthus. Our results demonstrate that previously identified specificity residues dictate binding affinity and that phosphatase specificity follows phosphotransfer specificity for cognate HK-RR pairs. The data

  9. A small protein that mediates the activation of a two-component system by another two-component system

    OpenAIRE

    Kox, Linda F.F.; Wösten, Marc M. S. M.; Groisman, Eduardo A.

    2000-01-01

    The PmrA–PmrB two-component system of Salmonella enterica controls resistance to the peptide antibiotic polymyxin B and to several antimicrobial proteins from human neutrophils. Transcription of PmrA-activated genes is induced by high iron, but can also be promoted by growth in low magnesium in a process that requires another two-component system, PhoP–PhoQ. Here, we define the genetic basis for the interaction between the PhoP–PhoQ and PmrA–PmrB systems. We have identified pmrD as a PhoP-act...

  10. Two-component signal transduction system SaeRS positively regulates Staphylococcus epidermidis glucose metabolism.

    Science.gov (United States)

    Lou, Qiang; Qi, Yijun; Ma, Yuanfang; Qu, Di

    2014-01-01

    Staphylococcus epidermidis, which is a causative pathogen of nosocomial infection, expresses its virulent traits such as biofilm and autolysis regulated by two-component signal transduction system SaeRS. In this study, we performed a proteomic analysis of differences in expression between the S. epidermidis 1457 wild-type and saeRS mutant to identify candidates regulated by saeRS using two-dimensional gel electrophoresis (2-DE) combined with matrix-assisted laser desorption/lonization mass spectrometry (MALDI-TOF-MS). Of 55 identified proteins that significantly differed in expression between the two strains, 15 were upregulated and 40 were downregulated. The downregulated proteins included enzymes related to glycolysis and TCA cycle, suggesting that glucose is not properly utilized in S. epidermidis when saeRS was deleted. The study will be helpful for treatment of S. epidermidis infection from the viewpoint of metabolic modulation dependent on two-component signal transduction system SaeRS.

  11. Structural insight into partner specificity and phosphoryl transfer in two-component signal transduction.

    Science.gov (United States)

    Casino, Patricia; Rubio, Vicente; Marina, Alberto

    2009-10-16

    The chief mechanism used by bacteria for sensing their environment is based on two conserved proteins: a sensor histidine kinase (HK) and an effector response regulator (RR). The signal transduction process involves highly conserved domains of both proteins that mediate autokinase, phosphotransfer, and phosphatase activities whose output is a finely tuned RR phosphorylation level. Here, we report the structure of the complex between the entire cytoplasmic portion of Thermotoga maritima class I HK853 and its cognate, RR468, as well as the structure of the isolated RR468, both free and BeF(3)(-) bound. Our results provide insight into partner specificity in two-component systems, recognition of the phosphorylation state of each partner, and the catalytic mechanism of the phosphatase reaction. Biochemical analysis shows that the HK853-catalyzed autokinase reaction proceeds by a cis autophosphorylation mechanism within the HK subunit. The results suggest a model for the signal transduction mechanism in two-component systems.

  12. In vivo study of the two-component signaling network in Escherichia coli

    OpenAIRE

    Sommer, Erik

    2012-01-01

    Microorganisms commonly use ‘two-component’ signaling systems for sensing environmental conditions, with members being present in nearly all bacterial and archaeal genomes in different numbers. Prototypical two-component systems are comprised of a sensory histidine kinase and a response regulator protein that is phosphorylated by the kinase. The regulator typically acts as a transcription factor regulating gene expression. Due to their prevalence in microorganisms, a basic understanding of th...

  13. Using structural information to change the phosphotransfer specificity of a two-component chemotaxis signalling complex.

    Directory of Open Access Journals (Sweden)

    Christian H Bell

    2010-02-01

    Full Text Available Two-component signal transduction pathways comprising histidine protein kinases (HPKs and their response regulators (RRs are widely used to control bacterial responses to environmental challenges. Some bacteria have over 150 different two-component pathways, and the specificity of the phosphotransfer reactions within these systems is tightly controlled to prevent unwanted crosstalk. One of the best understood two-component signalling pathways is the chemotaxis pathway. Here, we present the 1.40 A crystal structure of the histidine-containing phosphotransfer domain of the chemotaxis HPK, CheA(3, in complex with its cognate RR, CheY(6. A methionine finger on CheY(6 that nestles in a hydrophobic pocket in CheA(3 was shown to be important for the interaction and was found to only occur in the cognate RRs of CheA(3, CheY(6, and CheB(2. Site-directed mutagenesis of this methionine in combination with two adjacent residues abolished binding, as shown by surface plasmon resonance studies, and phosphotransfer from CheA(3-P to CheY(6. Introduction of this methionine and an adjacent alanine residue into a range of noncognate CheYs, dramatically changed their specificity, allowing protein interaction and rapid phosphotransfer from CheA(3-P. The structure presented here has allowed us to identify specificity determinants for the CheA-CheY interaction and subsequently to successfully reengineer phosphotransfer signalling. In summary, our results provide valuable insight into how cells mediate specificity in one of the most abundant signalling pathways in biology, two-component signal transduction.

  14. Cross-talk and specificity in two-component signal transduction pathways.

    Science.gov (United States)

    Agrawal, Ruchi; Sahoo, Bikash Kumar; Saini, Deepak Kumar

    2016-05-01

    Two-component signaling systems (TCSs) are composed of two proteins, sensor kinases and response regulators, which can cross-talk and integrate information between them by virtue of high-sequence conservation and modular nature, to generate concerted and diversified responses. However, TCSs have been shown to be insulated, to facilitate linear signal transmission and response generation. Here, we discuss various mechanisms that confer specificity or cross-talk among TCSs. The presented models are supported with evidence that indicate the physiological significance of the observed TCS signaling architecture. Overall, we propose that the signaling topology of any TCSs cannot be predicted using obvious sequence or structural rules, as TCS signaling is regulated by multiple factors, including spatial and temporal distribution of the participating proteins.

  15. Phosphate sink containing two-component signaling systems as tunable threshold devices.

    Directory of Open Access Journals (Sweden)

    Munia Amin

    2014-10-01

    Full Text Available Synthetic biology aims to design de novo biological systems and reengineer existing ones. These efforts have mostly focused on transcriptional circuits, with reengineering of signaling circuits hampered by limited understanding of their systems dynamics and experimental challenges. Bacterial two-component signaling systems offer a rich diversity of sensory systems that are built around a core phosphotransfer reaction between histidine kinases and their output response regulator proteins, and thus are a good target for reengineering through synthetic biology. Here, we explore the signal-response relationship arising from a specific motif found in two-component signaling. In this motif, a single histidine kinase (HK phosphotransfers reversibly to two separate output response regulator (RR proteins. We show that, under the experimentally observed parameters from bacteria and yeast, this motif not only allows rapid signal termination, whereby one of the RRs acts as a phosphate sink towards the other RR (i.e. the output RR, but also implements a sigmoidal signal-response relationship. We identify two mathematical conditions on system parameters that are necessary for sigmoidal signal-response relationships and define key parameters that control threshold levels and sensitivity of the signal-response curve. We confirm these findings experimentally, by in vitro reconstitution of the one HK-two RR motif found in the Sinorhizobium meliloti chemotaxis pathway and measuring the resulting signal-response curve. We find that the level of sigmoidality in this system can be experimentally controlled by the presence of the sink RR, and also through an auxiliary protein that is shown to bind to the HK (yielding Hill coefficients of above 7. These findings show that the one HK-two RR motif allows bacteria and yeast to implement tunable switch-like signal processing and provides an ideal basis for developing threshold devices for synthetic biology applications.

  16. Signal integration by the two-component signal transduction response regulator CpxR.

    Science.gov (United States)

    Wolfe, Alan J; Parikh, Niyati; Lima, Bruno P; Zemaitaitis, Bozena

    2008-04-01

    The CpxAR two-component signal transduction system in Escherichia coli and other pathogens senses diverse envelope stresses and promotes the transcription of a variety of genes that remedy these stresses. An important member of the CpxAR regulon is cpxP. The CpxA-dependent transcription of cpxP has been linked to stresses such as misfolded proteins and alkaline pH. It also has been proposed that acetyl phosphate, the intermediate of the phosphotransacetylase (Pta)-acetate kinase (AckA) pathway, can activate the transcription of cpxP in a CpxA-independent manner by donating its phosphoryl group to CpxR. We tested this hypothesis by measuring the transcription of cpxP using mutants with mutations in the CpxAR pathway, mutants with mutations in the Pta-AckA pathway, and mutants with a combination of both types of mutations. From this epistasis analysis, we learned that CpxR integrates diverse stimuli. The stimuli that originate in the envelope depend on CpxA, while those associated with growth and central metabolism depend on the Pta-AckA pathway. While CpxR could receive a phosphoryl group from acetyl phosphate, this global signal was not the primary trigger for CpxR activation associated with the Pta-AckA pathway. On the strength of these results, we contend that the interactions between central metabolism and signal transduction can be quite complex and that successful investigations of such interactions must include a complete epistatic analysis.

  17. Mechanism of activation of PhoQ/PhoP two-component signal transduction by SafA, an auxiliary protein of PhoQ histidine kinase in Escherichia coli.

    Science.gov (United States)

    Ishii, Eiji; Eguchi, Yoko; Utsumi, Ryutaro

    2013-01-01

    The PhoQ/PhoP two-component signal transduction system in Escherichia coli is activated by SafA, a small membrane protein that modifies the PhoQ histidine kinase. The SafA C-terminal domain (41-65 aa) interacts directly with the sensory domain of PhoQ at the periplasm. We used in vitro and in vivo strategies to elucidate the way SafA modifies the PhoQ/PhoP phosphorelay system. First, the enzymatic activities of membranes from cells overexpressing PhoQ and cells expressing both PhoQ and SafA were compared in vitro. Increased autophosphorylation of PhoQ was observed in the presence of SafA, but it did not increase the dephosphorylation of phospho-PhoP by PhoQ. In addition, SafA increased the phospho-PhoP level on the phosphotransfer assay. We confirmed that induction of SafA results in an accumulation of phospho-PhoP in vivo by the Phos-tag system. Our results suggest that the accumulation of phospho-PhoP is linked to activation of PhoQ autophosphorylation by SafA.

  18. Two-Component Signal Transduction System SaeRS Positively Regulates Staphylococcus epidermidis Glucose Metabolism

    Directory of Open Access Journals (Sweden)

    Qiang Lou

    2014-01-01

    Full Text Available Staphylococcus epidermidis, which is a causative pathogen of nosocomial infection, expresses its virulent traits such as biofilm and autolysis regulated by two-component signal transduction system SaeRS. In this study, we performed a proteomic analysis of differences in expression between the S. epidermidis 1457 wild-type and saeRS mutant to identify candidates regulated by saeRS using two-dimensional gel electrophoresis (2-DE combined with matrix-assisted laser desorption/lonization mass spectrometry (MALDI-TOF-MS. Of 55 identified proteins that significantly differed in expression between the two strains, 15 were upregulated and 40 were downregulated. The downregulated proteins included enzymes related to glycolysis and TCA cycle, suggesting that glucose is not properly utilized in S. epidermidis when saeRS was deleted. The study will be helpful for treatment of S. epidermidis infection from the viewpoint of metabolic modulation dependent on two-component signal transduction system SaeRS.

  19. P2CS: a two-component system resource for prokaryotic signal transduction research

    Directory of Open Access Journals (Sweden)

    Méjean Vincent

    2009-07-01

    Full Text Available Abstract Background With the escalation of high throughput prokaryotic genome sequencing, there is an ever-increasing need for databases that characterise, catalogue and present data relating to particular gene sets and genomes/metagenomes. Two-component system (TCS signal transduction pathways are the dominant mechanisms by which micro-organisms sense and respond to external as well as internal environmental changes. These systems respond to a wide range of stimuli by triggering diverse physiological adjustments, including alterations in gene expression, enzymatic reactions, or protein-protein interactions. Description We present P2CS (Prokaryotic 2-Component Systems, an integrated and comprehensive database of TCS signal transduction proteins, which contains a compilation of the TCS genes within 755 completely sequenced prokaryotic genomes and 39 metagenomes. P2CS provides detailed annotation of each TCS gene including family classification, sequence features, functional domains, as well as genomic context visualization. To bypass the generic problem of gene underestimation during genome annotation, we also constituted and searched an ORFeome, which improves the recovery of TCS proteins compared to searches on the equivalent proteomes. Conclusion P2CS has been developed for computational analysis of the modular TCSs of prokaryotic genomes and metagenomes. It provides a complete overview of information on TCSs, including predicted candidate proteins and probable proteins, which need further curation/validation. The database can be browsed and queried with a user-friendly web interface at http://www.p2cs.org/.

  20. P2CS: a two-component system resource for prokaryotic signal transduction research.

    Science.gov (United States)

    Barakat, Mohamed; Ortet, Philippe; Jourlin-Castelli, Cécile; Ansaldi, Mireille; Méjean, Vincent; Whitworth, David E

    2009-07-15

    With the escalation of high throughput prokaryotic genome sequencing, there is an ever-increasing need for databases that characterise, catalogue and present data relating to particular gene sets and genomes/metagenomes. Two-component system (TCS) signal transduction pathways are the dominant mechanisms by which micro-organisms sense and respond to external as well as internal environmental changes. These systems respond to a wide range of stimuli by triggering diverse physiological adjustments, including alterations in gene expression, enzymatic reactions, or protein-protein interactions. We present P2CS (Prokaryotic 2-Component Systems), an integrated and comprehensive database of TCS signal transduction proteins, which contains a compilation of the TCS genes within 755 completely sequenced prokaryotic genomes and 39 metagenomes. P2CS provides detailed annotation of each TCS gene including family classification, sequence features, functional domains, as well as genomic context visualization. To bypass the generic problem of gene underestimation during genome annotation, we also constituted and searched an ORFeome, which improves the recovery of TCS proteins compared to searches on the equivalent proteomes. P2CS has been developed for computational analysis of the modular TCSs of prokaryotic genomes and metagenomes. It provides a complete overview of information on TCSs, including predicted candidate proteins and probable proteins, which need further curation/validation. The database can be browsed and queried with a user-friendly web interface at http://www.p2cs.org/.

  1. Diversity of two-component systems: insights into the signal transduction mechanism by the  Staphylococcus aureus two-component system GraSR.

    Science.gov (United States)

    Muzamal, Uzma; Gomez, Daniel; Kapadia, Fenika; Golemi-Kotra, Dasantila

    2014-01-01

    The response to cationic antimicrobial peptides (CAMPs) in Staphylococcus aureus relies on a two-component system (TCS), GraSR, an auxiliary protein GraX and an ATP-binding cassette (ABC) transporter, VraF/G. To understand the signal transduction mechanism by GraSR, we investigated the kinase activity of the cytoplasmic domain of histidine kinase GraS and the interaction with its cognate response regulator GraR. We also investigated interactions among the auxiliary protein GraX, GraS/R and the ATPase protein of the ABC transporter, VraF. We found that GraS lacks autophosphorylation activity, unlike a similar histidine kinase, BceS, of Bacillus subtilis. In addition, the interaction between GraS and GraR is very weak in comparison to the stronger interaction observed between BceS and its conjugated response regulator, BceR, suggesting that CAMP signaling may not flow directly from GraS to GraR. We found that the auxiliary protein GraX interacts with VraF and GraR, and requires the histidine phosphotransfer and dimerization domain of GraS to interact with this protein. Further, VraF requires the GraS region that connects the membrane-bound domain with the cytoplasmic domain of this protein for interaction with GraS. The interactions of GraX with GraS/R and VraF indicate that GraX may serve as a scaffold to bring these proteins in close proximity to GraS, plausibly to facilitate activation of GraS to ultimately transduce the signal to GraR.

  2. Role of functionality in two-component signal transduction: A stochastic study

    Science.gov (United States)

    Maity, Alok Kumar; Bandyopadhyay, Arnab; Chaudhury, Pinaki; Banik, Suman K.

    2014-03-01

    We present a stochastic formalism for signal transduction processes in a bacterial two-component system. Using elementary mass action kinetics, the proposed model takes care of signal transduction in terms of a phosphotransfer mechanism between the cognate partners of a two-component system, viz., the sensor kinase and the response regulator. Based on the difference in functionality of the sensor kinase, the noisy phosphotransfer mechanism has been studied for monofunctional and bifunctional two-component systems using the formalism of the linear noise approximation. Steady-state analysis of both models quantifies different physically realizable quantities, e.g., the variance, the Fano factor (variance/mean), and mutual information. The resultant data reveal that both systems reliably transfer information of extracellular environment under low external stimulus and in a high-kinase-and-phosphatase regime. We extend our analysis further by studying the role of the two-component system in downstream gene regulation.

  3. Evolution and phyletic distribution of two-component signal transduction systems.

    Science.gov (United States)

    Wuichet, Kristin; Cantwell, Brian J; Zhulin, Igor B

    2010-04-01

    Two-component signal transduction systems are abundant in prokaryotes. They enable cells to adjust multiple cellular functions in response to changing environmental conditions. These systems are also found, although in much smaller numbers, in lower eukaryotes and plants, where they appear to control a few very specific functions. Two-component systems have evolved in Bacteria from much simpler one-component systems bringing about the benefit of extracellular versus intracellular sensing. We review reports establishing the origins of two-component systems and documenting their occurrence in major lineages of Life. Copyright 2010 Elsevier Ltd. All rights reserved.

  4. Evolution and phyletic distribution of two-component signal transduction systems

    OpenAIRE

    Wuichet, Kristin; Cantwell, Brian J.; Zhulin, Igor B.

    2010-01-01

    Two-component signal transduction systems are abundant in prokaryotes. They enable cells to adjust multiple cellular functions in response to changing environmental conditions. These systems are also found, although in much smaller numbers, in lower eukaryotes and plants, where they appear to control a few very specific functions. Two-component systems have evolved in Bacteria from much simpler one-component systems bringing about the benefit of extracellular versus intracellular sensing. We ...

  5. Targeting two-component signal transduction: a novel drug discovery system.

    Science.gov (United States)

    Okada, Ario; Gotoh, Yasuhiro; Watanabe, Takafumi; Furuta, Eiji; Yamamoto, Kaneyoshi; Utsumi, Ryutaro

    2007-01-01

    We have developed two screening systems for isolating inhibitors that target bacterial two-component signal transduction: (1) a differential growth assay using a temperature-sensitive yycF mutant (CNM2000) of Bacillus subtilis, which is supersensitive to histidine kinase inhibitors, and (2) a high-throughput genetic system for targeting the homodimerization of histidine kinases essential for the bacterial two-component signal transduction. By using these methods, we have been able to identify various types of inhibitors that block the autophosphorylation of histidine kinases with different modes of actions.

  6. A novel "four-component" two-component signal transduction mechanism regulates developmental progression in Myxococcus xanthus.

    Science.gov (United States)

    Jagadeesan, Sakthimala; Mann, Petra; Schink, Christian W; Higgs, Penelope I

    2009-08-07

    Histidine-aspartate phosphorelays are employed by two-component signal transduction family proteins to mediate responses to specific signals or stimuli in microorganisms and plants. The RedCDEF proteins constitute a novel signaling system in which four two-component proteins comprising a histidine kinase, a histidine-kinase like protein, and two response regulators function together to regulate progression through the elaborate developmental program of Myxococcus xanthus. A combination of in vivo phenotypic analyses of in-frame deletions and non-functional point mutations in each gene as well as in vitro autophosphorylation and phosphotransfer analyses of recombinant proteins indicate that the RedC histidine kinase protein autophosphorylates and donates a phosphoryl group to the single domain response regulator, RedF, to repress progression through the developmental program. To relieve this developmental repression, RedC instead phosphorylates RedD, a dual receiver response regulator protein. Surprisingly, RedD transfers the phosphoryl group to the histidine kinase-like protein RedE, which itself appears to be incapable of autophosphorylation. Phosphorylation of RedE may render RedE accessible to RedF, where it removes the phosphoryl group from RedF-P, which is otherwise an unusually stable phosphoprotein. These analyses reveal a novel "four-component" signaling mechanism that has probably arisen to temporally coordinate signals controlling the developmental program in M. xanthus. The RedCDEF signaling system provides an important example of how the inherent plasticity and modularity of the basic two-component signaling domains comprise a highly adaptable framework well suited to expansion into complex signaling mechanisms.

  7. Connecting two-component regulatory systems by a protein that protects a response regulator from dephosphorylation by its cognate sensor

    OpenAIRE

    Kato, Akinori; Groisman, Eduardo A.

    2004-01-01

    A fundamental question in signal transduction is how an organism integrates multiple signals into a cellular response. Here we report the mechanism by which the Salmonella PmrA/PmrB two-component system responds to the signal controlling the PhoP/PhoQ two-component system. We establish that the PhoP-activated PmrD protein binds to the phosphorylated form of the response regulator PmrA, preventing both its intrinsic dephosphorylation and that promoted by its cognate sensor kinase PmrB. This re...

  8. Regulation of acid resistance by connectors of two-component signal transduction systems in Escherichia coli.

    Science.gov (United States)

    Eguchi, Yoko; Ishii, Eiji; Hata, Kensuke; Utsumi, Ryutaro

    2011-03-01

    Two-component signal transduction systems (TCSs), utilized extensively by bacteria and archaea, are involved in the rapid adaptation of the organisms to fluctuating environments. A typical TCS transduces the signal by a phosphorelay between the sensor histidine kinase and its cognate response regulator. Recently, small-sized proteins that link TCSs have been reported and are called "connectors." Their physiological roles, however, have remained elusive. SafA (sensor associating factor A) (formerly B1500), a small (65-amino-acid [65-aa]) membrane protein, is among such connectors and links Escherichia coli TCSs EvgS/EvgA and PhoQ/PhoP. Since the activation of the EvgS/EvgA system induces acid resistance, we examined whether the SafA-activated PhoQ/PhoP system is also involved in the acid resistance induced by EvgS/EvgA. Using a constitutively active evgS1 mutant for the activation of EvgS/EvgA, we found that SafA, PhoQ, and PhoP all contributed to the acid resistance phenotype. Moreover, EvgS/EvgA activation resulted in the accumulation of cellular RpoS in the exponential-phase cells in a SafA-, PhoQ-, and PhoP-dependent manner. This RpoS accumulation was caused by another connector, IraM, expression of which was induced by the activation of the PhoQ/PhoP system, thus preventing RpoS degradation by trapping response regulator RssB. Acid resistance assays demonstrated that IraM also participated in the EvgS/EvgA-induced acid resistance. Therefore, we propose a model of a signal transduction cascade proceeding from EvgS/EvgA to PhoQ/PhoP and then to RssB (connected by SafA and IraM) and discuss its contribution to the acid resistance phenotype.

  9. Functional characterization of WalRK: A two-component signal transduction system from Bacillus anthracis

    Directory of Open Access Journals (Sweden)

    Alisha Dhiman

    2014-01-01

    Full Text Available Two-component signal transduction systems (TCS, consisting of a sensor histidine protein kinase and its cognate response regulator, are an important mode of environmental sensing in bacteria. Additionally, they have been found to regulate virulence determinants in several pathogens. Bacillus anthracis, the causative agent of anthrax and a bioterrorism agent, harbours 41 pairs of TCS. However, their role in its pathogenicity has remained largely unexplored. Here, we show that WalRK of B. anthracis forms a functional TCS which exhibits some species-specific functions. Biochemical studies showed that domain variants of WalK, the histidine kinase, exhibit classical properties of autophosphorylation and phosphotransfer to its cognate response regulator WalR. Interestingly, these domain variants also show phosphatase activity towards phosphorylated WalR, thereby making WalK a bifunctional histidine kinase/phosphatase. An in silico regulon determination approach, using a consensus binding sequence from Bacillus subtilis, provided a list of 30 genes that could form a putative WalR regulon in B. anthracis. Further, electrophoretic mobility shift assay was used to show direct binding of purified WalR to the upstream regions of three putative regulon candidates, an S-layer protein EA1, a cell division ABC transporter FtsE and a sporulation histidine kinase KinB3. Our work lends insight into the species-specific functions and mode of action of B. anthracis WalRK.

  10. Functional characterization of WalRK: A two-component signal transduction system from Bacillus anthracis.

    Science.gov (United States)

    Dhiman, Alisha; Bhatnagar, Sonika; Kulshreshtha, Parul; Bhatnagar, Rakesh

    2014-01-01

    Two-component signal transduction systems (TCS), consisting of a sensor histidine protein kinase and its cognate response regulator, are an important mode of environmental sensing in bacteria. Additionally, they have been found to regulate virulence determinants in several pathogens. Bacillus anthracis, the causative agent of anthrax and a bioterrorism agent, harbours 41 pairs of TCS. However, their role in its pathogenicity has remained largely unexplored. Here, we show that WalRK of B. anthracis forms a functional TCS which exhibits some species-specific functions. Biochemical studies showed that domain variants of WalK, the histidine kinase, exhibit classical properties of autophosphorylation and phosphotransfer to its cognate response regulator WalR. Interestingly, these domain variants also show phosphatase activity towards phosphorylated WalR, thereby making WalK a bifunctional histidine kinase/phosphatase. An in silico regulon determination approach, using a consensus binding sequence from Bacillus subtilis, provided a list of 30 genes that could form a putative WalR regulon in B. anthracis. Further, electrophoretic mobility shift assay was used to show direct binding of purified WalR to the upstream regions of three putative regulon candidates, an S-layer protein EA1, a cell division ABC transporter FtsE and a sporulation histidine kinase KinB3. Our work lends insight into the species-specific functions and mode of action of B. anthracis WalRK.

  11. Two-component signal transduction pathways regulating growth and cell cycle progression in a bacterium: a system-level analysis

    National Research Council Canada - National Science Library

    Skerker, Jeffrey M; Prasol, Melanie S; Perchuk, Barrett S; Biondi, Emanuele G; Laub, Michael T

    2005-01-01

    Two-component signal transduction systems, comprised of histidine kinases and their response regulator substrates, are the predominant means by which bacteria sense and respond to extracellular signals...

  12. Comparative Genomic Analysis of Two-Component Signal Transduction Systems in Probiotic Lactobacillus casei.

    Science.gov (United States)

    Yu, Shuijing; Peng, Yanping; Chen, Wanyi; Deng, Yangwu; Guo, Yanhua

    2014-09-01

    Lactobacillus casei has traditionally been recognized as a probiotic, thus needing to survive the industrial production processes and transit through the gastrointestinal tract before providing benefit to human health. The two-component signal transduction system (TCS) plays important roles in sensing and reacting to environmental changes, which consists of a histidine kinase (HK) and a response regulator (RR). In this study we identified HKs and RRs of six sequenced L. casei strains. Ortholog analysis revealed 15 TCS clusters (HK-RR pairs), one orphan HKs and three orphan RRs, of which 12 TCS clusters were common to all six strains, three were absent in one strain. Further classification of the predicted HKs and RRs revealed interesting aspects of their putative functions. Some TCS clusters are involved with the response under the stress of the bile salts, acid, or oxidative, which contribute to survive the difficult journey through the human gastrointestinal tract. Computational predictions of 15 TCSs were verified by PCR experiments. This genomic level study of TCSs should provide valuable insights into the conservation and divergence of TCS proteins in the L. casei strains.

  13. General aspects of two-component regulatory circuits in bacteria: Domains, signals and roles.

    Science.gov (United States)

    Padilla-Vaca, Felipe; Mondragón-Jaimes, Verónica; Franco, Bernardo

    2016-08-09

    All living organisms are subject to changing environments, which must be sensed in order to respond swiftly and efficiently. Two-component systems (TCS) are signal transduction regulatory circuits based typically on a membrane bound sensor kinase and a cytoplasmic response regulator, that is activated through a histidine to aspartate phosphorelay reactions. Activated response regulator acts usually as a transcription factor. The best known examples were identified in bacteria, but they are also found in fungi, algae and plants. Thus far, they are not found in mammals. Regulatory circuits coupled to two-component systems exhibit a myriad of responses to environmental stimuli such as: redox potential, pH, specific metabolites, pressure, light and more recently to specific antimicrobial peptides that activate a sensor kinase responsible for expressing virulence factors through the active response regulator. In this review we explore general aspects on two-component systems that ultimately can play a role on virulence regulation, also the intriguing domain properties of the sensor kinases that can be a potential target for antimicrobial compounds. Only a handful of sensor kinases are extensively characterized, the vast majority belong to what we call 'the dark matter of bacterial signal transduction' since no known signal, structure and biochemical properties are available. Regulatory circuits from vertebrate pathogenic organisms can explain virulence in terms of either response to environmental factors or specific niche occupancy. Hopefully, knowledge on these signal transduction systems can lead to identify novel molecules that target two-component systems, since the increase of drug resistant microorganisms is worrisome.

  14. Imidazole as a Small Molecule Analogue in Two-Component Signal Transduction.

    Science.gov (United States)

    Page, Stephani C; Silversmith, Ruth E; Collins, Edward J; Bourret, Robert B

    2015-12-15

    In two-component signal transduction systems (TCSs), responses to stimuli are mediated through phosphotransfer between protein components. Canonical TCSs use His → Asp phosphotransfer in which phosphoryl groups are transferred from a conserved His on a sensory histidine kinase (HK) to a conserved Asp on a response regulator (RR). RRs contain the catalytic core of His → Asp phosphotransfer, evidenced by the ability of RRs to autophosphorylate with small molecule analogues of phospho-His proteins. Phosphorelays are a more complex variation of TCSs that additionally utilize Asp → His phosphotransfer through the use of an additional component, the histidine-containing phosphotransfer domain (Hpt), which reacts with RRs both as phosphodonors and phosphoacceptors. Here we show that imidazole has features of a rudimentary Hpt. Imidazole acted as a nucleophile and attacked phosphorylated RRs (RR-P) to produce monophosphoimidazole (MPI) and unphosphorylated RR. Phosphotransfer from RR-P to imidazole required the intact RR active site, indicating that the RR provided the core catalytic machinery for Asp → His phosphotransfer. Imidazole functioned in an artificial phosphorelay to transfer phosphoryl groups between unrelated RRs. The X-ray crystal structure of an activated RR·imidazole complex showed imidazole oriented in the RR active site similarly to the His of an Hpt. Imidazole interacted with RR nonconserved active site residues, which influenced the relative reactivity of RR-P with imidazole versus water. Rate constants for reaction of imidazole or MPI with chimeric RRs suggested that the RR active site contributes to the kinetic preferences exhibited by the YPD1 Hpt.

  15. Two-component signal transduction pathways regulating growth and cell cycle progression in a bacterium: a system-level analysis.

    Directory of Open Access Journals (Sweden)

    Jeffrey M Skerker

    2005-10-01

    Full Text Available Two-component signal transduction systems, comprised of histidine kinases and their response regulator substrates, are the predominant means by which bacteria sense and respond to extracellular signals. These systems allow cells to adapt to prevailing conditions by modifying cellular physiology, including initiating programs of gene expression, catalyzing reactions, or modifying protein-protein interactions. These signaling pathways have also been demonstrated to play a role in coordinating bacterial cell cycle progression and development. Here we report a system-level investigation of two-component pathways in the model organism Caulobacter crescentus. First, by a comprehensive deletion analysis we show that at least 39 of the 106 two-component genes are required for cell cycle progression, growth, or morphogenesis. These include nine genes essential for growth or viability of the organism. We then use a systematic biochemical approach, called phosphotransfer profiling, to map the connectivity of histidine kinases and response regulators. Combining these genetic and biochemical approaches, we identify a new, highly conserved essential signaling pathway from the histidine kinase CenK to the response regulator CenR, which plays a critical role in controlling cell envelope biogenesis and structure. Depletion of either cenK or cenR leads to an unusual, severe blebbing of cell envelope material, whereas constitutive activation of the pathway compromises cell envelope integrity, resulting in cell lysis and death. We propose that the CenK-CenR pathway may be a suitable target for new antibiotic development, given previous successes in targeting the bacterial cell wall. Finally, the ability of our in vitro phosphotransfer profiling method to identify signaling pathways that operate in vivo takes advantage of an observation that histidine kinases are endowed with a global kinetic preference for their cognate response regulators. We propose that this

  16. Systematic Dissection and Trajectory-Scanning Mutagenesis of the Molecular Interface That Ensures Specificity of Two-Component Signaling Pathways

    Science.gov (United States)

    Lubin, Emma A.; Ashenberg, Orr; Skerker, Jeffrey M.; Laub, Michael T.

    2010-01-01

    Two-component signal transduction systems enable bacteria to sense and respond to a wide range of environmental stimuli. Sensor histidine kinases transmit signals to their cognate response regulators via phosphorylation. The faithful transmission of information through two-component pathways and the avoidance of unwanted cross-talk require exquisite specificity of histidine kinase-response regulator interactions to ensure that cells mount the appropriate response to external signals. To identify putative specificity-determining residues, we have analyzed amino acid coevolution in two-component proteins and identified a set of residues that can be used to rationally rewire a model signaling pathway, EnvZ-OmpR. To explore how a relatively small set of residues can dictate partner selectivity, we combined alanine-scanning mutagenesis with an approach we call trajectory-scanning mutagenesis, in which all mutational intermediates between the specificity residues of EnvZ and another kinase, RstB, were systematically examined for phosphotransfer specificity. The same approach was used for the response regulators OmpR and RstA. Collectively, the results begin to reveal the molecular mechanism by which a small set of amino acids enables an individual kinase to discriminate amongst a large set of highly-related response regulators and vice versa. Our results also suggest that the mutational trajectories taken by two-component signaling proteins following gene or pathway duplication may be constrained and subject to differential selective pressures. Only some trajectories allow both the maintenance of phosphotransfer and the avoidance of unwanted cross-talk. PMID:21124821

  17. Distribution, structure and diversity of “bacterial” genes encoding two-component proteins in the Euryarchaeota

    Directory of Open Access Journals (Sweden)

    Mark K. Ashby

    2006-01-01

    Full Text Available The publicly available annotated archaeal genome sequences (23 complete and three partial annotations, October 2005 were searched for the presence of potential two-component open reading frames (ORFs using gene category lists and BLASTP. A total of 489 potential two-component genes were identified from the gene category lists and BLASTP. Two-component genes were found in 14 of the 21 Euryarchaeal sequences (October 2005 and in neither the Crenarchaeota nor the Nanoarchaeota. A total of 20 predicted protein domains were identified in the putative two-component ORFs that, in addition to the histidine kinase and receiver domains, also includes sensor and signalling domains. The detailed structure of these putative proteins is shown, as is the distribution of each class of two-component genes in each species. Potential members of orthologous groups have been identified, as have any potential operons containing two or more two-component genes. The number of two-component genes in those Euryarchaeal species which have them seems to be linked more to lifestyle and habitat than to genome complexity, with most examples being found in Methanospirillum hungatei, Haloarcula marismortui, Methanococcoides burtonii and the mesophilic Methanosarcinales group. The large numbers of two-component genes in these species may reflect a greater requirement for internal regulation. Phylogenetic analysis of orthologous groups of five different protein classes, three probably involved in regulating taxis, suggests that most of these ORFs have been inherited vertically from an ancestral Euryarchaeal species and point to a limited number of key horizontal gene transfer events.

  18. Conformational transition of response regulator RR468 in a two-component system signal transduction process.

    Science.gov (United States)

    Banerjee, Rahul; Yan, Honggao; Cukier, Robert I

    2014-05-08

    Signal transduction can be accomplished via a two-component system (TCS) consisting of a histidine kinase (HK) and a response regulator (RR). In this work, we simulate the response regulator RR468 from Thermotoga maritima, in which phosphorylation and dephosphorylation of a conserved aspartate residue acts as a switch via a large conformational change concentrated in three proximal loops. A detailed view of the conformational transition is obscured by the lack of stability of the intermediate states, which are difficult to detect using common structural biology techniques. Molecular dynamics (MD) trajectories of the inactive and active conformations were run, and show that the inactive (or active) trajectories do not exhibit sampling of the active (or inactive) conformations on this time scale. Targeted MD (TMD) was used to generate trajectories that span the inactive and active conformations and provide a view of how a localized event like phosphorylation can lead to conformational changes elsewhere in the protein, especially in the three proximal loops. The TMD trajectories are clustered to identify stages along the transition path. Residue interaction networks are identified that point to key residues having to rearrange in the process of transition. These are identified using both hydrogen bond analysis and residue interaction strength measurements. Potentials of mean force are generated for key residue rearrangements to ascertain their free energy barriers. We introduce methods that attempt to extrapolate from one conformation to the other and find that the most fluctuating proximal loop can transit part way from one to the other, suggesting that this conformational information is embedded in the sequence.

  19. Two-Component Signal Transduction Pathways Regulating Growth and Cell Cycle Progression in a Bacterium: A System-Level Analysis

    Science.gov (United States)

    Skerker, Jeffrey M; Prasol, Melanie S; Perchuk, Barrett S; Biondi, Emanuele G

    2005-01-01

    Two-component signal transduction systems, comprised of histidine kinases and their response regulator substrates, are the predominant means by which bacteria sense and respond to extracellular signals. These systems allow cells to adapt to prevailing conditions by modifying cellular physiology, including initiating programs of gene expression, catalyzing reactions, or modifying protein–protein interactions. These signaling pathways have also been demonstrated to play a role in coordinating bacterial cell cycle progression and development. Here we report a system-level investigation of two-component pathways in the model organism Caulobacter crescentus. First, by a comprehensive deletion analysis we show that at least 39 of the 106 two-component genes are required for cell cycle progression, growth, or morphogenesis. These include nine genes essential for growth or viability of the organism. We then use a systematic biochemical approach, called phosphotransfer profiling, to map the connectivity of histidine kinases and response regulators. Combining these genetic and biochemical approaches, we identify a new, highly conserved essential signaling pathway from the histidine kinase CenK to the response regulator CenR, which plays a critical role in controlling cell envelope biogenesis and structure. Depletion of either cenK or cenR leads to an unusual, severe blebbing of cell envelope material, whereas constitutive activation of the pathway compromises cell envelope integrity, resulting in cell lysis and death. We propose that the CenK–CenR pathway may be a suitable target for new antibiotic development, given previous successes in targeting the bacterial cell wall. Finally, the ability of our in vitro phosphotransfer profiling method to identify signaling pathways that operate in vivo takes advantage of an observation that histidine kinases are endowed with a global kinetic preference for their cognate response regulators. We propose that this system

  20. Two-component signal transduction in Corynebacterium glutamicum and other corynebacteria: on the way towards stimuli and targets.

    Science.gov (United States)

    Bott, Michael; Brocker, Melanie

    2012-06-01

    In bacteria, adaptation to changing environmental conditions is often mediated by two-component signal transduction systems. In the prototypical case, a specific stimulus is sensed by a membrane-bound histidine kinase and triggers autophosphorylation of a histidine residue. Subsequently, the phosphoryl group is transferred to an aspartate residue of the cognate response regulator, which then becomes active and mediates a specific response, usually by activating and/or repressing a set of target genes. In this review, we summarize the current knowledge on two-component signal transduction in Corynebacterium glutamicum. This Gram-positive soil bacterium is used for the large-scale biotechnological production of amino acids and can also be applied for the synthesis of a wide variety of other products, such as organic acids, biofuels, or proteins. Therefore, C. glutamicum has become an important model organism in industrial biotechnology and in systems biology. The type strain ATCC 13032 possesses 13 two-component systems and the role of five has been elucidated in recent years. They are involved in citrate utilization (CitAB), osmoregulation and cell wall homeostasis (MtrAB), adaptation to phosphate starvation (PhoSR), adaptation to copper stress (CopSR), and heme homeostasis (HrrSA). As C. glutamicum does not only face changing conditions in its natural environment, but also during cultivation in industrial bioreactors of up to 500 m(3) volume, adaptability can also be crucial for good performance in biotechnological production processes. Detailed knowledge on two-component signal transduction and regulatory networks therefore will contribute to both the application and the systemic understanding of C. glutamicum and related species.

  1. An intimate link: two-component signal transduction systems and metal transport systems in bacteria.

    Science.gov (United States)

    Singh, Kamna; Senadheera, Dilani B; Cvitkovitch, Dennis G

    2014-01-01

    Bacteria have evolved various strategies to contend with high concentrations of environmental heavy metal ions for rapid, adaptive responses to maintain cell viability. Evidence gathered in the past two decades suggests that bacterial two-component signal transduction systems (TCSTSs) are intimately involved in monitoring cation accumulation, and can regulate the expression of related metabolic and virulence genes to elicit adaptive responses to changes in the concentration of these ions. Using examples garnered from recent studies, we summarize the cross-regulatory relationships between metal ions and TCSTSs. We present evidence of how bacterial TCSTSs modulate metal ion homeostasis and also how metal ions, in turn, function to control the activities of these signaling systems linked with bacterial survival and virulence.

  2. Characterization of the Arc two-component signal transduction system of the capnophilic rumen bacterium Mannheimia succiniciproducens.

    Science.gov (United States)

    Jung, Won Seok; Jung, Young Ryul; Oh, Doo-Byoung; Kang, Hyun Ah; Lee, Sang Yup; Chavez-Canales, Maria; Georgellis, Dimitris; Kwon, Ohsuk

    2008-07-01

    The ArcB/A two-component signal transduction system of Escherichia coli modulates the expression of numerous operons in response to redox conditions of growth. We demonstrate that the putative arcA and arcB genes of Mannheimia succiniciproducens MBEL55E, a capnophilic (CO2-loving) rumen bacterium, encode functional proteins that specify a two-component system. The Arc proteins of the two bacterial species sufficiently resemble each other that they can participate in heterologous transphosphorylation in vitro, and the arcA and arcB genes of M. succiniciproducens confer toluidine blue resistance to E. coli arcA and arcB mutants. However, neither the quinone analogs (ubiquinone 0 and menadione) nor the cytosolic effectors (d-lactate, acetate, and pyruvate) affect the net phosphorylation of M. succiniciproducens ArcB. Our results indicate that different types of signaling molecules and distinct modes of kinase regulation are used by the ArcB proteins of E. coli and M. succiniciproducens.

  3. Negative control in two-component signal transduction by transmitter phosphatase activity.

    Science.gov (United States)

    Huynh, TuAnh Ngoc; Stewart, Valley

    2011-10-01

    Bifunctional sensor transmitter modules of two-component systems exert both positive and negative control on the receiver domain of the cognate response regulator. In negative control, the transmitter module accelerates the rate of phospho-receiver dephosphorylation. This transmitter phosphatase reaction serves the important physiological functions of resetting response regulator phosphorylation level and suppressing cross-talk. Although the biochemical reactions underlying positive control are reasonably well understood, the mechanism for transmitter phosphatase activity has been unknown. A recent hypothesis is that the transmitter phosphatase reaction is catalysed by a conserved Gln, Asn or Thr residue, via a hydrogen bond between the amide or hydroxyl group and the nucleophilic water molecule in acyl-phosphate hydrolysis. This hypothetical mechanism closely resembles the established mechanisms of auxiliary phosphatases such as CheZ and CheX, and may be widely conserved in two-component signal transduction. In addition to the proposed catalytic residues, transmitter phosphatase activity also requires the correct transmitter conformation and appropriate interactions with the receiver. Evidence suggests that the phosphatase-competent and autokinase-competent states are mutually exclusive, and the corresponding negative and positive activities are likely to be reciprocally regulated through dynamic control of transmitter conformations. © 2011 Blackwell Publishing Ltd.

  4. Evolutionary tuning of protein expression levels of a positively autoregulated two-component system.

    Directory of Open Access Journals (Sweden)

    Rong Gao

    2013-10-01

    Full Text Available Cellular adaptation relies on the development of proper regulatory schemes for accurate control of gene expression levels in response to environmental cues. Over- or under-expression can lead to diminished cell fitness due to increased costs or insufficient benefits. Positive autoregulation is a common regulatory scheme that controls protein expression levels and gives rise to essential features in diverse signaling systems, yet its roles in cell fitness are less understood. It remains largely unknown how much protein expression is 'appropriate' for optimal cell fitness under specific extracellular conditions and how the dynamic environment shapes the regulatory scheme to reach appropriate expression levels. Here, we investigate the correlation of cell fitness and output response with protein expression levels of the E. coli PhoB/PhoR two-component system (TCS. In response to phosphate (Pi-depletion, the PhoB/PhoR system activates genes involved in phosphorus assimilation as well as genes encoding themselves, similarly to many other positively autoregulated TCSs. We developed a bacteria competition assay in continuous cultures and discovered that different Pi conditions have conflicting requirements of protein expression levels for optimal cell fitness. Pi-replete conditions favored cells with low levels of PhoB/PhoR while Pi-deplete conditions selected for cells with high levels of PhoB/PhoR. These two levels matched PhoB/PhoR concentrations achieved via positive autoregulation in wild-type cells under Pi-replete and -deplete conditions, respectively. The fitness optimum correlates with the wild-type expression level, above which the phosphorylation output saturates, thus further increase in expression presumably provides no additional benefits. Laboratory evolution experiments further indicate that cells with non-ideal protein levels can evolve toward the optimal levels with diverse mutational strategies. Our results suggest that the natural

  5. Two-component signal transduction as potential drug targets in pathogenic bacteria.

    Science.gov (United States)

    Gotoh, Yasuhiro; Eguchi, Yoko; Watanabe, Takafumi; Okamoto, Sho; Doi, Akihiro; Utsumi, Ryutaro

    2010-04-01

    Gene clusters contributing to processes such as cell growth and pathogenicity are often controlled by two-component signal transduction systems (TCSs). Specific inhibitors against TCS systems work differently from conventional antibiotics, and developing them into new drugs that are effective against various drug-resistant bacteria may be possible. Furthermore, inhibitors of TCSs that control virulence factors may reduce virulence without killing the pathogenic bacteria. Previous TCS inhibitors targeting the kinase domain of the histidine kinase sensor suffered from poor selectivity. Recent TCS inhibitors, however, target the sensory domains of the sensors blocking the quorum sensing system, or target the essential response regulator. These new targets are introduced, together with several specific TCSs that have the potential to serve as effective drug targets. Copyright 2010 Elsevier Ltd. All rights reserved.

  6. Two-Component Signal Transduction Systems That Regulate the Temporal and Spatial Expression of Myxococcus xanthus Sporulation Genes.

    Science.gov (United States)

    Sarwar, Zaara; Garza, Anthony G

    2015-09-14

    When starved for nutrients, Myxococcus xanthus produces a biofilm that contains a mat of rod-shaped cells, known as peripheral rods, and aerial structures called fruiting bodies, which house thousands of dormant and stress-resistant spherical spores. Because rod-shaped cells differentiate into spherical, stress-resistant spores and spore differentiation occurs only in nascent fruiting bodies, many genes and multiple levels of regulation are required. Over the past 2 decades, many regulators of the temporal and spatial expression of M. xanthus sporulation genes have been uncovered. Of these sporulation gene regulators, two-component signal transduction circuits, which typically contain a histidine kinase sensor protein and a transcriptional regulator known as response regulator, are among the best characterized. In this review, we discuss prototypical two-component systems (Nla6S/Nla6 and Nla28S/Nla28) that regulate an early, preaggregation phase of sporulation gene expression during fruiting body development. We also discuss orphan response regulators (ActB and FruA) that regulate a later phase of sporulation gene expression, which begins during the aggregation stage of fruiting body development. In addition, we summarize the research on a complex two-component system (Esp) that is important for the spatial regulation of sporulation. Copyright © 2016, American Society for Microbiology. All Rights Reserved.

  7. Comparative Analysis of Two-component Signal Transduction System in Two Streptomycete Genomes

    Institute of Scientific and Technical Information of China (English)

    Wu WEI; Yixue LI; Weihua WANG; Zhiwei CAO; Hong YU; Xiaojing WANG; Jing ZHAO; Hao TAN; Hao XU; Weihong JIANG

    2007-01-01

    Species of the genus Streptomyces are major bacteria responsible for producing most natural antibiotics. Streptomyces coelicolor A3(2) and Streptomyces avermitilis were sequenced in 2002 and 2003,respectively. Two-component signal transduction systems (TCSs), consisting of a histidine sensor kinase (SK) and a cognate response regulator (RR), form the most common mechanism of transmembrane signal transduction in prokaryotes. TCSs in S. coelicolor A3(2) have been analyzed in detail. Here, we identify and classify the SK and RR of S. avermitilis and compare the TCSs with those of S. coelicolor A3(2) by computational approaches. Phylogenetic analysis of the cognate SK-RR pairs of the two species indicated that the cognate SK-RR pairs fall into four classes according to the distribution of their orthologs in other organisms. In addition to the cognate SK-RR pairs, some potential partners of non-cognate SK-RR were found, including those of unpaired SK and orphan RR and the cross-talk between different components in either strain. Our study provides new clues for further exploration of the molecular regulation mechanism of streptomycetes with industrial importance.

  8. The Role of Two-Component Signal Transduction Systems in Staphylococcus aureus Virulence Regulation.

    Science.gov (United States)

    Haag, Andreas F; Bagnoli, Fabio

    2016-01-05

    Staphylococcus aureus is a versatile, opportunistic human pathogen that can asymptomatically colonize a human host but can also cause a variety of cutaneous and systemic infections. The ability of S. aureus to adapt to such diverse environments is reflected in the presence of complex regulatory networks fine-tuning metabolic and virulence gene expression. One of the most widely distributed mechanisms is the two-component signal transduction system (TCS) which allows a pathogen to alter its gene expression profile in response to environmental stimuli. The simpler TCSs consist of only a transmembrane histidine kinase (HK) and a cytosolic response regulator. S. aureus encodes a total of 16 conserved pairs of TCSs that are involved in diverse signalling cascades ranging from global virulence gene regulation (e.g. quorum sensing by the Agr system), the bacterial response to antimicrobial agents, cell wall metabolism, respiration and nutrient sensing. These regulatory circuits are often interconnected and affect each other's expression, thus fine-tuning staphylococcal gene regulation. This manuscript gives an overview of the current knowledge of staphylococcal environmental sensing by TCS and its influence on virulence gene expression and virulence itself. Understanding bacterial gene regulation by TCS can give major insights into staphylococcal pathogenicity and has important implications for knowledge-based drug design and vaccine formulation.

  9. [Two-component signal transduction as attractive drug targets in pathogenic bacteria].

    Science.gov (United States)

    Utsumi, Ryutaro; Igarashi, Masayuki

    2012-01-01

    Gene clusters contributing to processes such as cell growth and pathogenicity are often controlled by two-component signal transduction systems (TCSs). TCS consists of a histidine kinase (HK) and a response regulator (RR). TCSs are attractive as drug targets for antimicrobials because many HK and RR genes are coded on the bacterial genome though few are found in lower eukaryotes. The HK/RR signal transduction system is distinct from serine/threonine and tyrosine phosphorylation in higher eukaryotes. Specific inhibitors against TCS systems work differently from conventional antibiotics, and developing them into new drugs that are effective against various drug-resistant bacteria may be possible. Furthermore, inhibitors of TCSs that control virulence factors may reduce virulence without killing the pathogenic bacteria. Previous TCS inhibitors targeting the kinase domain of the histidine kinase sensor suffered from poor selectivity. Recent TCS inhibitors, however, target the sensory domains of the sensors blocking the quorum sensing system, or target the essential response regulator. These new targets are introduced, together with several specific TCSs that have the potential to serve as effective drug targets.

  10. Phosphate sink containing two-component signaling systems as tunable threshold devices

    DEFF Research Database (Denmark)

    Amin, Munia; Kothamachu, Varun B; Feliu, Elisenda

    2014-01-01

    -component signaling systems offer a rich diversity of sensory systems that are built around a core phosphotransfer reaction between histidine kinases and their output response regulator proteins, and thus are a good target for reengineering through synthetic biology. Here, we explore the signal-response relationship...... and define key parameters that control threshold levels and sensitivity of the signal-response curve. We confirm these findings experimentally, by in vitro reconstitution of the one HK-two RR motif found in the Sinorhizobium meliloti chemotaxis pathway and measuring the resulting signal-response curve. We...... to implement tunable switch-like signal processing and provides an ideal basis for developing threshold devices for synthetic biology applications....

  11. The Two-Component Signal Transduction System VxrAB Positively Regulates Vibrio cholerae Biofilm Formation.

    Science.gov (United States)

    Teschler, Jennifer K; Cheng, Andrew T; Yildiz, Fitnat H

    2017-09-15

    Two-component signal transduction systems (TCSs), typically composed of a sensor histidine kinase (HK) and a response regulator (RR), are the primary mechanism by which pathogenic bacteria sense and respond to extracellular signals. The pathogenic bacterium Vibrio cholerae is no exception and harbors 52 RR genes. Using in-frame deletion mutants of each RR gene, we performed a systematic analysis of their role in V. cholerae biofilm formation. We determined that 7 RRs impacted the expression of an essential biofilm gene and found that the recently characterized RR, VxrB, regulates the expression of key structural and regulatory biofilm genes in V. choleraevxrB is part of a 5-gene operon, which contains the cognate HK vxrA and three genes of unknown function. Strains carrying ΔvxrA and ΔvxrB mutations are deficient in biofilm formation, while the ΔvxrC mutation enhances biofilm formation. The overexpression of VxrB led to a decrease in motility. We also observed a small but reproducible effect of the absence of VxrB on the levels of cyclic di-GMP (c-di-GMP). Our work reveals a new function for the Vxr TCS as a regulator of biofilm formation and suggests that this regulation may act through key biofilm regulators and the modulation of cellular c-di-GMP levels.IMPORTANCE Biofilms play an important role in the Vibrio cholerae life cycle, providing protection from environmental stresses and contributing to the transmission of V. cholerae to the human host. V. cholerae can utilize two-component systems (TCS), composed of a histidine kinase (HK) and a response regulator (RR), to regulate biofilm formation in response to external cues. We performed a systematic analysis of V. cholerae RRs and identified a new regulator of biofilm formation, VxrB. We demonstrated that the VxrAB TCS is essential for robust biofilm formation and that this system may regulate biofilm formation via its regulation of key biofilm regulators and cyclic di-GMP levels. This research furthers our

  12. The Nla28S/Nla28 two-component signal transduction system regulates sporulation in Myxococcus xanthus.

    Science.gov (United States)

    Sarwar, Zaara; Garza, Anthony G

    2012-09-01

    The response regulator Nla28 is a key component in a cascade of transcriptional activators that modulates expression of many important developmental genes in Myxococcus xanthus. In this study, we identified and characterized Nla28S, a histidine kinase that modulates the activity of this important regulator of M. xanthus developmental genes. We show that the putative cytoplasmic domain of Nla28S has the in vitro biochemical properties of a histidine kinase protein: it hydrolyzes ATP and undergoes an ATP-dependent autophosphorylation that is acid labile and base stable. We also show that the putative cytoplasmic domain of Nla28S transfers a phosphoryl group to Nla28 in vitro, that the phosphotransfer is specific, and that a substitution in the predicted site of Nla28 phosphorylation (aspartate 53) abolishes the phosphotransfer reaction. In phenotypic studies, we found that a mutation in nla28S produces a developmental phenotype similar to, but weaker than, that produced by a mutation in nla28; both mutations primarily affect sporulation. Together, these data indicate that Nla28S is the in vivo histidine kinase partner of Nla28 and that the primary function of the Nla28S/Nla28 two-component signal transduction system is to regulate sporulation genes. The results of genetic studies suggest that phosphorylation of Nla28S is important for the in vivo sporulation function of the Nla28S/Nla28 two-component system. In addition, the quorum signal known as A-signal is important for full developmental expression of the nla28S-nla28 operon, suggesting that quorum signaling regulates the availability of the Nla28S/Nla28 signal transduction circuit in developing cells.

  13. Stochastic simulation of prokaryotic two-component signalling indicates stochasticity-induced active-state locking and growth-rate dependent bistability.

    Science.gov (United States)

    Wei, Katy; Moinat, Maxim; Maarleveld, Timo R; Bruggeman, Frank J

    2014-07-29

    Signal transduction by prokaryotes almost exclusively relies on two-component systems for sensing and responding to (extracellular) signals. Here, we use stochastic models of two-component systems to better understand the impact of stochasticity on the fidelity and robustness of signal transmission, the outcome of autoregulatory gene expression and the influence of cell growth and division. We report that two-component systems are remarkably robust against copy number fluctuations of the signalling proteins they are composed of, which enhances signal transmission fidelity. Furthermore, we find that due to stochasticity these systems can get locked in an active state for extended time periods when (initially high) signal levels drop to zero. This behaviour can contribute to a bet-hedging adaptation strategy, aiding survival in fluctuating environments. Additionally, autoregulatory gene expression can cause two-component systems to become bistable at realistic parameter values. As a result, two sub-populations of cells can co-exist-active and inactive cells, which contributes to fitness in unpredictable environments. Bistability proved robust with respect to cell growth and division, and is tunable by the growth rate. In conclusion, our results indicate how single cells can cope with the inevitable stochasticity occurring in the activity of their two-component systems. They are robust to disadvantageous fluctuations that scramble signal transduction and they exploit beneficial stochasticity that generates fitness-enhancing heterogeneity across an isogenic population of cells.

  14. Physiological Role of Two-Component Signal Transduction Systems in Food-Associated Lactic Acid Bacteria.

    Science.gov (United States)

    Monedero, Vicente; Revilla-Guarinos, Ainhoa; Zúñiga, Manuel

    2017-01-01

    Two-component systems (TCSs) are widespread signal transduction pathways mainly found in bacteria where they play a major role in adaptation to changing environmental conditions. TCSs generally consist of sensor histidine kinases that autophosphorylate in response to a specific stimulus and subsequently transfer the phosphate group to their cognate response regulators thus modulating their activity, usually as transcriptional regulators. In this review we present the current knowledge on the physiological role of TCSs in species of the families Lactobacillaceae and Leuconostocaceae of the group of lactic acid bacteria (LAB). LAB are microorganisms of great relevance for health and food production as the group spans from starter organisms to pathogens. Whereas the role of TCSs in pathogenic LAB (most of them belonging to the family Streptococcaceae) has focused the attention, the roles of TCSs in commensal LAB, such as most species of Lactobacillaceae and Leuconostocaceae, have been somewhat neglected. However, evidence available indicates that TCSs are key players in the regulation of the physiology of these bacteria. The first studies in food-associated LAB showed the involvement of some TCSs in quorum sensing and production of bacteriocins, but subsequent studies have shown that TCSs participate in other physiological processes, such as stress response, regulation of nitrogen metabolism, regulation of malate metabolism, and resistance to antimicrobial peptides, among others. Copyright © 2017 Elsevier Inc. All rights reserved.

  15. Identification of a two-component signal transduction system that regulates maltose genes in Clostridium perfringens.

    Science.gov (United States)

    Hiscox, Thomas J; Ohtani, Kaori; Shimizu, Tohru; Cheung, Jackie K; Rood, Julian I

    2014-12-01

    Clostridium perfringens is a Gram-positive rod that is widely distributed in nature and is the etiological agent of several human and animal diseases. The complete genome sequence of C. perfringens strain 13 has been determined and multiple two-component signal transduction systems identified. One of these systems, designated here as the MalNO system, was analyzed in this study. Microarray analysis was used to carry out functional analysis of a malO mutant. The results, which were confirmed by quantitative reverse-transcriptase PCR, indicated that genes putatively involved in the uptake and metabolism of maltose were up-regulated in the malO mutant. These effects were reversed by complementation with the wild-type malO gene. Growth of these isogenic strains in medium with and without maltose showed that the malO mutant recovered more quickly from maltose deprivation when compared to the wild-type and complemented strains, leading to the conclusion that the MalNO system regulates maltose utilization in C. perfringens. It is postulated that this regulatory network may allow this soil bacterium and opportunistic pathogen to respond to environmental conditions where there are higher concentrations of maltose or maltodextrins, such as in the presence of decaying plant material in rich soil. Copyright © 2014 Elsevier Ltd. All rights reserved.

  16. Adaptation to environmental stimuli within the host: two-component signal transduction systems of Mycobacterium tuberculosis.

    Science.gov (United States)

    Bretl, Daniel J; Demetriadou, Chrystalla; Zahrt, Thomas C

    2011-12-01

    Pathogenic microorganisms encounter a variety of environmental stresses following infection of their respective hosts. Mycobacterium tuberculosis, the etiological agent of tuberculosis, is an unusual bacterial pathogen in that it is able to establish lifelong infections in individuals within granulomatous lesions that are formed following a productive immune response. Adaptation to this highly dynamic environment is thought to be mediated primarily through transcriptional reprogramming initiated in response to recognition of stimuli, including low-oxygen tension, nutrient depletion, reactive oxygen and nitrogen species, altered pH, toxic lipid moieties, cell wall/cell membrane-perturbing agents, and other environmental cues. To survive continued exposure to these potentially adverse factors, M. tuberculosis encodes a variety of regulatory factors, including 11 complete two-component signal transduction systems (TCSSs) and several orphaned response regulators (RRs) and sensor kinases (SKs). This report reviews our current knowledge of the TCSSs present in M. tuberculosis. In particular, we discuss the biochemical and functional characteristics of individual RRs and SKs, the environmental stimuli regulating their activation, the regulons controlled by the various TCSSs, and the known or postulated role(s) of individual TCSSs in the context of M. tuberculosis physiology and/or pathogenesis.

  17. STAC--A New Domain Associated with Transmembrane Solute Transport and Two-Component Signal Transduction Systems.

    Science.gov (United States)

    Korycinski, Mateusz; Albrecht, Reinhard; Ursinus, Astrid; Hartmann, Marcus D; Coles, Murray; Martin, Jörg; Dunin-Horkawicz, Stanislaw; Lupas, Andrei N

    2015-10-09

    Transmembrane receptors are integral components of sensory pathways in prokaryotes. These receptors share a common dimeric architecture, consisting in its basic form of an N-terminal extracellular sensor, transmembrane helices, and an intracellular effector. As an exception, we have identified an archaeal receptor family--exemplified by Af1503 from Archaeoglobus fulgidus--that is C-terminally shortened, lacking a recognizable effector module. Instead, a HAMP domain forms the sole extension for signal transduction in the cytosol. Here, we examine the gene environment of Af1503-like receptors and find a frequent association with transmembrane transport proteins. Furthermore, we identify and define a closely associated new protein domain family, which we characterize structurally using Af1502 from A. fulgidus. Members of this family are found both as stand-alone proteins and as domains within extant receptors. In general, the latter appear as connectors between the solute carrier 5 (SLC5)-like transmembrane domains and two-component signal transduction (TCST) domains. This is seen, for example, in the histidine kinase CbrA, which is a global regulator of metabolism, virulence, and antibiotic resistance in Pseudomonads. We propose that this newly identified domain family mediates signal transduction in systems regulating transport processes and name it STAC, for SLC and TCST-Associated Component. Copyright © 2015 MRC Laboratory of Molecular Biology. Published by Elsevier Ltd.. All rights reserved.

  18. Analysis of proteins regulated by two-component signal transduction system saeRS in Staphylococcus epidermidis%表皮葡萄球菌双组分信号转导系统saeRS对相关蛋白调控的研究

    Institute of Scientific and Technical Information of China (English)

    娄强; 王艳歌; 瞿涤

    2012-01-01

    目的 利用双向电泳技术对表皮葡萄球菌菌体蛋白进行蛋白质组学分析,寻找双组分信号转导系统saeRS的调控网络.方法 对表皮葡萄球菌1457双组分信号转导系统saeRS删除株与野生株菌体蛋白进行双向电泳差异比较;电泳图谱采用Image Master 2D Platinum软件分析;免疫印迹法验证saeRS调控的差异蛋白.结果 在表皮葡萄球菌1457双组分信号转导系统saeRS删除株与野生株蛋白质图谱中共发现23个差异表达的蛋白点(10个下调,13个上调).结论 蛋白质双向电泳技术可以成功应用于分析表皮葡萄球菌双组分信号转导系统saeRS的调控网络;此图谱为进一步研究saeRS在表皮葡萄球菌中的调控机制奠定了基础.%Objective To search for new proteins regulated by two-component signal transduction system saeRS, the proteomics of whole cellular proteins between saeRS deletion mutant and wild-type strain of Staphylococcus epidermidis ( S. epidermidis) using two-dimensional electrophoresis. Methods Immobilized pH gradient (IPG) two-dimensional electrophoresis was adopted and the gels were analyzed with Image Master 2D Platinum software. Western blot was used to analyze the differentially expressed proteins. Results We found 23 differentially expressed protein spots (10 down-regulated and 13 up-regulated) in 2-DE gels of saeRS deletion mutant and wild-type strain of S. epidermidis. Conclusion Two-dimensional electrophoresis of protein can be used to analyze the regulation network of two-component signal transduction system saeRS in S. epidermidis, and this provides basis for further study on saeRS regulation mechanism.

  19. Cloning of a two-component signal transduction system of Xanthomonas campestris pv. phaseoli var. fuscans strain BXPF65

    DEFF Research Database (Denmark)

    Chan, JWYF; Maynard, Scott; Goodwin, PH

    1998-01-01

    A putative two-component signal transduction system was amplified and cloned from the plant pathogenic bacterium Xanthomonas campestris pv. phaseoli var. fuscans isolate BXPF65. The 620 bp amplified fragment was sequenced and analyzed with the BLAST Enhanced Alignment Utility (BEAUTY). BEAUTY ana...

  20. Comparative analysis of two-component signal transduction systems of Bacillus cereus, Bacillus thuringiensis and Bacillus anthracis

    NARCIS (Netherlands)

    Been, de M.W.H.J.; Francke, C.; Moezelaar, R.; Abee, T.; Siezen, R.J.

    2006-01-01

    Members of the Bacillus cereus group are ubiquitously present in the environment and can adapt to a wide range of environmental fluctuations. In bacteria, these adaptive responses are generally mediated by two-component signal transduction systems (TCSs), which consist of a histidine kinase (HK) and

  1. Comparative analysis of two-component signal transduction systems of Bacillus cereus, Bacillus thuringiensis and Bacillus anthracis

    NARCIS (Netherlands)

    Been, M.W.H.J. de; Francke, C.; Moezelaar, R.; Abee, T.; Siezen, R.J.

    2006-01-01

    Members of the Bacillus cereus group are ubiquitously present in the environment and can adapt to a wide range of environmental fluctuations. In bacteria, these adaptive responses are generally mediated by two-component signal transduction systems (TCSs), which consist of a histidine kinase (HK) and

  2. A hybrid two-component system protein from Azospirillum brasilense Sp7 was involved in chemotaxis.

    Science.gov (United States)

    Cui, Yanhua; Tu, Ran; Wu, Lixian; Hong, Yuanyuan; Chen, Sanfeng

    2011-09-20

    We here report the sequence and functional analysis of org35 of Azospirillum brasilense Sp7, which was originally identified to be able to interact with NifA in yeast-two-hybrid system. The org35 encodes a hybrid two-component system protein, including N-terminal PAS domains, a histidine kinase (HPK) domain and a response regulator (RR) domain in C-terminal. To determine the function of the Org35, a deletion-insertion mutant in PAS domain [named Sp7353] and a complemental strain Sp7353C were constructed. The mutant had reduced chemotaxis ability compared to that of wild-type, and the complemental strain was similar to the wild-type strain. These data suggested that the A. brasilense org35 played a key role in chemotaxis. Variants containing different domains of the org35 were expressed, and the functions of these domains were studied in vitro. Phosphorylation assays in vitro demonstrated that the HPK domain of Org35 possessed the autokinase activity and that the phosphorylated HPK was able to transfer phosphate groups to the RR domain. The result indicated Org35 was a phosphorylation-communicating protein.

  3. Positive autoregulation shapes response timing and intensity in two-component signal transduction systems.

    Science.gov (United States)

    Mitrophanov, Alexander Y; Hadley, Tricia J; Groisman, Eduardo A

    2010-08-27

    Positive feedback loops are regulatory elements that can modulate expression output, kinetics and noise in genetic circuits. Transcriptional regulators participating in such loops are often expressed from two promoters, one constitutive and one autoregulated. Here, we investigate the interplay of promoter strengths and the intensity of the stimulus activating the transcriptional regulator in defining the output of a positively autoregulated genetic circuit. Using a mathematical model of two-component regulatory systems, which are present in all domains of life, we establish that positive feedback strongly affects the steady-state output levels at both low and high levels of stimulus if the constitutive promoter of the regulator is weak. By contrast, the effect of positive feedback is negligible when the constitutive promoter is sufficiently strong, unless the stimulus intensity is very high. Furthermore, we determine that positive feedback can affect both transient and steady state output levels even in the simplest genetic regulatory systems. We tested our modeling predictions by abolishing the positive feedback loop in the two-component regulatory system PhoP/PhoQ of Salmonella enterica, which resulted in diminished induction of PhoP-activated genes. Copyright (c) 2010 Elsevier Ltd. All rights reserved.

  4. The oligomeric assembly of the novel haem-degrading protein HbpS is essential for interaction with its cognate two-component sensor kinase

    NARCIS (Netherlands)

    Ortiz de Orué Lucana, Darío; Bogel, Gabriele; Zou, Peijian; Groves, Matthew R

    2009-01-01

    HbpS, a novel protein of previously unknown function from Streptomyces reticuli, is up-regulated in response to haemin- and peroxide-based oxidative stress and interacts with the SenS/SenR two-component signal transduction system. In this study, we report the high-resolution crystal structures (2.2

  5. The oligomeric assembly of the novel haem-degrading protein HbpS is essential for interaction with its cognate two-component sensor kinase

    NARCIS (Netherlands)

    Ortiz de Orué Lucana, Darío; Bogel, Gabriele; Zou, Peijian; Groves, Matthew R

    2009-01-01

    HbpS, a novel protein of previously unknown function from Streptomyces reticuli, is up-regulated in response to haemin- and peroxide-based oxidative stress and interacts with the SenS/SenR two-component signal transduction system. In this study, we report the high-resolution crystal structures (2.2

  6. Purification and assays of Rhodobacter capsulatus RegB-RegA two-component signal transduction system.

    Science.gov (United States)

    Swem, Lee R; Swem, Danielle L; Wu, Jiang; Bauer, Carl E

    2007-01-01

    Two-component signal-transduction systems, composed of a histidine-sensor kinase and a DNA-binding response regulator, allow bacteria to detect environmental changes and adjust cellular physiology to live more efficiently in a broad distribution of niches. Although many two-component signal-transduction systems are known, a limited number of signals that stimulate these systems have been discovered. This chapter describes the purification and characterization of the predominant two-component signal-transduction system utilized by Rhodobacter capsulatus, a nonsulfur purple photosynthetic bacterium. Specifically, we explain the overexpression, detergent solubilization, and purification of the full-length membrane-spanning histidine-sensor kinase RegB. We also provide a method to measure autophosphorylation of RegB and discern the effect of its signal molecule, ubiquinone, on autophosphorylation levels. In addition we describe the overexpression and purification of the cognate response regulator RegA and a technique used to visualize the phosphotransfer reaction from RegB to RegA.

  7. The Physarum polycephalum Genome Reveals Extensive Use of Prokaryotic Two-Component and Metazoan-Type Tyrosine Kinase Signaling.

    Science.gov (United States)

    Schaap, Pauline; Barrantes, Israel; Minx, Pat; Sasaki, Narie; Anderson, Roger W; Bénard, Marianne; Biggar, Kyle K; Buchler, Nicolas E; Bundschuh, Ralf; Chen, Xiao; Fronick, Catrina; Fulton, Lucinda; Golderer, Georg; Jahn, Niels; Knoop, Volker; Landweber, Laura F; Maric, Chrystelle; Miller, Dennis; Noegel, Angelika A; Peace, Rob; Pierron, Gérard; Sasaki, Taeko; Schallenberg-Rüdinger, Mareike; Schleicher, Michael; Singh, Reema; Spaller, Thomas; Storey, Kenneth B; Suzuki, Takamasa; Tomlinson, Chad; Tyson, John J; Warren, Wesley C; Werner, Ernst R; Werner-Felmayer, Gabriele; Wilson, Richard K; Winckler, Thomas; Gott, Jonatha M; Glöckner, Gernot; Marwan, Wolfgang

    2015-11-27

    Physarum polycephalum is a well-studied microbial eukaryote with unique experimental attributes relative to other experimental model organisms. It has a sophisticated life cycle with several distinct stages including amoebal, flagellated, and plasmodial cells. It is unusual in switching between open and closed mitosis according to specific life-cycle stages. Here we present the analysis of the genome of this enigmatic and important model organism and compare it with closely related species. The genome is littered with simple and complex repeats and the coding regions are frequently interrupted by introns with a mean size of 100 bases. Complemented with extensive transcriptome data, we define approximately 31,000 gene loci, providing unexpected insights into early eukaryote evolution. We describe extensive use of histidine kinase-based two-component systems and tyrosine kinase signaling, the presence of bacterial and plant type photoreceptors (phytochromes, cryptochrome, and phototropin) and of plant-type pentatricopeptide repeat proteins, as well as metabolic pathways, and a cell cycle control system typically found in more complex eukaryotes. Our analysis characterizes P. polycephalum as a prototypical eukaryote with features attributed to the last common ancestor of Amorphea, that is, the Amoebozoa and Opisthokonts. Specifically, the presence of tyrosine kinases in Acanthamoeba and Physarum as representatives of two distantly related subdivisions of Amoebozoa argues against the later emergence of tyrosine kinase signaling in the opisthokont lineage and also against the acquisition by horizontal gene transfer. © The Author(s) 2015. Published by Oxford University Press on behalf of the Society for Molecular Biology and Evolution.

  8. Influence of two-component signal transduction systems of Lactobacillus casei BL23 on tolerance to stress conditions.

    Science.gov (United States)

    Alcántara, Cristina; Revilla-Guarinos, Ainhoa; Zúñiga, Manuel

    2011-02-01

    Lactobacillus casei BL23 carries 17 two-component signal transduction systems. Insertional mutations were introduced into each gene encoding the cognate response regulators, and their effects on growth under different conditions were assayed. Inactivation of systems TC01, TC06, and TC12 (LCABL_02080-LCABL_02090, LCABL_12050-LCABL_12060, and LCABL_19600-LCABL_19610, respectively) led to major growth defects under the conditions assayed.

  9. Analysis of the virulence-associated RevSR two-component signal transduction system of Clostridium perfringens.

    Science.gov (United States)

    Cheung, Jackie K; Wisniewski, Jessica A; Adams, Vicki M; Quinsey, Noelene S; Rood, Julian I

    2016-09-01

    Clostridium perfringens is a Gram-positive, anaerobic, spore-forming bacterium that causes human gas gangrene (clostridial myonecrosis) and food poisoning. Early studies showed that virulence was regulated by the VirSR two-component signal transduction system. However, our identification of the RevR orphan response regulator indicated that more than one system was involved in controlling virulence. To further characterize this virulence-associated regulator, gel mobility shift experiments, coupled with DNase I footprinting, were used to identify the RevR DNA binding sequence. Bioinformatics analysis suggested that an orphan sensor histidine kinase, CPE1757 (renamed RevS), was the cognate sensor of RevR. Interaction between RevS and RevR was demonstrated by use of a bacterial two-hybrid system and validated by protein-protein interaction studies using biolayer interferometry. To assess the involvement of RevS in virulence regulation, the revS gene was inactivated by Targetron insertion. When isogenic wild-type, revS and complemented revS strains were tested in a mouse myonecrosis model, the revS mutant was found to be attenuated in virulence, which was similar to the attenuation observed previously with the revR mutant. However, transcriptional analysis of selected RevR-regulated genes in the revS mutant revealed a different pattern of expression to a revR mutant, suggesting that the RevSR system is more complex than originally thought. Taken together, the results have led to the identification and characterization of the two essential parts of a new regulatory network that is involved in the regulation of virulence in C. perfringens.

  10. The periplasmic regulator ExoR inhibits ExoS/ChvI two-component signaling in Sinorhizobium meliloti

    OpenAIRE

    Chen, Esther J.; Sabio, Erich A.; Long, Sharon R

    2008-01-01

    Sinorhizobium meliloti requires ExoS/ChvI two-component signaling to establish a nitrogen-fixing symbiosis with legume hosts. The importance of ExoS/ChvI signaling in microbe-host interactions is underscored by the requirement of ExoS/ChvI orthologs for virulence of the related α-proteobacteria Agrobacterium tumefaciens and Brucella abortus. In S. meliloti, ExoS/ChvI is a key regulator of gene expression for exopolysaccharide synthesis, biofilm formation, motility, nutrient utilization, and f...

  11. Dopamine reward prediction-error signalling: a two-component response

    Science.gov (United States)

    Schultz, Wolfram

    2017-01-01

    Environmental stimuli and objects, including rewards, are often processed sequentially in the brain. Recent work suggests that the phasic dopamine reward prediction-error response follows a similar sequential pattern. An initial brief, unselective and highly sensitive increase in activity unspecifically detects a wide range of environmental stimuli, then quickly evolves into the main response component, which reflects subjective reward value and utility. This temporal evolution allows the dopamine reward prediction-error signal to optimally combine speed and accuracy. PMID:26865020

  12. A subfamily of putative cytokinin receptors is revealed by an analysis of the evolution of the two-component signaling system of plants.

    Science.gov (United States)

    Gruhn, Nijuscha; Halawa, Mhyeddeen; Snel, Berend; Seidl, Michael F; Heyl, Alexander

    2014-05-01

    The two-component signaling system--the major signaling pathway of bacteria--is found among higher eukaryotes only in plants, where it regulates diverse processes, such as the signaling of the phytohormone cytokinin. Cytokinin is perceived by a hybrid histidine (His) kinase receptor, and the signal is transduced by a multistep phosphorelay system of His phosphotransfer proteins and different classes of response regulators (RRs). To shed light on the origin and evolution of the two-component signaling system members in plants, we conducted a comprehensive domain-based phylogenetic study across the relevant kingdoms, including Charophyceae algae, the group of green algae giving rise to land plants. Surprisingly, we identified a subfamily of cytokinin receptors with members only from the early diverging land plants Marchantia polymorpha and Physcomitrella patens and then experimentally characterized two members of this subfamily. His phosphotransfer proteins of Charophyceae seemed to be more closely related to land plants than to other groups of green algae. Farther down the signaling pathway, the type-B RRs were found across all plant clades, but many members lack either the canonical Asp residue or the DNA binding domain. In contrast, the type-A RRs seemed to be limited to land plants. Finally, the analysis provided hints that one additional group of RRs, the type-C RRs, might be degenerated receptors and thus, of a different evolutionary origin than bona fide RRs.

  13. Identification of DNA binding motifs of the Mycobacterium tuberculosis PhoP/PhoR two-component signal transduction system.

    Science.gov (United States)

    Cimino, Mena; Thomas, Christophe; Namouchi, Amine; Dubrac, Sarah; Gicquel, Brigitte; Gopaul, Deshmukh N

    2012-01-01

    The Mycobacterium tuberculosis PhoP/PhoR two-component signal transduction system controls the expression of about 2% of the genome and plays a major role in pathogenicity. However, its regulon has not been well characterized. The binding site of PhoP transcription regulator was identified in the upstream regions of msl3, pks2, lipF and fadD21 genes, by using gene fusions, electrophoretic mobility shift assays and DNase I footprinting experiments. A consensus sequence for PhoP binding was deduced. It consists of two direct repeats, DR1/DR2, associated with a third repeat, DR3, important in some cases for PhoP binding to DR1/DR2 but located at a variable distance from these direct repeats. DR1/DR2 and DR3 consensus sequences were used to screen the whole-genome sequence for other putative binding sites potentially corresponding to genes directly regulated by PhoP. The identified 87 genes, encoding transcription regulators, and proteins involved in secondary metabolites biosynthesis, transport and catabolism are proposed to belong to the PhoP regulon. A consensus sequence derived from the analysis of PhoP binding to four gene promoter regions is proposed. We show for the first time the involvement of a third direct repeat motif in this binding reaction. The consensus sequence was instrumented to study the global regulation mediated by PhoP in M. tuberculosis. This analysis leads to the identification of several genes that are potentially regulated by this key player.

  14. Stochastic simulation of prokaryotic two-component signalling indicates stochasticity-induced active-state locking and growth-rate dependent bistability

    NARCIS (Netherlands)

    K. Wei (Katy); M. Moinat (Maxim); T.R. Maarleveld (Timo); F.J. Bruggeman (Frank)

    2014-01-01

    htmlabstractSignal transduction by prokaryotes almost exclusively relies on two-component systems for sensing and responding to (extracellular) signals. Here, we use stochastic models of two-component systems to better understand the impact of stochasticity on the fidelity and robustness of signal

  15. Stochastic kinetic model of two component system signalling reveals all-or-none, graded and mixed mode stochastic switching responses.

    Science.gov (United States)

    Kierzek, Andrzej M; Zhou, Lu; Wanner, Barry L

    2010-03-01

    Two-component systems (TCSs) are prevalent signal transduction systems in bacteria that control innumerable adaptive responses to environmental cues and host-pathogen interactions. We constructed a detailed stochastic kinetic model of two component signalling based on published data. Our model has been validated with flow cytometry data and used to examine reporter gene expression in response to extracellular signal strength. The model shows that, depending on the actual kinetic parameters, TCSs exhibit all-or-none, graded or mixed mode responses. In accordance with other studies, positively autoregulated TCSs exhibit all-or-none responses. Unexpectedly, our model revealed that TCSs lacking a positive feedback loop exhibit not only graded but also mixed mode responses, in which variation of the signal strength alters the level of gene expression in induced cells while the regulated gene continues to be expressed at the basal level in a substantial fraction of cells. The graded response of the TCS changes to mixed mode response by an increase of the translation initiation rate of the histidine kinase. Thus, a TCS is an evolvable design pattern capable of implementing deterministic regulation and stochastic switches associated with both graded and threshold responses. This has implications for understanding the emergence of population diversity in pathogenic bacteria and the design of genetic circuits in synthetic biology applications. The model is available in systems biology markup language (SBML) and systems biology graphical notation (SBGN) formats and can be used as a component of large-scale biochemical reaction network models.

  16. Structure and mechanism of the essential two-component signal-transduction system WalKR in Staphylococcus aureus.

    Science.gov (United States)

    Ji, Quanjiang; Chen, Peter J; Qin, Guangrong; Deng, Xin; Hao, Ziyang; Wawrzak, Zdzislaw; Yeo, Won-Sik; Quang, Jenny Winjing; Cho, Hoonsik; Luo, Guan-Zheng; Weng, Xiaocheng; You, Qiancheng; Luan, Chi-Hao; Yang, Xiaojing; Bae, Taeok; Yu, Kunqian; Jiang, Hualiang; He, Chuan

    2016-03-18

    Most low GC Gram-positive bacteria possess an essential walKR two-component system (TCS) for signal transduction involved in regulating cell wall homoeostasis. Despite the well-established intracellular regulatory mechanism, the role of this TCS in extracellular signal recognition and factors that modulate the activity of this TCS remain largely unknown. Here we identify the extracellular receptor of the kinase 'WalK' (erWalK) as a key hub for bridging extracellular signal input and intracellular kinase activity modulation in Staphylococcus aureus. Characterization of the crystal structure of erWalK revealed a canonical Per-Arnt-Sim (PAS) domain for signal sensing. Single amino-acid mutation of potential signal-transduction residues resulted in severely impaired function of WalKR. A small molecule derived from structure-based virtual screening against erWalK is capable of selectively activating the walKR TCS. The molecular level characterization of erWalK will not only facilitate exploration of natural signal(s) but also provide a template for rational design of erWalK inhibitors.

  17. An Overview of Two-Component Signal Transduction Systems Implicated in Extra-Intestinal Pathogenic E. coli Infections

    Directory of Open Access Journals (Sweden)

    Erin J. Breland

    2017-05-01

    Full Text Available Extra-intestinal pathogenic E. coli (ExPEC infections are common in mammals and birds. The predominant ExPEC types are avian pathogenic E. coli (APEC, neonatal meningitis causing E. coli/meningitis associated E. coli (NMEC/MAEC, and uropathogenic E. coli (UPEC. Many reviews have described current knowledge on ExPEC infection strategies and virulence factors, especially for UPEC. However, surprisingly little has been reported on the regulatory modules that have been identified as critical in ExPEC pathogenesis. Two-component systems (TCSs comprise the predominant method by which bacteria respond to changing environments and play significant roles in modulating bacterial fitness in diverse niches. Recent studies have highlighted the potential of manipulating signal transduction systems as a means to chemically re-wire bacterial pathogens, thereby reducing selective pressure and avoiding the emergence of antibiotic resistance. This review begins by providing a brief introduction to characterized infection strategies and common virulence factors among APEC, NMEC, and UPEC and continues with a comprehensive overview of two-component signal transduction networks that have been shown to influence ExPEC pathogenesis.

  18. An Overview of Two-Component Signal Transduction Systems Implicated in Extra-Intestinal Pathogenic E. coli Infections.

    Science.gov (United States)

    Breland, Erin J; Eberly, Allison R; Hadjifrangiskou, Maria

    2017-01-01

    Extra-intestinal pathogenic E. coli (ExPEC) infections are common in mammals and birds. The predominant ExPEC types are avian pathogenic E. coli (APEC), neonatal meningitis causing E. coli/meningitis associated E. coli (NMEC/MAEC), and uropathogenic E. coli (UPEC). Many reviews have described current knowledge on ExPEC infection strategies and virulence factors, especially for UPEC. However, surprisingly little has been reported on the regulatory modules that have been identified as critical in ExPEC pathogenesis. Two-component systems (TCSs) comprise the predominant method by which bacteria respond to changing environments and play significant roles in modulating bacterial fitness in diverse niches. Recent studies have highlighted the potential of manipulating signal transduction systems as a means to chemically re-wire bacterial pathogens, thereby reducing selective pressure and avoiding the emergence of antibiotic resistance. This review begins by providing a brief introduction to characterized infection strategies and common virulence factors among APEC, NMEC, and UPEC and continues with a comprehensive overview of two-component signal transduction networks that have been shown to influence ExPEC pathogenesis.

  19. Two-Component Signal Transduction Systems of Desulfovibrio Vulgaris: Structural and Phylogenetic Analysis and Deduction of Putative Cognate Pairs

    Energy Technology Data Exchange (ETDEWEB)

    Zhang, Weiwen; Culley, David E.; Wu, Gang; Brockman, Fred J.

    2006-01-20

    ABSTRACT-Two-component signal transduction systems (TCSTS) composed of sensory histidine kinases (HK) and response regulators (RR), constitute a key element of the mechanism by which bacteria sense and respond to changes in environments. A large number of TCSTSs including 59 putative HKs and 55 RRs were identified from the Desulfovibrio vulgaris genome, indicating their important roles in regulation of cellular metabolism. In this study, the structural and phylogenetic analysis of all putative TCSTSs in D. vulgaris was performed. The results showed D. vulgaris contained an unexpectedly large number of hybrid-type HKs, implying that multiple-step phosphorelay may be a common signal transduction mechanism in D. vulgaris. Most TCSTS components of D. vulgaris were found clustered into several subfamilies previously recognized in other bacteria and extensive co-evolution between D. vulgaris HKs and RRs was observed, suggesting that the concordance of HKs and RRs in cognate phylogenetic groups could be indicative of cognate TCSTSs...

  20. Identification of a second two-component signal transduction system that controls fosfomycin tolerance and glycerol-3-phosphate uptake.

    Science.gov (United States)

    Kurabayashi, Kumiko; Hirakawa, Yuko; Tanimoto, Koichi; Tomita, Haruyoshi; Hirakawa, Hidetada

    2015-03-01

    Particular interest in fosfomycin has resurfaced because it is a highly beneficial antibiotic for the treatment of refractory infectious diseases caused by pathogens that are resistant to other commonly used antibiotics. The biological cost to cells of resistance to fosfomycin because of chromosomal mutation is high. We previously found that a bacterial two-component system, CpxAR, induces fosfomycin tolerance in enterohemorrhagic Escherichia coli (EHEC) O157:H7. This mechanism does not rely on irreversible genetic modification and allows EHEC to relieve the fitness burden that results from fosfomycin resistance in the absence of fosfomycin. Here we show that another two-component system, TorSRT, which was originally characterized as a regulatory system for anaerobic respiration utilizing trimethylamine-N-oxide (TMAO), also induces fosfomycin tolerance. Activation of the Tor regulatory pathway by overexpression of torR, which encodes the response regulator, or addition of TMAO increased fosfomycin tolerance in EHEC. We also show that phosphorylated TorR directly represses the expression of glpT, a gene that encodes a symporter of fosfomycin and glycerol-3-phosphate, and activation of the TorR protein results in the reduced uptake of fosfomycin by cells. However, cells in which the Tor pathway was activated had an impaired growth phenotype when cultured with glycerol-3-phosphate as a carbon substrate. These observations suggest that the TorSRT pathway is the second two-component system to reversibly control fosfomycin tolerance and glycerol-3-phosphate uptake in EHEC, and this may be beneficial for bacteria by alleviating the biological cost. We expect that this mechanism could be a potential target to enhance the utility of fosfomycin as chemotherapy against multidrug-resistant pathogens. Copyright © 2015, American Society for Microbiology. All Rights Reserved.

  1. Comparative genomic analysis of two-component regulatory proteins in Pseudomonas syringae

    DEFF Research Database (Denmark)

    Lavin, J.L.; Kiil, Kristoffer; Resano, O.

    2007-01-01

    important differences in TCS proteins among the three P. syringae pathovars. Conclusion: In this article we present a thorough analysis of the identification and distribution of TCS proteins among the sequenced genomes of P. syringae. We have identified differences in TCS proteins among the three P...... requires a complex array of TCS proteins to cope with diverse plant hosts, host responses, and environmental conditions. Results: Based on the genomic data, pattern searches with Hidden Markov Model (HMM) profiles have been used to identify putative HKs and RRs. The genomes of Psy B728a, Pto DC3000 and Pph...... 1448A were found to contain a large number of genes encoding TCS proteins, and a core of complete TCS proteins were shared between these genomes: 30 putative TCS clusters, 11 orphan HKs, 33 orphan RRs, and 16 hybrid HKs. A close analysis of the distribution of genes encoding TCS proteins revealed...

  2. Two-component signal transduction system SaeRS is involved in competence and penicillin susceptibility in Staphylococcus epidermidis.

    Science.gov (United States)

    Lou, Qiang; Ma, Yuanfang; Qu, Di

    2016-04-01

    Staphylococcus epidermidis, which is a causative pathogen of nosocomial infection, expresses its virulent traits such as biofilm and autolysis regulated by two-component signal transduction system SaeRS. In this study, the S. epidermidis SaeRS was identified to negatively regulate the expression of genes involved in competence (comF, murF), cytolysis (lrgA), and autolysis (lytS) by DNA microarray or real-time RT-PCR analysis. In addition, saeRS mutant showed increased competence and higher susceptibility to antibiotics such as penicillin and oxacillin than the wild-type strain. The study will be helpful for understanding the characterization of the SaeRS in S. epidermidis. © 2016 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.

  3. 454 Transcriptome sequencing suggests a role for two-component signalling in cellularization and differentiation of barley endosperm transfer cells.

    Directory of Open Access Journals (Sweden)

    Johannes Thiel

    Full Text Available BACKGROUND: Cell specification and differentiation in the endosperm of cereals starts at the maternal-filial boundary and generates the endosperm transfer cells (ETCs. Besides the importance in assimilate transfer, ETCs are proposed to play an essential role in the regulation of endosperm differentiation by affecting development of proximate endosperm tissues. We attempted to identify signalling elements involved in early endosperm differentiation by using a combination of laser-assisted microdissection and 454 transcriptome sequencing. PRINCIPAL FINDINGS: 454 sequencing of the differentiating ETC region from the syncytial state until functionality in transfer processes captured a high proportion of novel transcripts which are not available in existing barley EST databases. Intriguingly, the ETC-transcriptome showed a high abundance of elements of the two-component signalling (TCS system suggesting an outstanding role in ETC differentiation. All components and subfamilies of the TCS, including distinct kinds of membrane-bound receptors, have been identified to be expressed in ETCs. The TCS system represents an ancient signal transduction system firstly discovered in bacteria and has previously been shown to be co-opted by eukaryotes, like fungi and plants, whereas in animals and humans this signalling route does not exist. Transcript profiling of TCS elements by qRT-PCR suggested pivotal roles for specific phosphorelays activated in a coordinated time flow during ETC cellularization and differentiation. ETC-specificity of transcriptionally activated TCS phosphorelays was assessed for early differentiation and cellularization contrasting to an extension of expression to other grain tissues at the beginning of ETC maturation. Features of candidate genes of distinct phosphorelays and transcriptional activation of genes putatively implicated in hormone signalling pathways hint at a crosstalk of hormonal influences, putatively ABA and ethylene, and

  4. Searching for Potential Drug Targets in Two-component and Phosphorelay Signal-transduction Systems using Three-dimensional Cluster Analysis

    Institute of Scientific and Technical Information of China (English)

    Xiao-Hui CAI; Qing ZHANG; Shuo-Yong SHI; Da-Fu DING

    2005-01-01

    Two-component and phosphorelay signal transduction systems are central components in the virulence and antimicrobial resistance responses of a number of bacterial and fungal pathogens; in some cases, these systems are essential for bacterial growth and viability. Herein, we analyze in detail the conserved surface residue clusters in the phosphotransferase domain of histidine kinases and the regulatory domain of response regulators by using complex structure-based three-dimensional cluster analysis. We also investigatethe protein-protein interactions that these residue clusters participate in. The Spo0B-SpoOF complex structure was used as the reference structure, and the multiple aligned sequences of phosphotransferases and response regulators were paired correspondingly. The results show that a contiguous conserved residue cluster is formed around the active site, which crosses the interface of histidine kinases and response regulators. The conserved residue clusters of phosphotransferase and the regulatory domains are directly involved in the functional implementation of two-component signal transduction systems and are good targets for the development of novel antimicrobial agents.

  5. Autoregulation of lantibiotic bovicin HJ50 biosynthesis by the BovK-BovR two-component signal transduction system in Streptococcus bovis HJ50.

    Science.gov (United States)

    Ni, Jianqiang; Teng, Kunling; Liu, Gang; Qiao, Caixia; Huan, Liandong; Zhong, Jin

    2011-01-01

    Streptococcus bovis HJ50 produces a lacticin 481-like 33-amino-acid-residue lantibiotic, designated bovicin HJ50. bovK-bovR in the bovicin HJ50 biosynthetic gene cluster is predicted to be a two-component signal transduction system involved in sensing signals and regulating gene expression. Disruption of bovK or bovR resulted in the abrogation of bovicin HJ50 production, suggesting both genes play important roles in bovicin HJ50 biosynthesis. Addition of exogenous bovicin HJ50 peptide to cultures of a bovM mutant that lost the capability for bovicin HJ50 production and structural gene bovA transcription in S. bovis HJ50 induced dose-dependent transcription of the bovA gene, demonstrating that bovicin HJ50 production was normally autoregulated. The transcription of bovA was no longer induced by bovicin HJ50 in bovK and bovR disruption mutants, suggesting that BovK-BovR plays an essential role in the signal transduction regulating bovicin HJ50 biosynthesis. A phosphorylation assay indicated that BovK has the ability to autophosphorylate and subsequently transfer the phosphoryl group to the downstream BovR protein to carry on signal transduction. Electromobility shift assays (EMSA) and green fluorescent protein (GFP) reporter gene expression assays showed the specific binding of BovR to the bovA promoter, indicating that BovR regulates bovA expression by direct binding between them. Taken together, bovicin HJ50 biosynthesis is induced by bovicin HJ50 itself and regulated via the two-component signal transduction system BovK-BovR.

  6. Two-component magnetic structure of iron oxide nanoparticles mineralized in Listeria innocua protein cages

    Science.gov (United States)

    Usselman, Robert J.; Klem, Michael T.; Russek, Stephen E.; Young, Mark; Douglas, Trevor; Goldfarb, Ron B.

    2010-06-01

    Magnetometry was used to determine the magnetic properties of maghemite (γ-Fe2O3) nanoparticles formed within Listeria innocua protein cage. The electron magnetic resonance spectrum shows the presence of at least two magnetization components. The magnetization curves are explained by a sum of two Langevin functions in which each filled protein cage contains both a large magnetic iron oxide core plus an amorphous surface consisting of small noncoupled iron oxide spin clusters. This model qualitatively explains the observed decrease in the temperature dependent saturation moment and removes an unrealistic temperature dependent increase in the particle moment often observed in nanoparticle magnetization measurements.

  7. The VirSR two-component signal transduction system regulates NetB toxin production in Clostridium perfringens.

    Science.gov (United States)

    Cheung, Jackie K; Keyburn, Anthony L; Carter, Glen P; Lanckriet, Anouk L; Van Immerseel, Filip; Moore, Robert J; Rood, Julian I

    2010-07-01

    Clostridium perfringens causes several diseases in domestic livestock, including necrotic enteritis in chickens, which is of concern to the poultry industry due to its health implications and associated economic cost. The novel pore-forming toxin NetB is a critical virulence factor in the pathogenesis of this disease. In this study, we have examined the regulation of NetB toxin production. In C. perfringens, the quorum sensing-dependent VirSR two-component signal transduction system regulates genes encoding several toxins and extracellular enzymes. Analysis of the sequence upstream of the netB gene revealed the presence of potential DNA binding sites, or VirR boxes, that are recognized by the VirR response regulator. In vitro binding experiments showed that purified VirR was able to recognize and bind to these netB-associated VirR boxes. Furthermore, using a reporter gene assay, the netB VirR boxes were shown to be functional. Mutation of the virR gene in two avian C. perfringens strains was shown to significantly reduce the production of the NetB toxin; culture supernatants derived from these strains were no longer cytotoxic to Leghorn male hepatoma cells. Complementation with the virRS operon restored the toxin phenotypes to wild type. The results also showed that the VirSR two-component system regulates the expression of netB at the level of transcription. We postulate that in the gastrointestinal tract of infected birds, NetB production is upregulated when the population of C. perfringens cells reaches a threshold level that leads to activation of the VirSR system.

  8. The VirSR Two-Component Signal Transduction System Regulates NetB Toxin Production in Clostridium perfringens▿

    Science.gov (United States)

    Cheung, Jackie K.; Keyburn, Anthony L.; Carter, Glen P.; Lanckriet, Anouk L.; Van Immerseel, Filip; Moore, Robert J.; Rood, Julian I.

    2010-01-01

    Clostridium perfringens causes several diseases in domestic livestock, including necrotic enteritis in chickens, which is of concern to the poultry industry due to its health implications and associated economic cost. The novel pore-forming toxin NetB is a critical virulence factor in the pathogenesis of this disease. In this study, we have examined the regulation of NetB toxin production. In C. perfringens, the quorum sensing-dependent VirSR two-component signal transduction system regulates genes encoding several toxins and extracellular enzymes. Analysis of the sequence upstream of the netB gene revealed the presence of potential DNA binding sites, or VirR boxes, that are recognized by the VirR response regulator. In vitro binding experiments showed that purified VirR was able to recognize and bind to these netB-associated VirR boxes. Furthermore, using a reporter gene assay, the netB VirR boxes were shown to be functional. Mutation of the virR gene in two avian C. perfringens strains was shown to significantly reduce the production of the NetB toxin; culture supernatants derived from these strains were no longer cytotoxic to Leghorn male hepatoma cells. Complementation with the virRS operon restored the toxin phenotypes to wild type. The results also showed that the VirSR two-component system regulates the expression of netB at the level of transcription. We postulate that in the gastrointestinal tract of infected birds, NetB production is upregulated when the population of C. perfringens cells reaches a threshold level that leads to activation of the VirSR system. PMID:20457789

  9. Two-component signal transduction in Synechocystis sp. PCC 6803 under phosphate limitation: role of acetyl phosphate.

    Science.gov (United States)

    Juntarajumnong, Waraporn; Eaton-Rye, Julian J; Incharoensakdi, Aran

    2007-09-30

    The two-component signal transduction, which typically consists of a histidine kinase and a response regulator, is used by bacterial cells to sense changes in their environment. Previously, the SphS-SphR histidine kinase and response regulator pair of phosphate sensing signal transduction has been identified in Synechocystis sp. PCC 6803. In addition, some response regulators in bacteria have been shown to be cross regulated by low molecular weight phosphorylated compounds in the absence of the cognate histidine kinase. The ability of an endogenous acetyl phosphate to phosphorylate the response regulator, SphR in the absence of the cognate histidine kinase, SphS was therefore tested in Synechocystis sp. PCC 6803. The mutant lacking functional SphS and acetate kinase showed no detectable alkaline phosphatase activity under phosphate-limiting growth conditions. The results suggested that the endogenous acetyl phosphate accumulated inside the mutants could not activate the SphR via phosphorylation. On the other hand, exogenous acetyl phosphate could allow the mutant lacking functional acetate kinase and phosphotransacetylase to grow under phosphate-limiting conditions suggesting the role of acetyl phosphate as an energy source. Reverse transcription PCR demonstrated that the transcripts of acetate kinase and phosphotransacetylase genes in Synechocystis sp. PCC 6803 is upregulated in response to phosphate limitation suggesting the importance of these two enzymes for energy metabolism in Synechocystis cells.

  10. A simple, two-component buffer enhances use of chromatofocusing for processing of therapeutic proteins.

    Science.gov (United States)

    Logan, K A; Lagerlund, I; Chamow, S M

    1999-01-20

    To extend the feasibility of chromatofocusing to industrial use, we have developed a simple chromatofocusing buffer system capable of generating a smooth pH gradient without the use of an external gradient maker. Using two cationic buffering components, an internal pH gradient is produced on appropriate chromatography media over a broad pH range (9.5 to 5.0). The utility of this buffer system is demonstrated with PBE94 and DEAE Sepharose fast flow ion-exchangers, as well as with experimental fast flow chromatofocusing gels. Using a rapid flow rate, we evaluated this buffer system for recovery of a therapeutic protein from a bacterial cell extract. The simplicity of the buffer system requiring no external gradient maker, coupled with the use of fast flow chromatographic media to produce broad-range pH gradients, improves the scalability of chromatofocusing for processing of therapeutic proteins. Copyright 1999 John Wiley & Sons, Inc.

  11. Signal transduction and adaptive regulation through bacterial two-component systems: the Escherichia coli AtoSC paradigm.

    Science.gov (United States)

    Kyriakidis, Dimitrios A; Tiligada, Ekaterini

    2009-09-01

    Adaptive signal transduction within microbial cells involves a multi-faceted regulated phosphotransfer mechanism that comprises structural rearrangements of sensor histidine kinases upon ligand-binding and phosphorylation-induced conformational changes in response regulators of versatile two-component systems (TCS), arisen early in bacterial evolution. In Escherichia coli, cross-talk between the AtoS histidine kinase and the AtoC response regulator, forming the AtoSC TCS, through His --> Asp phosphotransfer, activates AtoC directly to induce atoDAEB operon expression, thus modulating diverse fundamental cellular processes such as short-chain fatty acid catabolism, poly-(R)-3-hydroxybutyrate biosynthesis and chemotaxis. Among the inducers hitherto identified, acetoacetate is the classical activator. The AtoSC TCS functional modulation by polyamines, histamine and Ca(2+), as well as the role of AtoC as transcriptional regulator, add new promising perspectives in the physiological significance and potential pharmacological exploitation of this TCS in cell proliferation, bacteria-host interactions, chemotaxis, and adaptation.

  12. The two-component signal transduction system YvcPQ regulates the bacterial resistance to bacitracin in Bacillus thuringiensis.

    Science.gov (United States)

    Zhang, Shumeng; Li, Xinfeng; Wang, Xun; Li, Zhou; He, Jin

    2016-10-01

    YvcPQ is one of the two-component signal transduction systems that respond to specific stimuli and enable cells to adjust multiple cellular functions. It consists of a histidine kinase YvcQ and a response regulator YvcP. In this study, through searching the consensus sequence recognized by YvcP, we found four YvcP-binding motifs in the promoter regions of genes yvcR (BMB171_C4100), BMB171_C4385, kapD (BMB171_C4525) and BMB171_C4835 in Bacillus thuringiensis BMB171 which is a representative of Bacillus cereus group, and confirmed that these genes are regulated by YvcP. We compared the sequence of yvcPQ and its downstream genes in genus Bacillus, and found two different kinds of yvc locus, one was the yvcPQ-RS in B. subtilis species and the other was the yvcPQ-R-S1S2 in B. cereus group. Furthermore, we found that YvcP activates the transcription of yvcS1S2 (downstream of yvcR) to promote bacterial resistance to bacitracin and deletion of either yvcPQ operon or yvcS1S2 operon renders the bacterial cells more sensitive to bacitracin. This study enriched our understanding of both the YvcPQ's function and the mechanism of bacterial resistance to bacitracin.

  13. A conserved two-component signal transduction system controls the response to phosphate starvation in Bifidobacterium breve UCC2003.

    Science.gov (United States)

    Alvarez-Martin, Pablo; Fernández, Matilde; O'Connell-Motherway, Mary; O'Connell, Kerry Joan; Sauvageot, Nicolas; Fitzgerald, Gerald F; MacSharry, John; Zomer, Aldert; van Sinderen, Douwe

    2012-08-01

    This work reports on the identification and molecular characterization of the two-component regulatory system (2CRS) PhoRP, which controls the response to inorganic phosphate (P(i)) starvation in Bifidobacterium breve UCC2003. The response regulator PhoP was shown to bind to the promoter region of pstSCAB, specifying a predicted P(i) transporter system, as well as that of phoU, which encodes a putative P(i)-responsive regulatory protein. This interaction is assumed to cause transcriptional modulation under conditions of P(i) limitation. Our data suggest that the phoRP genes are subject to positive autoregulation and, together with pstSCAB and presumably phoU, represent the complete regulon controlled by the phoRP-encoded 2CRS in B. breve UCC2003. Determination of the minimal PhoP binding region combined with bioinformatic analysis revealed the probable recognition sequence of PhoP, designated here as the PHO box, which together with phoRP is conserved among many high-GC-content Gram-positive bacteria. The importance of the phoRP 2CRS in the response of B. breve to P(i) starvation conditions was confirmed by analysis of a B. breve phoP insertion mutant which exhibited decreased growth under phosphate-limiting conditions compared to its parent strain UCC2003.

  14. A genome-wide study of two-component signal transduction systems in eight newly sequenced mutans streptococci strains.

    Science.gov (United States)

    Song, Lifu; Sudhakar, Padhmanand; Wang, Wei; Conrads, Georg; Brock, Anke; Sun, Jibin; Wagner-Döbler, Irene; Zeng, An-Ping

    2012-04-04

    Mutans streptococci are a group of gram-positive bacteria including the primary cariogenic dental pathogen Streptococcus mutans and closely related species. Two component systems (TCSs) composed of a signal sensing histidine kinase (HK) and a response regulator (RR) play key roles in pathogenicity, but have not been comparatively studied for these oral bacterial pathogens. HKs and RRs of 8 newly sequenced mutans streptococci strains, including S. sobrinus DSM20742, S. ratti DSM20564 and six S. mutans strains, were identified and compared to the TCSs of S. mutans UA159 and NN2025, two previously genome sequenced S. mutans strains. Ortholog analysis revealed 18 TCS clusters (HK-RR pairs), 2 orphan HKs and 2 orphan RRs, of which 8 TCS clusters were common to all 10 strains, 6 were absent in one or more strains, and the other 4 were exclusive to individual strains. Further classification of the predicted HKs and RRs revealed interesting aspects of their putative functions. While TCS complements were comparable within the six S. mutans strains, S. sobrinus DSM20742 lacked TCSs possibly involved in acid tolerance and fructan catabolism, and S. ratti DSM20564 possessed 3 unique TCSs but lacked the quorum-sensing related TCS (ComDE). Selected computational predictions were verified by PCR experiments. Differences in the TCS repertoires of mutans streptococci strains, especially those of S. sobrinus and S. ratti in comparison to S. mutans, imply differences in their response mechanisms for survival in the dynamic oral environment. This genomic level study of TCSs should help in understanding the pathogenicity of these mutans streptococci strains.

  15. Evolutionary history of the OmpR/IIIA family of signal transduction two component systems in Lactobacillaceae and Leuconostocaceae

    Directory of Open Access Journals (Sweden)

    González-Candelas Fernando

    2011-02-01

    Full Text Available Abstract Background Two component systems (TCS are signal transduction pathways which typically consist of a sensor histidine kinase (HK and a response regulator (RR. In this study, we have analyzed the evolution of TCS of the OmpR/IIIA family in Lactobacillaceae and Leuconostocaceae, two families belonging to the group of lactic acid bacteria (LAB. LAB colonize nutrient-rich environments such as foodstuffs, plant materials and the gastrointestinal tract of animals thus driving the study of this group of both basic and applied interest. Results The genomes of 19 strains belonging to 16 different species have been analyzed. The number of TCS encoded by the strains considered in this study varied between 4 in Lactobacillus helveticus and 17 in Lactobacillus casei. The OmpR/IIIA family was the most prevalent in Lactobacillaceae accounting for 71% of the TCS present in this group. The phylogenetic analysis shows that no new TCS of this family has recently evolved in these Lactobacillaceae by either lineage-specific gene expansion or domain shuffling. Furthermore, no clear evidence of non-orthologous replacements of either RR or HK partners has been obtained, thus indicating that coevolution of cognate RR and HKs has been prevalent in Lactobacillaceae. Conclusions The results obtained suggest that vertical inheritance of TCS present in the last common ancestor and lineage-specific gene losses appear as the main evolutionary forces involved in their evolution in Lactobacillaceae, although some HGT events cannot be ruled out. This would agree with the genomic analyses of Lactobacillales which show that gene losses have been a major trend in the evolution of this group.

  16. A novel two-component signaling system facilitates uropathogenic Escherichia coli's ability to exploit abundant host metabolites.

    Directory of Open Access Journals (Sweden)

    Wentong Cai

    Full Text Available Two-component signaling systems (TCSs are major mechanisms by which bacteria adapt to environmental conditions. It follows then that TCSs would play important roles in the adaptation of pathogenic bacteria to host environments. However, no pathogen-associated TCS has been identified in uropathogenic Escherichia coli (UPEC. Here, we identified a novel TCS, which we termed KguS/KguR (KguS: α-ketoglutarate utilization sensor; KguR: α-ketoglutarate utilization regulator in UPEC CFT073, a strain isolated from human pyelonephritis. kguS/kguR was strongly associated with UPEC but was found only rarely among other E. coli including commensal and intestinal pathogenic strains. An in vivo competition assay in a mouse UTI model showed that deletion of kguS/kguR in UPEC CFT073 resulted in a significant reduction in its colonization of the bladders and kidneys of mice, suggesting that KguS/KguR contributed to UPEC fitness in vivo. Comparative proteomics identified the target gene products of KguS/KguR, and sequence analysis showed that TCS KguS/KguR and its targeted-genes, c5032 to c5039, are encoded on a genomic island, which is not present in intestinal pathogenic E. coli. Expression of the target genes was induced by α-ketoglutarate (α-KG. These genes were further shown to be involved in utilization of α-KG as a sole carbon source under anaerobic conditions. KguS/KguR contributed to the regulation of the target genes with the direct regulation by KguR verified using an electrophoretic mobility shift assay. In addition, oxygen deficiency positively modulated expression of kguS/kguR and its target genes. Taken altogether, this study describes the first UPEC-associated TCS that functions in controlling the utilization of α-ketoglutarate in vivo thereby facilitating UPEC adaptation to life inside the urinary tract.

  17. A novel two-component signaling system facilitates uropathogenic Escherichia coli's ability to exploit abundant host metabolites.

    Science.gov (United States)

    Cai, Wentong; Wannemuehler, Yvonne; Dell'anna, Giuseppe; Nicholson, Bryon; Barbieri, Nicolle L; Kariyawasam, Subhashinie; Feng, Yaping; Logue, Catherine M; Nolan, Lisa K; Li, Ganwu

    2013-01-01

    Two-component signaling systems (TCSs) are major mechanisms by which bacteria adapt to environmental conditions. It follows then that TCSs would play important roles in the adaptation of pathogenic bacteria to host environments. However, no pathogen-associated TCS has been identified in uropathogenic Escherichia coli (UPEC). Here, we identified a novel TCS, which we termed KguS/KguR (KguS: α-ketoglutarate utilization sensor; KguR: α-ketoglutarate utilization regulator) in UPEC CFT073, a strain isolated from human pyelonephritis. kguS/kguR was strongly associated with UPEC but was found only rarely among other E. coli including commensal and intestinal pathogenic strains. An in vivo competition assay in a mouse UTI model showed that deletion of kguS/kguR in UPEC CFT073 resulted in a significant reduction in its colonization of the bladders and kidneys of mice, suggesting that KguS/KguR contributed to UPEC fitness in vivo. Comparative proteomics identified the target gene products of KguS/KguR, and sequence analysis showed that TCS KguS/KguR and its targeted-genes, c5032 to c5039, are encoded on a genomic island, which is not present in intestinal pathogenic E. coli. Expression of the target genes was induced by α-ketoglutarate (α-KG). These genes were further shown to be involved in utilization of α-KG as a sole carbon source under anaerobic conditions. KguS/KguR contributed to the regulation of the target genes with the direct regulation by KguR verified using an electrophoretic mobility shift assay. In addition, oxygen deficiency positively modulated expression of kguS/kguR and its target genes. Taken altogether, this study describes the first UPEC-associated TCS that functions in controlling the utilization of α-ketoglutarate in vivo thereby facilitating UPEC adaptation to life inside the urinary tract.

  18. Functional characterization of the histidine kinase of the E. coli two-component signal transduction system AtoS-AtoC.

    Science.gov (United States)

    Filippou, Panagiota S; Kasemian, Lucy D; Panagiotidis, Christos A; Kyriakidis, Dimitrios A

    2008-09-01

    The Escherichia coli AtoS-AtoC two-component signal transduction system regulates the expression of the atoDAEB operon genes, whose products are required for short-chain fatty acid catabolism. In this study purified his-tagged wild-type and mutant AtoS proteins were used to prove that these proteins are true sensor kinases. The phosphorylated residue was identified as the histidine-398, which was located in a conserved Eta-box since AtoS carrying a mutation at this site failed to phosphorylate. This inability to phosphorylate was not due to gross structural alterations of AtoS since the H398L mutant retained its capability to bind ATP. Furthermore, the H398L mutant AtoS was competent to catalyze the trans-phosphorylation of an AtoS G-box (G565A) mutant protein which otherwise failed to autophosphorylate due to its inability to bind ATP. The formation of homodimers between the various AtoS proteins was also shown by cross-linking experiments both in vitro and in vivo.

  19. A survey of HK, HPt, and RR domains and their organization in two-component systems and phosphorelay proteins of organisms with fully sequenced genomes

    Science.gov (United States)

    Salvado, Baldiri; Vilaprinyo, Ester; Sorribas, Albert

    2015-01-01

    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 7,609 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. PMID:26339559

  20. A survey of HK, HPt, and RR domains and their organization in two-component systems and phosphorelay proteins of organisms with fully sequenced genomes

    Directory of Open Access Journals (Sweden)

    Baldiri Salvado

    2015-08-01

    Full Text Available 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 7,609 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.

  1. A Conserved Two-Component Signal Transduction System Controls the Response to Phosphate Starvation in Bifidobacterium breve UCC2003.

    NARCIS (Netherlands)

    Alvarez-Martin, P.; Fernandez, M.; O'Connell-Motherway, M.; O'Connell, K.J.; Sauvageot, N.; Fitzgerald, G.F.; Macsharry, J.; Zomer, A.L.; Sinderen, D. van

    2012-01-01

    This work reports on the identification and molecular characterization of the two-component regulatory system (2CRS) PhoRP, which controls the response to inorganic phosphate (P(i)) starvation in Bifidobacterium breve UCC2003. The response regulator PhoP was shown to bind to the promoter region of

  2. Two-component signal transduction in Agaricus bisporus: a comparative genomic analysis with other basidiomycetes through the web-based tool BASID2CS.

    Science.gov (United States)

    Lavín, José L; García-Yoldi, Alberto; Ramírez, Lucía; Pisabarro, Antonio G; Oguiza, José A

    2013-06-01

    Two-component systems (TCSs) are signal transduction mechanisms present in many eukaryotes, including fungi that play essential roles in the regulation of several cellular functions and responses. In this study, we carry out a genomic analysis of the TCS proteins in two varieties of the white button mushroom Agaricus bisporus. The genomes of both A. bisporus varieties contain eight genes coding for TCS proteins, which include four hybrid Histidine Kinases (HKs), a single histidine-containing phosphotransfer (HPt) protein and three Response Regulators (RRs). Comparison of the TCS proteins among A. bisporus and the sequenced basidiomycetes showed a conserved core complement of five TCS proteins including the Tco1/Nik1 hybrid HK, HPt protein and Ssk1, Skn7 and Rim15-like RRs. In addition, Dual-HKs, unusual hybrid HKs with 2 HK and 2 RR domains, are absent in A. bisporus and are limited to various species of basidiomycetes. Differential expression analysis showed no significant up- or down-regulation of the Agaricus TCS genes in the conditions/tissue analyzed with the exception of the Skn7-like RR gene (Agabi_varbisH97_2|198669) that is significantly up-regulated on compost compared to cultured mycelia. Furthermore, the pipeline web server BASID2CS (http://bioinformatics.unavarra.es:1000/B2CS/BASID2CS.htm) has been specifically designed for the identification, classification and functional annotation of putative TCS proteins from any predicted proteome of basidiomycetes using a combination of several bioinformatic approaches. Copyright © 2012 Elsevier Inc. All rights reserved.

  3. Biochemical Activities of Three Pairs of Ehrlichia chaffeensis Two-Component Regulatory System Proteins Involved in Inhibition of Lysosomal Fusion†

    Science.gov (United States)

    Kumagai, Yumi; Cheng, Zhihui; Lin, Mingqun; Rikihisa, Yasuko

    2006-01-01

    Ehrlichia chaffeensis, the etiologic agent of human monocytic ehrlichiosis, replicates in early endosomes by avoiding lysosomal fusion in monocytes and macrophages. In E. chaffeensis we predicted three pairs of putative two-component regulatory systems (TCSs) designated PleC-PleD, NtrY-NtrX, and CckA-CtrA based on amino acid sequence homology. In the present study to determine biochemical pairs and specificities of the TCSs, the recombinant proteins of the three putative histidine kinase (HK) kinase domains (rPleCHKD, rNtrYHKD, and MBP-rCckAHKD) and the full-length forms of three putative response regulators (RRs) (rPleD, rNtrX, and rCtrA) as well as the respective mutant recombinant proteins (rPleCHKDH244A, rNtrYHKDH498A, MBP-rCckAHKDH449A, rPleDD53A, rNtrXD59A, and rCtrAD53A) were expressed and purified as soluble proteins. The in vitro HK activity, the specific His residue-dependent autophosphorylation of the kinase domain, was demonstrated in the three HKs. The specific Asp residue-dependent in vitro phosphotransfer from the kinase domain to the putative cognate RR was demonstrated in each of the three RRs. Western blot analysis of E. chaffeensis membrane and soluble fractions using antibodies specific for each recombinant protein detected PleC and CckA in the membrane fraction, whereas it detected NtrY, NtrX, and PleD in the soluble fraction. CtrA was found in the two fractions at similar levels. E. chaffeensis was sensitive to closantel, an HK inhibitor. Closantel treatment induced lysosomal fusion of the E. chaffeensis inclusion in a human monocytic leukemia cell line, THP-1 cells, implying that functional TCSs are essential in preventing lysosomal fusion of the E. chaffeensis inclusion compartment. PMID:16926392

  4. A hybrid two-component system of Tannerella forsythia affects autoaggregation and post-translational modification of surface proteins.

    Science.gov (United States)

    Niwa, Daisuke; Nishikawa, Kiyoshi; Nakamura, Hiroshi

    2011-05-01

    Tannerella forsythia is a Gram-negative oral anaerobe closely associated with both periodontal and periapical diseases. The ORF TF0022 of strain ATCC 43037 encodes a hybrid two-component system consisting of an N-terminal histidine kinase and a C-terminal response regulator. Disruption of the TF0022 locus enhanced autoaggregation of the broth-cultured cells. Comparative proteome analyses revealed that two S-layer proteins in the TF0022 mutant exhibited decreased apparent masses by denaturing gel electrophoresis, suggesting a deficiency in post-translational modification. Furthermore, the mutant decreased the production of a glycosyltransferase encoded by TF1061 that is located in a putative glycosylation-related gene cluster. Quantitative real-time PCR revealed reduced transcription of TF1061 and the associated genes in the TF0022 mutant. These results indicate that TF0022 upregulates the expression of the glycosylation-related genes and suggest modulation of the autoaggregation of T. forsythia cells by a possible post-translational modification of cell-surface components.

  5. Conjugal transfer of the Sinorhizobium meliloti 1021 symbiotic plasmid is governed through the concerted action of one- and two-component signal transduction regulators.

    Science.gov (United States)

    Nogales, Joaquina; Blanca-Ordóñez, Helena; Olivares, José; Sanjuán, Juan

    2013-03-01

    Conjugal transfer of Sinorhizobium meliloti and Rhizobium etli symbiotic plasmids are repressed by the transcriptional regulator RctA. Here we report on new key players in the signal transduction cascade towards S. meliloti pSym conjugation. We have identified S. meliloti pSymA gene SMa0974 as an orthologue of the R. etli rctB gene which is required to antagonize repression by RctA. In S. meliloti two additional genes, rctR and rctC participate in control of rctB expression. rctR (SMa0955) encodes a protein of the GntR family of transcriptional regulators involved in repression of rctB. A rctR mutant promotes pSymA conjugal transfer and displays increased transcription of tra, virB and rctB genes even in presence of wild-type rctA gene. Among genes repressed by RctR, rctC (SMa0961) encodes a response regulator required to activate rctB transcription and therefore for derepression of plasmid conjugative functions. We conclude that in both R. etli and S. meliloti pSym conjugal transfer is derepressed via rctB, however the regulatory cascades to achieve activation of rctB are probably different. Upstream of rctB, the S. meliloti pSym conjugal transfer is regulated through the concerted action of genes representing one- (rctR) and two-component (rctC) signal transduction systems in response to yet unidentified signals.

  6. Systems level analysis of two-component signal transduction systems in Erwinia amylovora: Role in virulence, regulation of amylovoran biosynthesis and swarming motility

    Directory of Open Access Journals (Sweden)

    Sundin George W

    2009-05-01

    Full Text Available Abstract Background Two-component signal transduction systems (TCSTs, consisting of a histidine kinase (HK and a response regulator (RR, represent a major paradigm for signal transduction in prokaryotes. TCSTs play critical roles in sensing and responding to environmental conditions, and in bacterial pathogenesis. Most TCSTs in Erwinia amylovora have either not been identified or have not yet been studied. Results We used a systems approach to identify TCST and related signal transduction genes in the genome of E. amylovora. Comparative genomic analysis of TCSTs indicated that E. amylovora TCSTs were closely related to those of Erwinia tasmaniensis, a saprophytic enterobacterium isolated from apple flowers, and to other enterobacteria. Forty-six TCST genes in E. amylovora including 17 sensor kinases, three hybrid kinases, 20 DNA- or ligand-binding RRs, four RRs with enzymatic output domain (EAL-GGDEF proteins, and two kinases were characterized in this study. A systematic TCST gene-knockout experiment was conducted, generating a total of 59 single-, double-, and triple-mutants. Virulence assays revealed that five of these mutants were non-pathogenic on immature pear fruits. Results from phenotypic characterization and gene expression experiments indicated that several groups of TCST systems in E. amylovora control amylovoran biosynthesis, one of two major virulence factors in E. amylovora. Both negative and positive regulators of amylovoran biosynthesis were identified, indicating a complex network may control this important feature of pathogenesis. Positive (non-motile, EnvZ/OmpR, negative (hypermotile, GrrS/GrrA, and intermediate regulators for swarming motility in E. amylovora were also identified. Conclusion Our results demonstrated that TCSTs in E. amylovora played major roles in virulence on immature pear fruit and in regulating amylovoran biosynthesis and swarming motility. This suggested presence of regulatory networks governing

  7. Systems level analysis of two-component signal transduction systems in Erwinia amylovora: role in virulence, regulation of amylovoran biosynthesis and swarming motility.

    Science.gov (United States)

    Zhao, Youfu; Wang, Dongping; Nakka, Sridevi; Sundin, George W; Korban, Schuyler S

    2009-05-26

    Two-component signal transduction systems (TCSTs), consisting of a histidine kinase (HK) and a response regulator (RR), represent a major paradigm for signal transduction in prokaryotes. TCSTs play critical roles in sensing and responding to environmental conditions, and in bacterial pathogenesis. Most TCSTs in Erwinia amylovora have either not been identified or have not yet been studied. We used a systems approach to identify TCST and related signal transduction genes in the genome of E. amylovora. Comparative genomic analysis of TCSTs indicated that E. amylovora TCSTs were closely related to those of Erwinia tasmaniensis, a saprophytic enterobacterium isolated from apple flowers, and to other enterobacteria. Forty-six TCST genes in E. amylovora including 17 sensor kinases, three hybrid kinases, 20 DNA- or ligand-binding RRs, four RRs with enzymatic output domain (EAL-GGDEF proteins), and two kinases were characterized in this study. A systematic TCST gene-knockout experiment was conducted, generating a total of 59 single-, double-, and triple-mutants. Virulence assays revealed that five of these mutants were non-pathogenic on immature pear fruits. Results from phenotypic characterization and gene expression experiments indicated that several groups of TCST systems in E. amylovora control amylovoran biosynthesis, one of two major virulence factors in E. amylovora. Both negative and positive regulators of amylovoran biosynthesis were identified, indicating a complex network may control this important feature of pathogenesis. Positive (non-motile, EnvZ/OmpR), negative (hypermotile, GrrS/GrrA), and intermediate regulators for swarming motility in E. amylovora were also identified. Our results demonstrated that TCSTs in E. amylovora played major roles in virulence on immature pear fruit and in regulating amylovoran biosynthesis and swarming motility. This suggested presence of regulatory networks governing expression of critical virulence genes in E. amylovora.

  8. ‘Forcing the issue’ - Aromatic tuning facilitates stimulus-independent modulation of a two-component signaling circuit

    DEFF Research Database (Denmark)

    Nørholm, Morten; von Heijne, Gunnar; Draheim, Roger R.

    2014-01-01

    desirable because particular physiological or developmental processes could be controlled for biotechnological purposes without the need to identify the stimulus itself. Here, we demonstrate that aromatic tuning, i.e., repositioning the aromatic residues commonly found at the cytoplasmic end of the receptor...... receptors were not locked in a single conformation. We also noted that circuits containing aromatically tuned variants became more sensitive to changes in the receptor concentration than their wild-type counterpart, suggesting a new way to study mechanisms underpinning receptor concentration......-dependent robustness. We believe that aromatic tuning has several advantages compared to previous methods aimed at stimulus-independent modulation of receptors and that it will be generally applicable to a wide-range of two-component circuits....

  9. OmpR, a response regulator of the two-component signal transduction pathway, influences inv gene expression in Yersinia enterocolitica O9

    Directory of Open Access Journals (Sweden)

    Marta eBrzóstkowska

    2012-12-01

    Full Text Available The environmental control of invasin expression in Yersinia enterocolitica is mediated by a regulatory network composed of negative and positive regulators of inv gene transcription. Previously, we demonstrated that OmpR, a response regulator of the two-component signal transduction pathway EnvZ/OmpR, negatively regulates invasin gene expression in Y. enterocolitica O9 by direct interaction with the inv promoter region. This study was undertaken to clarify the role of OmpR in the inv regulatory circuit in which RovA protein has been shown to positively regulate inv transcription. Using ompR, rovA and ompR rovA Y. enterocolitica mutant backgrounds we showed that the inhibitory effect of OmpR on inv transcription may be observed only when RovA is present/active in Y. enterocolitica cells. To extend our research on inv regulation we examined the effect of OmpR on rovA gene expression. Analysis of rovA-lacZ transcriptional fusion in Y. enterocolitica wild-type and ompR background indicated that OmpR does not influence rovA expression. Thus, our results indicate that OmpR influences invasin expression directly via binding to the inv promoter, but not through modulation of rovA expression.

  10. OmpR, a response regulator of the two-component signal transduction pathway, influences inv gene expression in Yersinia enterocolitica O9.

    Science.gov (United States)

    Brzóstkowska, Marta; Raczkowska, Adrianna; Brzostek, Katarzyna

    2012-01-01

    The environmental control of invasin (inv) expression in Yersinia enterocolitica is mediated by a regulatory network composed of negative and positive regulators of inv gene transcription. Previously, we demonstrated that OmpR, a response regulator of the two-component signal transduction pathway EnvZ/OmpR, negatively regulates inv gene expression in Y. enterocolitica O9 by direct interaction with the inv promoter region. This study was undertaken to clarify the role of OmpR in the inv regulatory circuit in which RovA protein has been shown to positively regulate inv transcription. Using ompR, rovA, and ompR rovA Y. enterocolitica mutant backgrounds we showed that the inhibitory effect of OmpR on inv transcription may be observed only when RovA is present/active in Y. enterocolitica cells. To extend our research on inv regulation we examined the effect of OmpR on rovA gene expression. Analysis of rovA-lacZ transcriptional fusion in Y. enterocolitica wild-type and ompR background indicated that OmpR does not influence rovA expression. Thus, our results indicate that OmpR influences inv expression directly via binding to the inv promoter, but not through modulation of rovA expression.

  11. Two-component signal transduction system CBO0787/CBO0786 represses transcription from botulinum neurotoxin promoters in Clostridium botulinum ATCC 3502.

    Science.gov (United States)

    Zhang, Zhen; Korkeala, Hannu; Dahlsten, Elias; Sahala, Elina; Heap, John T; Minton, Nigel P; Lindström, Miia

    2013-03-01

    Blocking neurotransmission, botulinum neurotoxin is the most poisonous biological substance known to mankind. Despite its infamy as the scourge of the food industry, the neurotoxin is increasingly used as a pharmaceutical to treat an expanding range of muscle disorders. Whilst neurotoxin expression by the spore-forming bacterium Clostridium botulinum appears tightly regulated, to date only positive regulatory elements, such as the alternative sigma factor BotR, have been implicated in this control. The identification of negative regulators has proven to be elusive. Here, we show that the two-component signal transduction system CBO0787/CBO0786 negatively regulates botulinum neurotoxin expression. Single insertional inactivation of cbo0787 encoding a sensor histidine kinase, or of cbo0786 encoding a response regulator, resulted in significantly elevated neurotoxin gene expression levels and increased neurotoxin production. Recombinant CBO0786 regulator was shown to bind to the conserved -10 site of the core promoters of the ha and ntnh-botA operons, which encode the toxin structural and accessory proteins. Increasing concentration of CBO0786 inhibited BotR-directed transcription from the ha and ntnh-botA promoters, demonstrating direct transcriptional repression of the ha and ntnh-botA operons by CBO0786. Thus, we propose that CBO0786 represses neurotoxin gene expression by blocking BotR-directed transcription from the neurotoxin promoters. This is the first evidence of a negative regulator controlling botulinum neurotoxin production. Understanding the neurotoxin regulatory mechanisms is a major target of the food and pharmaceutical industries alike.

  12. A Moraxella catarrhalis two-component signal transduction system necessary for growth in liquid media affects production of two lysozyme inhibitors.

    Science.gov (United States)

    Joslin, Stephanie N; Pybus, Christine; Labandeira-Rey, Maria; Evans, Amanda S; Attia, Ahmed S; Brautigam, Chad A; Hansen, Eric J

    2015-01-01

    There are a paucity of data concerning gene products that could contribute to the ability of Moraxella catarrhalis to colonize the human nasopharynx. Inactivation of a gene (mesR) encoding a predicted response regulator of a two-component signal transduction system in M. catarrhalis yielded a mutant unable to grow in liquid media. This mesR mutant also exhibited increased sensitivity to certain stressors, including polymyxin B, SDS, and hydrogen peroxide. Inactivation of the gene (mesS) encoding the predicted cognate sensor (histidine) kinase yielded a mutant with the same inability to grow in liquid media as the mesR mutant. DNA microarray and real-time reverse transcriptase PCR analyses indicated that several genes previously shown to be involved in the ability of M. catarrhalis to persist in the chinchilla nasopharynx were upregulated in the mesR mutant. Two other open reading frames upregulated in the mesR mutant were shown to encode small proteins (LipA and LipB) that had amino acid sequence homology to bacterial adhesins and structural homology to bacterial lysozyme inhibitors. Inactivation of both lipA and lipB did not affect the ability of M. catarrhalis O35E to attach to a human bronchial epithelial cell line in vitro. Purified recombinant LipA and LipB fusion proteins were each shown to inhibit human lysozyme activity in vitro and in saliva. A lipA lipB deletion mutant was more sensitive than the wild-type parent strain to killing by human lysozyme in the presence of human apolactoferrin. This is the first report of the production of lysozyme inhibitors by M. catarrhalis. Copyright © 2015, American Society for Microbiology. All Rights Reserved.

  13. Genome-wide survey of two-component signal transduction systems in the plant growth-promoting bacterium Azospirillum.

    Science.gov (United States)

    Borland, Stéphanie; Oudart, Anne; Prigent-Combaret, Claire; Brochier-Armanet, Céline; Wisniewski-Dyé, Florence

    2015-10-22

    Two-component systems (TCS) play critical roles in sensing and responding to environmental cues. Azospirillum is a plant growth-promoting rhizobacterium living in the rhizosphere of many important crops. Despite numerous studies about its plant beneficial properties, little is known about how the bacterium senses and responds to its rhizospheric environment. The availability of complete genome sequenced from four Azospirillum strains (A. brasilense Sp245 and CBG 497, A. lipoferum 4B and Azospirillum sp. B510) offers the opportunity to conduct a comprehensive comparative analysis of the TCS gene family. Azospirillum genomes harbour a very large number of genes encoding TCS, and are especially enriched in hybrid histidine kinases (HyHK) genes compared to other plant-associated bacteria of similar genome sizes. We gained further insight into HyHK structure and architecture, revealing an intriguing complexity of these systems. An unusual proportion of TCS genes were orphaned or in complex clusters, and a high proportion of predicted soluble HKs compared to other plant-associated bacteria are reported. Phylogenetic analyses of the transmitter and receiver domains of A. lipoferum 4B HyHK indicate that expansion of this family mainly arose through horizontal gene transfer but also through gene duplications all along the diversification of the Azospirillum genus. By performing a genome-wide comparison of TCS, we unraveled important 'genus-defining' and 'plant-specifying' TCS. This study shed light on Azospirillum TCS which may confer important regulatory flexibility. Collectively, these findings highlight that Azospirillum genomes have broad potential for adaptation to fluctuating environments.

  14. Biophysical and physiological characterization of ZraP from Escherichia coli, the periplasmic accessory protein of the atypical ZraSR two-component system.

    Science.gov (United States)

    Petit-Härtlein, Isabelle; Rome, Kevin; de Rosny, Eve; Molton, Florian; Duboc, Carole; Gueguen, Erwan; Rodrigue, Agnès; Covès, Jacques

    2015-12-01

    The ZraSR system belongs to the family of TCSs (two-component signal transduction systems). In Escherichia coli, it was proposed to participate in zinc balance and to protect cytoplasmic zinc overload by sequestering this metal ion into the periplasm. This system controls the expression of the accessory protein ZraP that would be a periplasmic zinc scavenger. ZraPSR is functionally homologous with CpxPAR that integrates signals of envelope perturbation, including misfolded periplasmic proteins. The auxiliary periplasmic regulator CpxP inhibits the Cpx pathway by interacting with CpxA. Upon envelope stress sensing, the inhibitory function of CpxP is relieved, resulting in CpxR activation. Similarly to CpxPAR, ZraPSR probably plays a role in envelope stress response as a zinc-dependent chaperone activity was demonstrated for ZraP in Salmonella. We have purified ZraP from E. coli and shown that it is an octamer containing four interfacial metal-binding sites contributing to dimer stability. These sites are located close to the N-terminus, whereas the C-terminus is involved in polymerization of the protein to form a tetramer of dimers. In vitro, ZraP binds copper with a higher affinity than zinc and displays chaperone properties partially dependent on zinc binding. In vivo, zinc-bound ZraP is a repressor of the expression of the zraPSR operon. However, we have demonstrated that none of the Zra proteins are involved in zinc or copper resistance. We propose an integrated mechanism in which zinc is a marker of envelope stress perturbation and ZraPSR TCS is a sentinel sensing and responding to zinc entry into the periplasm.

  15. Expression and maintenance of ComD-ComE, the two-component signal-transduction system that controls competence of Streptococcus pneumoniae.

    Science.gov (United States)

    Martin, Bernard; Granadel, Chantal; Campo, Nathalie; Hénard, Vincent; Prudhomme, Marc; Claverys, Jean-Pierre

    2010-03-01

    A secreted competence-stimulating peptide (CSP), encoded by comC, constitutes, together with the two-component system ComD-ComE, the master switch for competence induction in Streptococcus pneumoniae. Interaction between CSP and its membrane-bound histidine-kinase receptor, ComD, is believed to lead to autophosphorylation of ComD, which then transphosphorylates the ComE response regulator to activate transcription of a limited set of genes, including the comCDE operon. This generates a positive feedback loop, amplifying the signal and co-ordinating competence throughout the population. On the other hand, the promoter(s) and proteins important for basal comCDE expression have not been defined. We now report that CSP-induced and basal comCDE transcription both initiate from the same promoter, P(E); that basal expression necessitates the presence of both ComD and a phosphate-accepting form of ComE, but not CSP; and that overexpression of ComE(R120S) triggers ComD-dependent transformation in the absence of CSP. These observations suggest that self-activation of ComD is required for basal comCDE expression. We also establish that transcriptional readthrough occurs across the tRNA(Arg5) terminator and contributes significantly to comCDE expression. Finally, we demonstrate by various means, including single-cell competence analysis with GFP, that readthrough is crucial to avoid the stochastic production of CSP non-responsive cells lacking ComD or ComE.

  16. The two-component signal transduction system CopRS of Corynebacterium glutamicum is required for adaptation to copper-excess stress.

    Science.gov (United States)

    Schelder, Stephanie; Zaade, Daniela; Litsanov, Boris; Bott, Michael; Brocker, Melanie

    2011-01-01

    Copper is an essential cofactor for many enzymes but at high concentrations it is toxic for the cell. Copper ion concentrations ≥50 µM inhibited growth of Corynebacterium glutamicum. The transcriptional response to 20 µM Cu(2+) was studied using DNA microarrays and revealed 20 genes that showed a ≥ 3-fold increased mRNA level, including cg3281-cg3289. Several genes in this genomic region code for proteins presumably involved in the adaption to copper-induced stress, e. g. a multicopper oxidase (CopO) and a copper-transport ATPase (CopB). In addition, this region includes the copRS genes (previously named cgtRS9) which encode a two-component signal transduction system composed of the histidine kinase CopS and the response regulator CopR. Deletion of the copRS genes increased the sensitivity of C. glutamicum towards copper ions, but not to other heavy metal ions. Using comparative transcriptome analysis of the ΔcopRS mutant and the wild type in combination with electrophoretic mobility shift assays and reporter gene studies the CopR regulon and the DNA-binding motif of CopR were identified. Evidence was obtained that CopR binds only to the intergenic region between cg3285 (copR) and cg3286 in the genome of C. glutamicum and activates expression of the divergently oriented gene clusters cg3285-cg3281 and cg3286-cg3289. Altogether, our data suggest that CopRS is the key regulatory system in C. glutamicum for the extracytoplasmic sensing of elevated copper ion concentrations and for induction of a set of genes capable of diminishing copper stress.

  17. Two-component signal transduction system CBO0787/CBO0786 represses transcription from botulinum neurotoxin promoters in Clostridium botulinum ATCC 3502.

    Directory of Open Access Journals (Sweden)

    Zhen Zhang

    2013-03-01

    Full Text Available Blocking neurotransmission, botulinum neurotoxin is the most poisonous biological substance known to mankind. Despite its infamy as the scourge of the food industry, the neurotoxin is increasingly used as a pharmaceutical to treat an expanding range of muscle disorders. Whilst neurotoxin expression by the spore-forming bacterium Clostridium botulinum appears tightly regulated, to date only positive regulatory elements, such as the alternative sigma factor BotR, have been implicated in this control. The identification of negative regulators has proven to be elusive. Here, we show that the two-component signal transduction system CBO0787/CBO0786 negatively regulates botulinum neurotoxin expression. Single insertional inactivation of cbo0787 encoding a sensor histidine kinase, or of cbo0786 encoding a response regulator, resulted in significantly elevated neurotoxin gene expression levels and increased neurotoxin production. Recombinant CBO0786 regulator was shown to bind to the conserved -10 site of the core promoters of the ha and ntnh-botA operons, which encode the toxin structural and accessory proteins. Increasing concentration of CBO0786 inhibited BotR-directed transcription from the ha and ntnh-botA promoters, demonstrating direct transcriptional repression of the ha and ntnh-botA operons by CBO0786. Thus, we propose that CBO0786 represses neurotoxin gene expression by blocking BotR-directed transcription from the neurotoxin promoters. This is the first evidence of a negative regulator controlling botulinum neurotoxin production. Understanding the neurotoxin regulatory mechanisms is a major target of the food and pharmaceutical industries alike.

  18. Learning and evolution in bacterial taxis: an operational amplifier circuit modeling the computational dynamics of the prokaryotic 'two component system' protein network.

    Science.gov (United States)

    Di Paola, Vieri; Marijuán, Pedro C; Lahoz-Beltra, Rafael

    2004-01-01

    Adaptive behavior in unicellular organisms (i.e., bacteria) depends on highly organized networks of proteins governing purposefully the myriad of molecular processes occurring within the cellular system. For instance, bacteria are able to explore the environment within which they develop by utilizing the motility of their flagellar system as well as a sophisticated biochemical navigation system that samples the environmental conditions surrounding the cell, searching for nutrients or moving away from toxic substances or dangerous physical conditions. In this paper we discuss how proteins of the intervening signal transduction network could be modeled as artificial neurons, simulating the dynamical aspects of the bacterial taxis. The model is based on the assumption that, in some important aspects, proteins can be considered as processing elements or McCulloch-Pitts artificial neurons that transfer and process information from the bacterium's membrane surface to the flagellar motor. This simulation of bacterial taxis has been carried out on a hardware realization of a McCulloch-Pitts artificial neuron using an operational amplifier. Based on the behavior of the operational amplifier we produce a model of the interaction between CheY and FliM, elements of the prokaryotic two component system controlling chemotaxis, as well as a simulation of learning and evolution processes in bacterial taxis. On the one side, our simulation results indicate that, computationally, these protein 'switches' are similar to McCulloch-Pitts artificial neurons, suggesting a bridge between evolution and learning in dynamical systems at cellular and molecular levels and the evolutive hardware approach. On the other side, important protein 'tactilizing' properties are not tapped by the model, and this suggests further complexity steps to explore in the approach to biological molecular computing.

  19. Feedback Control of Two-Component Regulatory Systems.

    Science.gov (United States)

    Groisman, Eduardo A

    2016-09-08

    Two-component systems are a dominant form of bacterial signal transduction. The prototypical two-component system consists of a sensor that responds to a specific input(s) by modifying the output of a cognate regulator. Because the output of a two-component system is the amount of phosphorylated regulator, feedback mechanisms may alter the amount of regulator, and/or modify the ability of a sensor or other proteins to alter the phosphorylation state of the regulator. Two-component systems may display intrinsic feedback whereby the amount of phosphorylated regulator changes under constant inducing conditions and without the participation of additional proteins. Feedback control allows a two-component system to achieve particular steady-state levels, to reach a given steady state with distinct dynamics, to express coregulated genes in a given order, and to activate a regulator to different extents, depending on the signal acting on the sensor.

  20. Role of the two component signal transduction system CpxAR in conferring cefepime and chloramphenicol resistance in Klebsiella pneumoniae NTUH-K2044.

    Directory of Open Access Journals (Sweden)

    Vijaya Bharathi Srinivasan

    Full Text Available BACKGROUND: Klebsiella pneumoniae is a gram-negative, non-motile, facultative anaerobe belonging to the Enterobacteriaceae family of the γ-Proteobacteria class in the phylum Proteobacteria. Multidrug resistant K. pneumoniae have caused major therapeutic problems worldwide due to emergence of extended-spectrum β-lactamase producing strains. Two-component systems serve as a basic stimulus-response coupling mechanism to allow organisms to sense and respond to changes in many different environmental conditions including antibiotic stress. PRINCIPAL FINDINGS: In the present study, we investigated the role of an uncharacterized cpxAR operon in bacterial physiology and antimicrobial resistance by generating isogenic mutant (ΔcpxAR deficient in the CpxA/CpxR component derived from the hyper mucoidal K1 strain K. pneumoniae NTUH-K2044. The behaviour of ΔcpxAR was determined under hostile conditions, reproducing stresses encountered in the gastrointestinal environment and deletion resulted in higher sensitivity to bile, osmotic and acid stresses. The ΔcpxAR was more susceptible to β-lactams and chloramphenicol than the wild-type strain, and complementation restored the altered phenotypes. The relative change in expression of acrB, acrD, eefB efflux genes were decreased in cpxAR mutant as evidenced by qRT-PCR. Comparison of outer membrane protein profiles indicated a conspicuous difference in the knock out background. Gel shift assays demonstrated direct binding of CpxR(KP to promoter region of ompC(KP in a concentration dependent manner. CONCLUSIONS AND SIGNIFICANCE: The Cpx envelope stress response system is known to be activated by alterations in pH, membrane composition and misfolded proteins, and this systematic investigation reveals its direct involvement in conferring antimicrobial resistance against clinically significant antibiotics for the very first time. Overall results displayed in this report reflect the pleiotropic role of the Cpx

  1. Characterization of the mrgRS locus of the opportunistic pathogen Burkholderia pseudomallei: temperature regulates the expression of a two-component signal transduction system

    Directory of Open Access Journals (Sweden)

    Dance David AB

    2006-08-01

    Full Text Available Abstract Background Burkholderia pseudomallei is a saprophyte in tropical environments and an opportunistic human pathogen. This versatility requires a sensing mechanism that allows the bacterium to respond rapidly to altered environmental conditions. We characterized a two-component signal transduction locus from B. pseudomallei 204, mrgR and mrgS, encoding products with extensive homology with response regulators and histidine protein kinases of Escherichia coli, Bordetella pertussis, and Vibrio cholerae. Results The locus was present and expressed in a variety of B. pseudomallei human and environmental isolates but was absent from other Burkholderia species, B. cepacia, B. cocovenenans, B. plantarii, B. thailandensis, B. vandii, and B. vietnamiensis. A 2128 bp sequence, including the full response regulator mrgR, but not the sensor kinase mrgS, was present in the B. mallei genome. Restriction fragment length polymorphism downstream from mrgRS showed two distinct groups were present among B. pseudomallei isolates. Our analysis of the open reading frames in this region of the genome revealed that transposase and bacteriophage activity may help explain this variation. MrgR and MrgS proteins were expressed in B. pseudomallei 204 cultured at different pH, salinity and temperatures and the expression was substantially reduced at 25°C compared with 37°C or 42°C but was mostly unaffected by pH or salinity, although at 25°C and 0.15% NaCl a small increase in MrgR expression was observed at pH 5. MrgR was recognized by antibodies in convalescent sera pooled from melioidosis patients. Conclusion The results suggest that mrgRS regulates an adaptive response to temperature that may be essential for pathogenesis, particularly during the initial phases of infection. B. pseudomallei and B. mallei are very closely related species that differ in their capacity to adapt to changing environmental conditions. Modifications in this region of the genome may

  2. Development and validation of a high-throughput cell-based screen to identify activators of a bacterial two-component signal transduction system.

    Science.gov (United States)

    van Rensburg, Julia J; Fortney, Kate R; Chen, Lan; Krieger, Andrew J; Lima, Bruno P; Wolfe, Alan J; Katz, Barry P; Zhang, Zhong-Yin; Spinola, Stanley M

    2015-07-01

    CpxRA is a two-component signal transduction system (2CSTS) found in many drug-resistant Gram-negative bacteria. In response to periplasmic stress, CpxA autophosphorylates and donates a phosphoryl group to its cognate response regulator, CpxR. Phosphorylated CpxR (CpxR-P) upregulates genes involved in membrane repair and downregulates multiple genes that encode virulence factors, which are trafficked across the cell membrane. Mutants that constitutively activate CpxRA in Salmonella enterica serovar Typhimurium and Haemophilus ducreyi are avirulent in mice and humans, respectively. Thus, the activation of CpxRA has high potential as a novel antimicrobial/antivirulence strategy. Using a series of Escherichia coli strains containing a CpxR-P-responsive lacZ reporter and deletions in genes encoding CpxRA system components, we developed and validated a novel cell-based high-throughput screen (HTS) for CpxRA activators. A screen of 36,000 compounds yielded one hit compound that increased reporter activity in wild-type cells. This is the first report of a compound that activates, rather than inhibits, a 2CSTS. The activity profile of the compound against CpxRA pathway mutants in the presence of glucose suggested that the compound inhibits CpxA phosphatase activity. We confirmed that the compound induced the accumulation of CpxR-P in treated cells. Although the hit compound contained a nitro group, a derivative lacking this group retained activity in serum and had lower cytotoxicity than that of the initial hit. This HTS is amenable for the screening of larger libraries to find compounds that activate CpxRA by other mechanisms, and it could be adapted to find activators of other two-component systems.

  3. Sentra, a database of signal transduction proteins.

    Energy Technology Data Exchange (ETDEWEB)

    Maltsev, N.; Marland, E.; Yu, G. X.; Bhatnagar, S.; Lusk, R.; Mathematics and Computer Science

    2002-01-01

    Sentra (http://www-wit.mcs.anl.gov/sentra) is a database of signal transduction proteins with the emphasis on microbial signal transduction. The database was updated to include classes of signal transduction systems modulated by either phosphorylation or methylation reactions such as PAS proteins and serine/threonine kinases, as well as the classical two-component histidine kinases and methyl-accepting chemotaxis proteins. Currently, Sentra contains signal transduction proteins from 43 completely sequenced prokaryotic genomes as well as sequences from SWISS-PROT and TrEMBL. Signal transduction proteins are annotated with information describing conserved domains, paralogous and orthologous sequences, and conserved chromosomal gene clusters. The newly developed user interface supports flexible search capabilities and extensive visualization of the data.

  4. SENTRA, a database of signal transduction proteins.

    Energy Technology Data Exchange (ETDEWEB)

    D' Souza, M.; Romine, M. F.; Maltsev, N.; Mathematics and Computer Science; PNNL

    2000-01-01

    SENTRA, available via URL http://wit.mcs.anl.gov/WIT2/Sentra/, is a database of proteins associated with microbial signal transduction. The database currently includes the classical two-component signal transduction pathway proteins and methyl-accepting chemotaxis proteins, but will be expanded to also include other classes of signal transduction systems that are modulated by phosphorylation or methylation reactions. Although the majority of database entries are from prokaryotic systems, eukaroytic proteins with bacterial-like signal transduction domains are also included. Currently SENTRA contains signal transduction proteins in 34 complete and almost completely sequenced prokaryotic genomes, as well as sequences from 243 organisms available in public databases (SWISS-PROT and EMBL). The analysis was carried out within the framework of the WIT2 system, which is designed and implemented to support genetic sequence analysis and comparative analysis of sequenced genomes.

  5. Equation-free analysis of two-component system signalling model reveals the emergence of co-existing phenotypes in the absence of multistationarity.

    Directory of Open Access Journals (Sweden)

    Rebecca B Hoyle

    Full Text Available Phenotypic differences of genetically identical cells under the same environmental conditions have been attributed to the inherent stochasticity of biochemical processes. Various mechanisms have been suggested, including the existence of alternative steady states in regulatory networks that are reached by means of stochastic fluctuations, long transient excursions from a stable state to an unstable excited state, and the switching on and off of a reaction network according to the availability of a constituent chemical species. Here we analyse a detailed stochastic kinetic model of two-component system signalling in bacteria, and show that alternative phenotypes emerge in the absence of these features. We perform a bifurcation analysis of deterministic reaction rate equations derived from the model, and find that they cannot reproduce the whole range of qualitative responses to external signals demonstrated by direct stochastic simulations. In particular, the mixed mode, where stochastic switching and a graded response are seen simultaneously, is absent. However, probabilistic and equation-free analyses of the stochastic model that calculate stationary states for the mean of an ensemble of stochastic trajectories reveal that slow transcription of either response regulator or histidine kinase leads to the coexistence of an approximate basal solution and a graded response that combine to produce the mixed mode, thus establishing its essential stochastic nature. The same techniques also show that stochasticity results in the observation of an all-or-none bistable response over a much wider range of external signals than would be expected on deterministic grounds. Thus we demonstrate the application of numerical equation-free methods to a detailed biochemical reaction network model, and show that it can provide new insight into the role of stochasticity in the emergence of phenotypic diversity.

  6. Crosstalk of two-component signal transduction systems in regulating central carbohydrate and energy metabolism during autotrophic and photomixotrophic growth of Synechocystis sp. PCC 6803.

    Science.gov (United States)

    Pei, Guangsheng; Niu, Xiangfeng; Zhou, Yuqing; Chen, Lei; Zhang, Weiwen

    2017-05-22

    Unicellular model cyanobacterium Synechocystis sp. PCC 6803 has received considerable attention as a sustainable energy resource because of its photosynthetic machinery. However, two-component signal transduction systems (TCSTSs) in regulating central carbohydrate and energy metabolism of cyanobacteria are still poorly understood due to their diversity and functional complication. In this study, by comparing the growth of knockout mutants of 44 response regulators (RRs) of TCSTSs in Synechocystis, several RR mutants demonstrating differential growth patterns were identified under auto- or photomixotrophic conditions. However, in spite of no growth difference observed for the remaining RR mutants, liquid chromatography-mass spectrometry based metabolomic profile analysis showed that a widespread crosstalk of TCSTSs in regulating central carbohydrate and energy metabolism of Synechocystis was identified, while most of them showed diverse patterns during different trophic types or growth stages. Furthermore, an integrative analysis between evolutionary relationships and metabolomic profiles revealed some pairs of paralogous RRs with highly functional convergence, suggesting the possible conserved functions of Synechocystis TCSTSs during evolution. This study laid an important basis for understanding the function of TCSTSs in photosynthetic cyanobacteria.

  7. A two-component signal-transduction cascade in Carnobacterium piscicola LV17B : two signaling peptides and one sensor-transmitter

    NARCIS (Netherlands)

    Kleerebezem, M; Kuipers, OP; de Vos, WM; Stiles, ME; Quadri, LEN; Vos, Willem M. de; Stiles, Michael E.; Quadri, Luis E.N.

    2001-01-01

    In the lactic acid bacterium Carnobacterium piscicola LV17B a peptide-pheromone dependent quorum-sensing mode is involved in the regulation of bacteriocin production. Bacteriocin CB2 was identified as an environmental signal that induces bacteriocin production. Here, we demonstrate that a second 24

  8. A two-component signal-transduction cascade in Carnobacterium piscicola LV17B : two signaling peptides and one sensor-transmitter

    NARCIS (Netherlands)

    Kleerebezem, M; Kuipers, OP; de Vos, WM; Stiles, ME; Quadri, LEN; Vos, Willem M. de; Stiles, Michael E.; Quadri, Luis E.N.

    2001-01-01

    In the lactic acid bacterium Carnobacterium piscicola LV17B a peptide-pheromone dependent quorum-sensing mode is involved in the regulation of bacteriocin production. Bacteriocin CB2 was identified as an environmental signal that induces bacteriocin production. Here, we demonstrate that a second 24

  9. The Hybrid Histidine Kinase LadS Forms a Multicomponent Signal Transduction System with the GacS/GacA Two-Component System in Pseudomonas aeruginosa

    Science.gov (United States)

    Redelberger, David; Fadel, Firas; Filloux, Alain; Sivaneson, Melissa; de Bentzmann, Sophie; Bordi, Christophe

    2016-01-01

    In response to environmental changes, Pseudomonas aeruginosa is able to switch from a planktonic (free swimming) to a sessile (biofilm) lifestyle. The two-component system (TCS) GacS/GacA activates the production of two small non-coding RNAs, RsmY and RsmZ, but four histidine kinases (HKs), RetS, GacS, LadS and PA1611, are instrumental in this process. RetS hybrid HK blocks GacS unorthodox HK autophosphorylation through the formation of a heterodimer. PA1611 hybrid HK, which is structurally related to GacS, interacts with RetS in P. aeruginosa in a very similar manner to GacS. LadS hybrid HK phenotypically antagonizes the function of RetS by a mechanism that has never been investigated. The four sensors are found in most Pseudomonas species but their characteristics and mode of signaling may differ from one species to another. Here, we demonstrated in P. aeruginosa that LadS controls both rsmY and rsmZ gene expression and that this regulation occurs through the GacS/GacA TCS. We additionally evidenced that in contrast to RetS, LadS signals through GacS/GacA without forming heterodimers, either with GacS or with RetS. Instead, we demonstrated that LadS is involved in a genuine phosphorelay, which requires both transmitter and receiver LadS domains. LadS signaling ultimately requires the alternative histidine-phosphotransfer domain of GacS, which is here used as an Hpt relay by the hybrid kinase. LadS HK thus forms, with the GacS/GacA TCS, a multicomponent signal transduction system with an original phosphorelay cascade, i.e. H1LadS→D1LadS→H2GacS→D2GacA. This highlights an original strategy in which a unique output, i.e. the modulation of sRNA levels, is controlled by a complex multi-sensing network to fine-tune an adapted biofilm and virulence response. PMID:27176226

  10. The Hybrid Histidine Kinase LadS Forms a Multicomponent Signal Transduction System with the GacS/GacA Two-Component System in Pseudomonas aeruginosa.

    Directory of Open Access Journals (Sweden)

    Gaël Chambonnier

    2016-05-01

    Full Text Available In response to environmental changes, Pseudomonas aeruginosa is able to switch from a planktonic (free swimming to a sessile (biofilm lifestyle. The two-component system (TCS GacS/GacA activates the production of two small non-coding RNAs, RsmY and RsmZ, but four histidine kinases (HKs, RetS, GacS, LadS and PA1611, are instrumental in this process. RetS hybrid HK blocks GacS unorthodox HK autophosphorylation through the formation of a heterodimer. PA1611 hybrid HK, which is structurally related to GacS, interacts with RetS in P. aeruginosa in a very similar manner to GacS. LadS hybrid HK phenotypically antagonizes the function of RetS by a mechanism that has never been investigated. The four sensors are found in most Pseudomonas species but their characteristics and mode of signaling may differ from one species to another. Here, we demonstrated in P. aeruginosa that LadS controls both rsmY and rsmZ gene expression and that this regulation occurs through the GacS/GacA TCS. We additionally evidenced that in contrast to RetS, LadS signals through GacS/GacA without forming heterodimers, either with GacS or with RetS. Instead, we demonstrated that LadS is involved in a genuine phosphorelay, which requires both transmitter and receiver LadS domains. LadS signaling ultimately requires the alternative histidine-phosphotransfer domain of GacS, which is here used as an Hpt relay by the hybrid kinase. LadS HK thus forms, with the GacS/GacA TCS, a multicomponent signal transduction system with an original phosphorelay cascade, i.e. H1LadS→D1LadS→H2GacS→D2GacA. This highlights an original strategy in which a unique output, i.e. the modulation of sRNA levels, is controlled by a complex multi-sensing network to fine-tune an adapted biofilm and virulence response.

  11. The Hybrid Histidine Kinase LadS Forms a Multicomponent Signal Transduction System with the GacS/GacA Two-Component System in Pseudomonas aeruginosa.

    Science.gov (United States)

    Chambonnier, Gaël; Roux, Lorène; Redelberger, David; Fadel, Firas; Filloux, Alain; Sivaneson, Melissa; de Bentzmann, Sophie; Bordi, Christophe

    2016-05-01

    In response to environmental changes, Pseudomonas aeruginosa is able to switch from a planktonic (free swimming) to a sessile (biofilm) lifestyle. The two-component system (TCS) GacS/GacA activates the production of two small non-coding RNAs, RsmY and RsmZ, but four histidine kinases (HKs), RetS, GacS, LadS and PA1611, are instrumental in this process. RetS hybrid HK blocks GacS unorthodox HK autophosphorylation through the formation of a heterodimer. PA1611 hybrid HK, which is structurally related to GacS, interacts with RetS in P. aeruginosa in a very similar manner to GacS. LadS hybrid HK phenotypically antagonizes the function of RetS by a mechanism that has never been investigated. The four sensors are found in most Pseudomonas species but their characteristics and mode of signaling may differ from one species to another. Here, we demonstrated in P. aeruginosa that LadS controls both rsmY and rsmZ gene expression and that this regulation occurs through the GacS/GacA TCS. We additionally evidenced that in contrast to RetS, LadS signals through GacS/GacA without forming heterodimers, either with GacS or with RetS. Instead, we demonstrated that LadS is involved in a genuine phosphorelay, which requires both transmitter and receiver LadS domains. LadS signaling ultimately requires the alternative histidine-phosphotransfer domain of GacS, which is here used as an Hpt relay by the hybrid kinase. LadS HK thus forms, with the GacS/GacA TCS, a multicomponent signal transduction system with an original phosphorelay cascade, i.e. H1LadS→D1LadS→H2GacS→D2GacA. This highlights an original strategy in which a unique output, i.e. the modulation of sRNA levels, is controlled by a complex multi-sensing network to fine-tune an adapted biofilm and virulence response.

  12. The essential yhcSR two-component signal transduction system directly regulates the lac and opuCABCD operons of Staphylococcus aureus.

    Directory of Open Access Journals (Sweden)

    Meiying Yan

    Full Text Available Our previous studies suggested that the essential two-component signal transduction system, YhcSR, regulates the opuCABCD operon at the transcriptional level, and the Pspac-driven opuCABCD partially complements the lethal effects of yhcS antisense RNA expression in Staphylococcus aureus. However, the reason why yhcSR regulon is required for growth is still unclear. In this report, we present that the lac and opuC operons are directly transcriptionally regulated by YhcSR. Using real-time RT-PCR we showed that the down-regulation of yhcSR expression affected the transcription of lacA encoding galactose-6-phosphotase isomerase subunit LacA, and opuCA encoding a subunit of a glycine betaine/carnitine/choline ABC transporter. Promoter-lux reporter fusion studies further confirmed the transcriptional regulation of lac by YhcSR. Gel shift assays revealed that YhcR binds to the promoter regions of the lac and opuC operons. Moreover, the Pspac-driven lacABC expression in trans was able to partially complement the lethal effect of induced yhcS antisense RNA. Likewise, the Pspac-driven opuCABCD expression in trans complemented the growth defect of S. aureus in a high osmotic strength medium during the depletion of YhcSR. Taken together, the above data indicate that the yhcSR system directly regulates the expression of lac and opuC operons, which, in turn, may be partially associated with the essentiality of yhcSR in S. aureus. These results provide a new insight into the biological functions of the yhcSR, a global regulator.

  13. The two-component signal transduction system ArlRS regulates Staphylococcus epidermidis biofilm formation in an ica-dependent manner.

    Directory of Open Access Journals (Sweden)

    Yang Wu

    Full Text Available Due to its ability to form biofilms on medical devices, Staphylococcus epidermidis has emerged as a major pathogen of nosocomial infections. In this study, we investigated the role of the two-component signal transduction system ArlRS in regulating S. epidermidis biofilm formation. An ArlRS-deficient mutant, WW06, was constructed using S. epidermidis strain 1457 as a parental strain. Although the growth curve of WW06 was similar to that of SE1457, the mutant strain was unable to form biofilms in vitro. In a rabbit subcutaneous infection model, sterile disks made of polymeric materials were implanted subcutaneously followed with inoculation of WW06 or SE1457. The viable bacteria cells of WW06 recovered from biofilms on the embedded disks were much lower than that of SE1457. Complementation of arlRS genes expression from plasmid in WW06 restored biofilm-forming phenotype both in vivo and in vitro. WW06 maintained the ability to undergo initial attachment. Transcription levels of several genes involved in biofilm formation, including icaADBC, sigB, and sarA, were decreased in WW06, compared to SE1457; and icaR expression was increased in WW06, detected by real-time reverse-transcription PCR. The biofilm-forming phenotype was restored by overexpressing icaADBC in WW06 but not by overexpressing sigB, indicating that ArlRS regulates biofilm formation through the regulation of icaADBC. Gel shift assay showed that ArlR can bind to the promoter region of the ica operon. In conclusion, ArlRS regulates S. epidermidis biofilm formation in an ica-dependent manner, distinct from its role in S. aureus.

  14. The two-component signal transduction system ArlRS regulates Staphylococcus epidermidis biofilm formation in an ica-dependent manner.

    Science.gov (United States)

    Wu, Yang; Wang, Jiaxue; Xu, Tao; Liu, Jingran; Yu, Wenqi; Lou, Qiang; Zhu, Tao; He, Nianan; Ben, Haijing; Hu, Jian; Götz, Friedrich; Qu, Di

    2012-01-01

    Due to its ability to form biofilms on medical devices, Staphylococcus epidermidis has emerged as a major pathogen of nosocomial infections. In this study, we investigated the role of the two-component signal transduction system ArlRS in regulating S. epidermidis biofilm formation. An ArlRS-deficient mutant, WW06, was constructed using S. epidermidis strain 1457 as a parental strain. Although the growth curve of WW06 was similar to that of SE1457, the mutant strain was unable to form biofilms in vitro. In a rabbit subcutaneous infection model, sterile disks made of polymeric materials were implanted subcutaneously followed with inoculation of WW06 or SE1457. The viable bacteria cells of WW06 recovered from biofilms on the embedded disks were much lower than that of SE1457. Complementation of arlRS genes expression from plasmid in WW06 restored biofilm-forming phenotype both in vivo and in vitro. WW06 maintained the ability to undergo initial attachment. Transcription levels of several genes involved in biofilm formation, including icaADBC, sigB, and sarA, were decreased in WW06, compared to SE1457; and icaR expression was increased in WW06, detected by real-time reverse-transcription PCR. The biofilm-forming phenotype was restored by overexpressing icaADBC in WW06 but not by overexpressing sigB, indicating that ArlRS regulates biofilm formation through the regulation of icaADBC. Gel shift assay showed that ArlR can bind to the promoter region of the ica operon. In conclusion, ArlRS regulates S. epidermidis biofilm formation in an ica-dependent manner, distinct from its role in S. aureus.

  15. The essential yhcSR two-component signal transduction system directly regulates the lac and opuCABCD operons of Staphylococcus aureus.

    Science.gov (United States)

    Yan, Meiying; Hall, Jeffrey W; Yang, Junshu; Ji, Yinduo

    2012-01-01

    Our previous studies suggested that the essential two-component signal transduction system, YhcSR, regulates the opuCABCD operon at the transcriptional level, and the Pspac-driven opuCABCD partially complements the lethal effects of yhcS antisense RNA expression in Staphylococcus aureus. However, the reason why yhcSR regulon is required for growth is still unclear. In this report, we present that the lac and opuC operons are directly transcriptionally regulated by YhcSR. Using real-time RT-PCR we showed that the down-regulation of yhcSR expression affected the transcription of lacA encoding galactose-6-phosphotase isomerase subunit LacA, and opuCA encoding a subunit of a glycine betaine/carnitine/choline ABC transporter. Promoter-lux reporter fusion studies further confirmed the transcriptional regulation of lac by YhcSR. Gel shift assays revealed that YhcR binds to the promoter regions of the lac and opuC operons. Moreover, the Pspac-driven lacABC expression in trans was able to partially complement the lethal effect of induced yhcS antisense RNA. Likewise, the Pspac-driven opuCABCD expression in trans complemented the growth defect of S. aureus in a high osmotic strength medium during the depletion of YhcSR. Taken together, the above data indicate that the yhcSR system directly regulates the expression of lac and opuC operons, which, in turn, may be partially associated with the essentiality of yhcSR in S. aureus. These results provide a new insight into the biological functions of the yhcSR, a global regulator.

  16. Role of the CpxAR two-component signal transduction system in control of fosfomycin resistance and carbon substrate uptake.

    Science.gov (United States)

    Kurabayashi, Kumiko; Hirakawa, Yuko; Tanimoto, Koichi; Tomita, Haruyoshi; Hirakawa, Hidetada

    2014-01-01

    Although fosfomycin is an old antibiotic, it has resurfaced with particular interest. The antibiotic is still effective against many pathogens that are resistant to other commonly used antibiotics. We have found that fosfomycin resistance of enterohemorrhagic Escherichia coli (EHEC) O157:H7 is controlled by the bacterial two-component signal transduction system CpxAR. A cpxA mutant lacking its phosphatase activity results in constitutive activation of its cognate response regulator, CpxR, and fosfomycin resistance. We have shown that fosfomycin resistance requires CpxR because deletion of the cpxR gene in the cpxA mutant restores fosfomycin sensitivity. We have also shown that CpxR directly represses the expression of two genes, glpT and uhpT, which encode transporters that cotransport fosfomycin with their native substrates glycerol-3-phosphate and glucose-6-phosphate, and repression of these genes leads to a decrease in fosfomycin transport into the cpxA mutant. However, the cpxA mutant had an impaired growth phenotype when cultured with glycerol-3-phosphate or glucose-6-phosphate as a sole carbon substrate and was outcompeted by the parent strain, even in nutrient-rich medium. This suggests a trade-off between fosfomycin resistance and the biological fitness associated with carbon substrate uptake. We propose a role for the CpxAR system in the reversible control of fosfomycin resistance. This may be a beneficial strategy for bacteria to relieve the fitness burden that results from fosfomycin resistance in the absence of fosfomycin.

  17. Perturbation of the two-component signal transduction system, BprRS, results in attenuated virulence and motility defects in Burkholderia pseudomallei.

    Science.gov (United States)

    Lazar Adler, Natalie R; Allwood, Elizabeth M; Deveson Lucas, Deanna; Harrison, Paul; Watts, Stephen; Dimitropoulos, Alexandra; Treerat, Puthayalai; Alwis, Priyangi; Devenish, Rodney J; Prescott, Mark; Govan, Brenda; Adler, Ben; Harper, Marina; Boyce, John D

    2016-05-04

    Burkholderia pseudomallei is the causative agent of melioidosis, a severe invasive disease of humans and animals. Initial screening of a B. pseudomallei signature-tagged mutagenesis library identified an attenuated mutant with a transposon insertion in a gene encoding the sensor component of an uncharacterised two-component signal transduction system (TCSTS), which we designated BprRS. Single gene inactivation of either the response regulator gene (bprR) or the sensor histidine kinase gene (bprS) resulted in mutants with reduced swarming motility and reduced virulence in mice. However, a bprRS double mutant was not attenuated for virulence and displayed wild-type levels of motility. The transcriptomes of the bprS, bprR and bprRS mutants were compared with the transcriptome of the parent strain K96243. Inactivation of the entire BprRS TCSTS (bprRS double mutant) resulted in altered expression of only nine genes, including both bprR and bprS, five phage-related genes and bpss0686, encoding a putative 5, 10-methylene tetrahydromethanopterin reductase involved in one carbon metabolism. In contrast, the transcriptomes of each of the bprR and bprS single gene mutants revealed more than 70 differentially expressed genes common to both mutants, including regulatory genes and those required for flagella assembly and for the biosynthesis of the cytotoxic polyketide, malleilactone. Inactivation of the entire BprRS TCSTS did not alter virulence or motility and very few genes were differentially expressed indicating that the definitive BprRS regulon is relatively small. However, loss of a single component, either the sensor histidine kinase BprS or its cognate response regulator BprR, resulted in significant transcriptomic and phenotypic differences from the wild-type strain. We hypothesize that the dramatically altered phenotypes of these single mutants are the result of cross-regulation with one or more other TCSTSs and concomitant dysregulation of other key regulatory genes.

  18. Preliminary Crystallographic Studies of the Regulatory Domain of Response Regulator YycF from an Essential Two-Component Signal Transduction System

    Energy Technology Data Exchange (ETDEWEB)

    Zhao, H.; Heroux, A; Sequeira, R; Tang, L

    2009-01-01

    YycGF is a crucial signal transduction system for the regulation of cell-wall metabolism in low-G+C Gram-positive bacteria, which include many important human pathogens. The response regulator YycF receives signals from its cognate histidine kinase YycG through a phosphotransfer reaction and elicits responses through regulation of gene expression. The N-terminal regulatory domain of YycF from Bacillus subtilis was overproduced and purified. The protein was crystallized and X-ray data were collected to 1.95 A resolution with a completeness of 97.7% and an overall Rmerge of 7.7%. The crystals belonged to space group P3121, with unit-cell parameters a = b = 59.50, c = 79.06 A.

  19. A two-component signal-transducing system is involved in competence and penicillin susceptibility in laboratory mutants of Streptococcus pneumoniae.

    Science.gov (United States)

    Guenzi, E; Gasc, A M; Sicard, M A; Hakenbeck, R

    1994-05-01

    Penicillin resistance in Streptococcus pneumoniae has been attributed so far to the production of penicillin-binding protein (PBP) variants with decreased affinities for beta-lactam antibiotics. Cefotaxime-resistant laboratory mutants, selected after several steps on increasing concentrations of this beta-lactam, become deficient in transformation as well. A DNA fragment conferring both cefotaxime resistance and transformation deficiency was isolated and cloned from the mutant C306. The cefotaxime resistance associated with this resistance determinant was not accompanied with apparent changes in PBP properties, and it mapped on the chromosome distinct from the known resistance determinants, genes encoding PBP2x, PBP1a or PBP2b. Determination of a 2265 bp DNA sequence of the resistance determinant revealed two open reading frames, ciaR and ciaH, whose deduced amino acid sequence identified the corresponding proteins as the response regulator and histidine kinase receptor, respectively (members of the two families of bacterial signal-transducing proteins). Two hydrophobic peptide regions divided the histidine kinase CiaH into two putative domains: an N-terminal extracellular sensor part, and an intracellular C-terminal domain with the conserved His-226 residue, the presumed phosphorylation site. The single point mutations responsible for cefotaxime-resistance and transformation deficiency of C306 and of another two independently isolated cefotaxime-resistant mutants were each located in the C-terminal half of CiaH. A small extracellular protein, the competence factor, is required for induction of competence. Neither C306 nor the transformants obtained with the mutated ciaH gene produced competence factor, and exogenous competence factor could not complement the transformation deficiency, indicating that the signal-transducing system cia is involved in early steps of competence regulation.

  20. Bioinformatics analysis of two-component signal transduction systems of Bacillus thuringiensis%苏云金芽孢杆菌双组份信号转导系统的生物信息学分析

    Institute of Scientific and Technical Information of China (English)

    张清仪; 王阶平; 程钢; 刘钟慧; 范文瑾; 何进

    2011-01-01

    苏云金芽孢杆菌(Bacillus thuringiensis)能产生杀虫晶体蛋白等多种活性成分,是目前应用最广泛的微生物杀虫剂.本文采用生物信息学方法,系统分析了由本实验室完成全基因组测序的苏云全芽孢杆菌YBT-1520、CT-43和BMB171 3个菌株的双组分信号转导系统(Two-component signal traducction system,TCS)的分布、结构及功能,并初步构建了部分TCS的调控网络关系图.本研究旨在为深入研究苏云金芽孢杆菌的生长、代谢以及毒力因子的表达与调控,全面了解伴孢晶体的形成机制开辟新的研究方向.%Bacillus thuringiensis (Bt) strains, which can produce insecticidal crystal proteins, were widely used as biological pesticides. In this paper, we comprehensively analyzed the distributions, structures and putative biological functions of two-component transduction systems (TCS) from the genomes of Bt strains YBT-1520, CT-43 and BMB171, which have been sequenced by our laboratory. And more importantly, we constructed a preliminary TCS regulatory networks. This study should open a novel research direction in Bt for the growth, metabolism, regulator of toxic gene expression, as well as the formation mechanism of parasporal crystals.

  1. A two-component micelle with emergent pH responsiveness by mixing dilauroyl phosphocholine and deoxycholic acid and its delivery of proteins into the cytosol.

    Science.gov (United States)

    Miyamoto, Noriko; Fujii, Shota; Mochizuki, Shinichi; Sakurai, Kazuo; Sakaguchi, Naoki; Koiwai, Kazunori

    2017-06-01

    Providing appropriate pH responsiveness for drug delivery nanoparticles is one of the major issues in developing a new generation of delivery systems. This paper reports that, when phosphocholine and a bile acid were mixed, the resultant two-component micelle gained pH responsiveness, while the individual components did not show any such responsiveness. The pH responsiveness was shown to be determined by the chemical structure, especially the positions and chirality of the OH groups, of the bile acid, and the sensitivity was determined by the alkyl chain length of the phosphocholine. The best combination for evading endocytosis was dilauroyl phosphocholine (DLPC) and deoxycholic acid (DA). Small-angle X-ray scattering revealed that the pH responsiveness was related to the change of surface hydrophobicity, namely, decreasing pH led to protonation of the carboxylic acid, resulting in aggregation of the preceding micelles. We assume that particles that become hydrophobic in this way can start interacting with the endocytotic bilayer, which eventually leads to rupture of the endocytotic vesicle. This mechanism is well supported by the finding that fluorescein-conjugated ovalbumin proteins were transported into the cytosol when they were co-administered with DLPC/DA. Copyright © 2017 Elsevier B.V. All rights reserved.

  2. Interaction Analysis of a Two-Component System Using Nanodiscs.

    Directory of Open Access Journals (Sweden)

    Patrick Hörnschemeyer

    Full Text Available Two-component systems are the major means by which bacteria couple adaptation to environmental changes. All utilize a phosphorylation cascade from a histidine kinase to a response regulator, and some also employ an accessory protein. The system-wide signaling fidelity of two-component systems is based on preferential binding between the signaling proteins. However, information on the interaction kinetics between membrane embedded histidine kinase and its partner proteins is lacking. Here, we report the first analysis of the interactions between the full-length membrane-bound histidine kinase CpxA, which was reconstituted in nanodiscs, and its cognate response regulator CpxR and accessory protein CpxP. Using surface plasmon resonance spectroscopy in combination with interaction map analysis, the affinity of membrane-embedded CpxA for CpxR was quantified, and found to increase by tenfold in the presence of ATP, suggesting that a considerable portion of phosphorylated CpxR might be stably associated with CpxA in vivo. Using microscale thermophoresis, the affinity between CpxA in nanodiscs and CpxP was determined to be substantially lower than that between CpxA and CpxR. Taken together, the quantitative interaction data extend our understanding of the signal transduction mechanism used by two-component systems.

  3. 隐藏嗜酸菌Acidiphilium cryptum JF-5双组分信号转导系统%Two-component signal transduction system of Acidiphilium cryptum JF-5

    Institute of Scientific and Technical Information of China (English)

    余水静; 彭艳平; 邓扬悟; 郭燕华; 刘荷英

    2013-01-01

      为了探索隐藏嗜酸菌(Acidiphilium cryptum)对多变极端矿山环境条件的感知和反应分子机制,预测和分析了隐藏嗜酸菌JF-5菌株的双组分信号转导系统(Two-component signal transduction system, TCS)的分布、结构及功能。鉴定了9对成对TCSs、2个杂合结构TCSs、3个孤儿组氨酸蛋白激酶(HK)和5个孤儿反应调节蛋白(RR);发现5个TCSs参与隐藏嗜酸菌对重金属响应转录调控;大多数HKs的N-末端具有接受信号的跨膜区、HAMP或PAS等结构域,RRs主要是OmpR亚家族,占总RRs的40%以上;从进化关系上来看,一些处在进化树同一分支上的共同聚簇TCS基因可能具有相同的进化途径。本研究结果可为研究隐藏嗜酸菌在极端环境中适应性分子机制提供新的方向。%In order to explore the molecular mechanism of Acidiphilium cryptum response to extreme conditions in the mine environment, the distributions, structures and putative biological functions of two-component signal transduction systems (TCSs) in A. cryptum JF-5 are predicated and analyzed. And 9 TCSs, 2 hybrid histidine kinases (HK), 3 orphan HKs and 5 orphan response regulator proteins (RR) were identified. Five TCSs were involved in the transcription regulation of A. cryptum response to heavy metal. The most N-terminal regions of HKs were characterized by the presence of transmembrane helices, HAMPs, or PAS domains. More than 40 percent of putative RRs were classified into OmpR subfamily. From the evolutionary relationship, the common clustering TCS genes in the same branch of the evolutionary tree may have the same evolutionary pathway. Our results should open a novel research direction in A. cryptum for molecular mechanism of the adaptation in the extreme environment.

  4. Structural characterization of the heme-based oxygen sensor, AfGcHK, its interactions with the cognate response regulator, and their combined mechanism of action in a bacterial two-component signaling system.

    Science.gov (United States)

    Stranava, Martin; Martínek, Václav; Man, Petr; Fojtikova, Veronika; Kavan, Daniel; Vaněk, Ondřej; Shimizu, Toru; Martinkova, Marketa

    2016-10-01

    The oxygen sensor histidine kinase AfGcHK from the bacterium Anaeromyxobacter sp. Fw 109-5 forms a two-component signal transduction system together with its cognate response regulator (RR). The binding of oxygen to the heme iron of its N-terminal sensor domain causes the C-terminal kinase domain of AfGcHK to autophosphorylate at His183 and then transfer this phosphate to Asp52 or Asp169 of the RR protein. Analytical ultracentrifugation revealed that AfGcHK and the RR protein form a complex with 2:1 stoichiometry. Hydrogen-deuterium exchange coupled to mass spectrometry (HDX-MS) suggested that the most flexible part of the whole AfGcHK protein is a loop that connects the two domains and that the heme distal side of AfGcHK, which is responsible for oxygen binding, is the only flexible part of the sensor domain. HDX-MS studies on the AfGcHK:RR complex also showed that the N-side of the H9 helix in the dimerization domain of the AfGcHK kinase domain interacts with the helix H1 and the β-strand B2 area of the RR protein's Rec1 domain, and that the C-side of the H8 helix region in the dimerization domain of the AfGcHK protein interacts mostly with the helix H5 and β-strand B6 area of the Rec1 domain. The Rec1 domain containing the phosphorylable Asp52 of the RR protein probably has a significantly higher affinity for AfGcHK than the Rec2 domain. We speculate that phosphorylation at Asp52 changes the overall structure of RR such that the Rec2 area containing the second phosphorylation site (Asp169) can also interact with AfGcHK. Proteins 2016; 84:1375-1389. © 2016 Wiley Periodicals, Inc. © 2016 Wiley Periodicals, Inc.

  5. Systematic mutation analysis of two-component signal transduction systems reveals EsrA-EsrB and PhoP-PhoQ as the major virulence regulators in Edwardsiella tarda.

    Science.gov (United States)

    Lv, Yuanzhi; Xiao, Jingfan; Liu, Qin; Wu, Haizhen; Zhang, Yuanxing; Wang, Qiyao

    2012-05-25

    Edwardsiella tarda is a Gram-negative broad-host-range pathogen that causes hemorrhagic septicemia in many commercially important fish species. Its ability to adapt to and thrive in diverse environments outside and inside of its hosts prompts us to investigate the roles of the previously identified 33 putative two-component signal transduction systems (TCSs) in E. tarda. In this work, we successfully constructed deletion mutations in each of the response regulator genes, suggesting that none of the TCSs are essential for cell viability in E. tarda. The mutants were further examined for roles in biofilm formation, antibiotic resistance, stress response, expression and secretion of proteins involved in either the type III secretion system (T3SS) or type VI secretion system (T6SS), as well as virulence. Through these assays, we identified four regulators of biofilm development, two regulators of antibiotic resistance, and four regulators involved in stress responses. We found that two regulators, EsrB and PhoP, are essential for the pathogenicity of E. tarda and further demonstrated that these two regulators have codependent and independent contributions to E. tarda virulence. Mutation of EsrB resulted in the complete loss of both the T3SS and T6SS proteins, while PhoP partially regulated the expression of T3SS and T6SS genes through EsrB, and was essential for resistance to antimicrobial peptides. This work suggested that these two response regulators are involved in the regulation of the complex virulence network of this bacterium and merit as candidate genes for live attenuated vaccine construction. Copyright © 2011 Elsevier B.V. All rights reserved.

  6. Relationship of the CreBC two-component regulatory system and inner membrane protein CreD with swimming motility in Stenotrophomonas maltophilia.

    Science.gov (United States)

    Huang, Hsin-Hui; Chen, Wei-Ching; Lin, Cheng-Wen; Lin, Yi-Tsung; Ning, Hsiao-Chen; Chang, Yi-Chih; Yang, Tsuey-Ching

    2017-01-01

    The CreBC two-component system (TCS) is a conserved regulatory system found in Escherichia coli, Aeromonas spp., Pseudomonas aeruginosa, and Stenotrophomonas maltophilia. In this study, we determined how CreBC TCS regulates secreted protease activities and swimming motility using creB, creC, and creBC in-frame deletion mutants (KJΔCreB, KJΔCreC, and KJΔBC) of S. maltophilia KJ. Compared to wild-type KJ, KJΔCreB had a comparable secreted protease activity; however, the secreted protease activities were obviously reduced in KJΔCreC and KJΔBC, suggesting that CreC works together with another unidentified response regulator (not CreB) to regulate secreted protease activity. Single gene inactivation of creB or creC resulted in mutants with an enhanced swimming motility, and this phenotype was exacerbated in a double mutant KJΔBC. To elucidate the underlying mechanism responsible for the ΔcreBC-mediated swimming enhancement, flagella morphology observation, RNA-seq based transcriptome assay, qRT-PCR, and membrane integrity and potential assessment were performed. Flagella morphological observation ruled out the possibility that swimming enhancement was due to altered flagella morphology. CreBC inactivation upregulated the expression of creD and flagella-associated genes encoding the basal body- and motor-associated proteins. Furthermore, KJΔBC had an increased membrane susceptibility to Triton X-100 and CreD upregulation in KJΔBC partially alleviated the compromise of membrane integrity. The impact of creBC TCS on bacterial membrane potential was assessed by carbonyl cyanide m-chlorophenyl hydrazine (CCCP50) concentration at which 50% of bacterial swimming is inhibited. CCCP50 of wild-type KJ increased when creBC was deleted, indicating an association between the higher membrane potential of KJΔBC cells and enhanced motility. Upregulation of the basal body- and motor-associated genes of flagella in KJΔBC cells may explain the increased membrane potential

  7. PhoP-PhoR two-component signal transduction systems in pathogenesis of Mycobacterium tuberculosis%PhoPR双组份系统在结核分枝杆菌致病机制中的研究进展

    Institute of Scientific and Technical Information of China (English)

    邬博; 张万江

    2014-01-01

    Two-component signal transduction system (TCS) exists extensively in prokaryotic cell ,which plays a key role of regulation in the growth ,differentiation ,metabolism ,virulence ,persistence ,and pathogenicity .PhoPR two-component signal transduction system ,as one of two-component systems ,is the most basic and the most importantly sensitive to the envi-ronmental changes and makes corresponding certain reaction system adapt to changes in the host microenvironment .Therefore , PhoPR TCS which is an important regulatory system of Mycobacterium tuberculosis (MTB) to adapt to environmental change has been increasing concern ,even PhoPR TCS is becoming the new front-burner issue in the pathogenesis of M TB regulation of research .%双组份信号转导系统(Two-component signal transduction system ,TCS)广泛存在于原核生物细胞内,对细胞生长、分化、代谢、毒力、持留性、致病性等方面的调控发挥着重要作用。PhoPR TCS作为双组份系统中最基本、最重要的感应外界环境变化,并作出相应反应的调控系统,能够调控结核分枝杆菌(Mycobacterium Tuberculosis ,MTB)更好的适应宿主微环境变化。因此PhoPR TCS作为M TB适应环境变化的重要调控系统已愈来愈受关注,关于PhoPR TCS在M TB的致病机制调控方面的研究正在成为新的热点。

  8. Cytokinin Regulates the Etioplast-Chloroplast Transition through the Two-Component Signaling System and Activation of Chloroplast-Related Genes1

    Science.gov (United States)

    Cortleven, Anne; Marg, Ingke; Schlicke, Hagen; Hill, Kristine; Schmülling, Thomas

    2016-01-01

    One of the classical functions of the plant hormone cytokinin is the regulation of plastid development, but the underlying molecular mechanisms remain elusive. In this study, we employed a genetic approach to evaluate the role of cytokinin and its signaling pathway in the light-induced development of chloroplasts from etioplasts in Arabidopsis (Arabidopsis thaliana). Cytokinin increases the rate of greening and stimulates ultrastructural changes characteristic for the etioplast-to-chloroplast transition. The steady-state levels of metabolites of the tetrapyrrole biosynthesis pathway leading to the production of chlorophyll are enhanced by cytokinin. This effect of cytokinin on metabolite levels arises due to the modulation of expression for chlorophyll biosynthesis genes such as HEMA1, GUN4, GUN5, and CHLM. Increased expression of HEMA1 is reflected in an enhanced level of the encoded glutamyl-tRNA reductase, which catalyzes one of the rate-limiting steps of chlorophyll biosynthesis. Mutant analysis indicates that the cytokinin receptors ARABIDOPSIS HIS KINASE2 (AHK2) and AHK3 play a central role in this process. Furthermore, the B-type ARABIDOPSIS RESPONSE REGULATOR1 (ARR1), ARR10, and ARR12 play an important role in mediating the transcriptional output during etioplast-chloroplast transition. B-type ARRs bind to the promotors of HEMA1 and LHCB6 genes, indicating that cytokinin-dependent transcription factors directly regulate genes of chlorophyll biosynthesis and the light harvesting complex. Together, these results demonstrate an important role for the cytokinin signaling pathway in chloroplast development, with the direct transcriptional regulation of chlorophyll biosynthesis genes as a key aspect for this hormonal control. PMID:27388681

  9. Cytokinin Regulates the Etioplast-Chloroplast Transition through the Two-Component Signaling System and Activation of Chloroplast-Related Genes.

    Science.gov (United States)

    Cortleven, Anne; Marg, Ingke; Yamburenko, Maria V; Schlicke, Hagen; Hill, Kristine; Grimm, Bernhard; Schaller, G Eric; Schmülling, Thomas

    2016-09-01

    One of the classical functions of the plant hormone cytokinin is the regulation of plastid development, but the underlying molecular mechanisms remain elusive. In this study, we employed a genetic approach to evaluate the role of cytokinin and its signaling pathway in the light-induced development of chloroplasts from etioplasts in Arabidopsis (Arabidopsis thaliana). Cytokinin increases the rate of greening and stimulates ultrastructural changes characteristic for the etioplast-to-chloroplast transition. The steady-state levels of metabolites of the tetrapyrrole biosynthesis pathway leading to the production of chlorophyll are enhanced by cytokinin. This effect of cytokinin on metabolite levels arises due to the modulation of expression for chlorophyll biosynthesis genes such as HEMA1, GUN4, GUN5, and CHLM Increased expression of HEMA1 is reflected in an enhanced level of the encoded glutamyl-tRNA reductase, which catalyzes one of the rate-limiting steps of chlorophyll biosynthesis. Mutant analysis indicates that the cytokinin receptors ARABIDOPSIS HIS KINASE2 (AHK2) and AHK3 play a central role in this process. Furthermore, the B-type ARABIDOPSIS RESPONSE REGULATOR1 (ARR1), ARR10, and ARR12 play an important role in mediating the transcriptional output during etioplast-chloroplast transition. B-type ARRs bind to the promotors of HEMA1 and LHCB6 genes, indicating that cytokinin-dependent transcription factors directly regulate genes of chlorophyll biosynthesis and the light harvesting complex. Together, these results demonstrate an important role for the cytokinin signaling pathway in chloroplast development, with the direct transcriptional regulation of chlorophyll biosynthesis genes as a key aspect for this hormonal control. © 2016 American Society of Plant Biologists. All rights reserved.

  10. Effect of histamine on the signal transduction of the AtoS-AtoC two component system and involvement in poly-(R)-3-hydroxybutyrate biosynthesis in Escherichia coli.

    Science.gov (United States)

    Kyriakidis, D A; Theodorou, M C; Filippou, P S; Kyriakidis, K D; Tiligada, E

    2008-06-01

    AtoS-AtoC two-component system acts directly on the atoDAEB operon transcription to regulate the biosynthesis of short-chain poly-(R)-3-hydroxybutyrate. This study sought to investigate the effect of histamine and compound 48/80 on the regulation of AtoS-AtoC two-component system in Escherichia coli K-12 MA255 (speC(-), speB(-)) and the isogenic E. coli strains BW25113 (atoSC(+)) and BW28878 (DeltaatoSC) transformed with plasmids carrying related genes. Histamine or compound 48/80 induced or tended to reduce atoC transcription, respectively, while neither compound showed any effect on atoDAEB operon transcription. Moreover, histamine down-regulated poly-(R)-3-hydroxybutyrate biosynthesis, whereas compound 48/80 up-regulated its biosynthesis, maximal induction being obtained in the presence of multiple copies of AtoS-AtoC. Interestingly, co-administration of histamine counteracted this inductive effect of compound 48/80. The reported data provide the first evidence for a differential modulator role of histamine and compound 48/80 on the AtoS-AtoC two-component system signaling in potentially pathogenic bacteria, leading to a new perspective on their symbiotic behavior.

  11. Two-Component Signaling System VgrRS Directly Senses Extracytoplasmic and Intracellular Iron to Control Bacterial Adaptation under Iron Depleted Stress.

    Directory of Open Access Journals (Sweden)

    Li Wang

    2016-12-01

    Full Text Available Both iron starvation and excess are detrimental to cellular life, especially for animal and plant pathogens since they always live in iron-limited environments produced by host immune responses. However, how organisms sense and respond to iron is incompletely understood. Herein, we reveal that in the phytopathogenic bacterium Xanthomonas campestris pv. campestris, VgrS (also named ColS is a membrane-bound receptor histidine kinase that senses extracytoplasmic iron limitation in the periplasm, while its cognate response regulator, VgrR (ColR, detects intracellular iron excess. Under iron-depleted conditions, dissociation of Fe3+ from the periplasmic sensor region of VgrS activates the VgrS autophosphorylation and subsequent phosphotransfer to VgrR, an OmpR-family transcription factor that regulates bacterial responses to take up iron. VgrR-VgrS regulon and the consensus DNA binding motif of the transcription factor VgrR were dissected by comparative proteomic and ChIP-seq analyses, which revealed that in reacting to iron-depleted environments, VgrR directly or indirectly controls the expressions of hundreds of genes that are involved in various physiological cascades, especially those associated with iron-uptake. Among them, we demonstrated that the phosphorylated VgrR tightly represses the transcription of a special TonB-dependent receptor gene, tdvA. This regulation is a critical prerequisite for efficient iron uptake and bacterial virulence since activation of tdvA transcription is detrimental to these processes. When the intracellular iron accumulates, the VgrR-Fe2+ interaction dissociates not only the binding between VgrR and the tdvA promoter, but also the interaction between VgrR and VgrS. This relieves the repression in tdvA transcription to impede continuous iron uptake and avoids possible toxic effects of excessive iron accumulation. Our results revealed a signaling system that directly senses both extracytoplasmic and intracellular

  12. Comparative analysis of wolbachia genomes reveals streamlining and divergence of minimalist two-component systems.

    Science.gov (United States)

    Christensen, Steen; Serbus, Laura Renee

    2015-03-24

    Two-component regulatory systems are commonly used by bacteria to coordinate intracellular responses with environmental cues. These systems are composed of functional protein pairs consisting of a sensor histidine kinase and cognate response regulator. In contrast to the well-studied Caulobacter crescentus system, which carries dozens of these pairs, the streamlined bacterial endosymbiont Wolbachia pipientis encodes only two pairs: CckA/CtrA and PleC/PleD. Here, we used bioinformatic tools to compare characterized two-component system relays from C. crescentus, the related Anaplasmataceae species Anaplasma phagocytophilum and Ehrlichia chaffeensis, and 12 sequenced Wolbachia strains. We found the core protein pairs and a subset of interacting partners to be highly conserved within Wolbachia and these other Anaplasmataceae. Genes involved in two-component signaling were positioned differently within the various Wolbachia genomes, whereas the local context of each gene was conserved. Unlike Anaplasma and Ehrlichia, Wolbachia two-component genes were more consistently found clustered with metabolic genes. The domain architecture and key functional residues standard for two-component system proteins were well-conserved in Wolbachia, although residues that specify cognate pairing diverged substantially from other Anaplasmataceae. These findings indicate that Wolbachia two-component signaling pairs share considerable functional overlap with other α-proteobacterial systems, whereas their divergence suggests the potential for regulatory differences and cross-talk.

  13. Immunological evaluation in nonhuman primates of formulations based on the chimeric protein P64k-domain III of dengue 2 and two components of Neisseria meningitidis.

    Science.gov (United States)

    Valdés, Iris; Hermida, Lisset; Martín, Jorge; Menéndez, Tamara; Gil, Lázaro; Lazo, Laura; Castro, Jorge; Niebla, Olivia; López, Carlos; Bernardo, Lídice; Sánchez, Jorge; Romero, Yaremis; Martínez, Rafael; Guzmán, María G; Guillén, Gerardo

    2009-02-11

    The main problem in the development of successful vaccines against dengue based on recombinant proteins is the necessity to use potent adjuvants to reach a proper functional immune response. Our group reported the expression, characterization and immunological evaluation of the recombinant protein PD5, which contains the domain III of the Envelope protein from dengue 2 virus fused to the carrier protein P64k. This construct completely protected monkeys against viral challenge when the Freund's adjuvant was employed. Therefore, to define suitable formulations for human use, the present work relies on the evaluation of PD5, produced with a high purity and under GMP conditions, when formulated either with outer membrane vesicles (OMV) or the serogroup A capsular polysaccharide (CPS-A) from Neisseria meningitidis, both adsorbed on aluminium hydroxide. The antibody response to the formulation containing the CPS-A was clearly superior to that of the formulation with OMV. The experiment of in vivo protection supported this evidence, since only the group immunized with PD5 and CPS-A was partially protected upon viral challenge. This is the first study in which the polysaccharide A of N. meningitidis is successfully employed as adjuvant for viral antigens.

  14. The auxiliary protein complex SaePQ activates the phosphatase activity of sensor kinase SaeS in the SaeRS two-component system of Staphylococcus aureus

    OpenAIRE

    Jeong, Do-Won; Cho, Hoonsik; Jones, Marcus B.; Shatzkes, Kenneth; Sun, Fei; Ji, Quanjiang; Liu, Qian; Scott N Peterson; He, Chuan; Bae, Taeok

    2012-01-01

    In bacterial two-component regulatory systems (TCSs), dephosphorylation of phosphorylated response regulators is essential for resetting the activated systems to the pre-activation state. However, in the SaeRS TCS, a major virulence TCS of Staphylococcus aureus, the mechanism for dephosphorylation of the response regulator SaeR has not been identified. Here we report that two auxiliary proteins from the sae operon, SaeP and SaeQ, form a protein complex with the sensor kinase SaeS and activate...

  15. Progression on the research of two-component signal transduction system in fungus and its inhibitors%真菌双组分信号转导系统及其抑制剂研究进展

    Institute of Scientific and Technical Information of China (English)

    徐西光; 张子平; 程波

    2011-01-01

    双组分信号转导系统存在于包括真菌在内的大部分低等真核生物、原核生物及一些植物中.真菌双组分信号转导蛋白在细胞新陈代谢、毒力以及致病性等方面具有重要作用,且目前在人类细胞中尚未发现双组分信号转导系统.因此,探明真菌双组分信号转导系统的机制,可为抑制剂的设计和寻找提供多个“靶点”,从而研制出能够抗致病性真菌而不对宿主细胞造成损伤的新型抗真菌药物.本文就近年来真菌双组分信号转导系统及其潜在抑制剂进行综述.%Two-component signal transduction system, which plays an important role in cell metabolism, virulence and pathoge-nicity, has been found in most lower eukaryotes, prokaryotes and some plants, yet not in human cells. Well-understanding of the mechanism may be helpful for inhibitor designing, which has antifungal effect without damage to host cell. Recent literatures about two-component signal transduction system in fungi and potential inhibitors are reviewed.

  16. Signal transduction meets systems biology: deciphering specificity determinants for protein-protein interactions

    Science.gov (United States)

    Bourret, Robert B.

    2008-01-01

    Two recent papers [Gao et al. Mol. Microbiol. 69, 1358 (2008); Skerker et al. Cell 133, 1043 (2008)] describe investigations into the specificity of protein-protein interactions that occur during signal transduction by two-component regulatory systems. This MicroCommentary summarizes and provides context for the reported findings. The results offer insights into molecular determinants that provide specificity to maintain signal separation and thus prevent deleterious crosstalk between pathways, as well as the potential extent and nature of interactions that may combine signals to achieve beneficial cross regulation among pathways. The methods employed are suitable for application to other systems. PMID:18694439

  17. Stochastic study of information transmission and population stability in a generic bacterial two-component system

    CERN Document Server

    Mapder, Tarunendu; Banik, Suman K

    2016-01-01

    Studies on the role of fluctuations in signal propagation and on gene regulation in monoclonal bacterial population have been extensively pursued based on the machinery of two-component system. The bacterial two-component system shows noise utilisation through its inherent plasticity. The fluctuations propagation takes place using the phosphotransfer module and the feedback mechanism during gene regulation. To delicately observe the noisy kinetics the generic cascade needs stochastic investigation at the mRNA and protein levels. To this end, we propose a theoretical framework to investigate the noisy signal transduction in a generic two-component system. The model shows reliability in information transmission through quantification of several statistical measures. We further extend our analysis to observe the protein distribution in a population of cells. Through numerical simulation, we identify the regime of the kinetic parameter set that generates a stability switch in the steady state distribution of prot...

  18. Two-component signal transduction system and the virulence-related characteristics of Streptococcus mutans%双组分信号传导系统与变异链球菌的致病相关特性

    Institute of Scientific and Technical Information of China (English)

    陈娇

    2011-01-01

    Two-component signal transduction system (TCSTS) plays an important role in the expression of virulence factors of Streptococcus mutans, such as the competence development, the formation of biofilm, the stability of structure, the production of bacteriocin, and the properties of acid production and resistance. It provides many ecological advantages to compete and survive in the dental plaque. This article is a review about the composition of TCSTS of Streptococcus mutans, the effect of TCSTS on the virulence-related characteristics of Streptococcus mutans, and the application significance of TCSTS.%双组分信号传导系统(TCSTS)在变异链球菌感受态形成、生物膜形成、结构稳定、菌素产生、产酸耐酸特性等毒力因子表达方面起着重要的作用,为其在菌斑生物膜中的竞争和生存提供了诸多生态性优势.本文就变异链球菌TCSTS的组成、TCSTS对变异链球菌致病相关特性的影响、TCSTS的应用意义等作一综述.

  19. VicRK two-component signal transduction system of Streptococcus mutans%变异链球菌的VicRK双组分信号传导系统

    Institute of Scientific and Technical Information of China (English)

    田媛媛

    2012-01-01

    Streptococcus mutans(S.nuaans), which is considered as the chief pathogen of human caries, possesses the ability to form biofilm via sucrose-dependent adhesion, genesis and endure acids in the biofilm that may ultimately lead to dental caries. VicRK is one of the 13 putative two-component signal transduction systems of S.mutans that modulate the expression of cariogeneisis related virulence factors. This review summarized the mechanism, structural organization, physiological characteristics and the impact on the cariogenesis capabilities of VicRK, as well as the correlation between VicRK and VicX in S.mutans.%变异链球菌是人类龋病的主要病原菌,它通过蔗糖依赖性黏附形成生物膜并在其中产酸耐酸,最终导致龋病.VicRK是变异链球菌13种双组分信号传导系统之一,可调节变异链球菌致龋性毒力相关因子的表达.本文就VicRK的作用机制、结构组成、生理特性,及其对变异链球菌致龋性的影响,VicRK和VicX间的关系等研究进展作一综述.

  20. Protein phosphorylation and its role in archaeal signal transduction.

    Science.gov (United States)

    Esser, Dominik; Hoffmann, Lena; Pham, Trong Khoa; Bräsen, Christopher; Qiu, Wen; Wright, Phillip C; Albers, Sonja-Verena; Siebers, Bettina

    2016-09-01

    Reversible protein phosphorylation is the main mechanism of signal transduction that enables cells to rapidly respond to environmental changes by controlling the functional properties of proteins in response to external stimuli. However, whereas signal transduction is well studied in Eukaryotes and Bacteria, the knowledge in Archaea is still rather scarce. Archaea are special with regard to protein phosphorylation, due to the fact that the two best studied phyla, the Euryarchaeota and Crenarchaeaota, seem to exhibit fundamental differences in regulatory systems. Euryarchaeota (e.g. halophiles, methanogens, thermophiles), like Bacteria and Eukaryotes, rely on bacterial-type two-component signal transduction systems (phosphorylation on His and Asp), as well as on the protein phosphorylation on Ser, Thr and Tyr by Hanks-type protein kinases. Instead, Crenarchaeota (e.g. acidophiles and (hyper)thermophiles) only depend on Hanks-type protein phosphorylation. In this review, the current knowledge of reversible protein phosphorylation in Archaea is presented. It combines results from identified phosphoproteins, biochemical characterization of protein kinases and protein phosphatases as well as target enzymes and first insights into archaeal signal transduction by biochemical, genetic and polyomic studies.

  1. Two component theory and electron magnetic moment

    NARCIS (Netherlands)

    Veltman, M.J.G.

    1998-01-01

    The two-component formulation of quantum electrodynamics is studied. The relation with the usual Dirac formulation is exhibited, and the Feynman rules for the two-component form of the theory are presented in terms of familiar objects. The transformation from the Dirac theory to the two-component th

  2. Two component theory and electron magnetic moment

    NARCIS (Netherlands)

    Veltman, M.J.G.

    1998-01-01

    The two-component formulation of quantum electrodynamics is studied. The relation with the usual Dirac formulation is exhibited, and the Feynman rules for the two-component form of the theory are presented in terms of familiar objects. The transformation from the Dirac theory to the two-component

  3. The Campylobacter jejuni PhosS/PhosR operon represents a non-classical phosphate-sensitive two-component system

    NARCIS (Netherlands)

    Wösten, MM; Parker, Craig T; van Mourik, Andries; Guilhabert, Magalie R; van Dijk, Linda; van Putten, Jos P M

    2006-01-01

    The bacterial pathogen Campylobacter jejuni carries several putative two-component signal transduction systems of unknown function. Here we report that the PhosS (Cj0889) and PhosR (Cj0890) proteins constitute a two-component system that is activated by phosphate limitation. Microarray analysis,

  4. Two component systems: physiological effect of a third component.

    Directory of Open Access Journals (Sweden)

    Baldiri Salvado

    Full Text Available Signal transduction systems mediate the response and adaptation of organisms to environmental changes. In prokaryotes, this signal transduction is often done through Two Component Systems (TCS. These TCS are phosphotransfer protein cascades, and in their prototypical form they are composed by a kinase that senses the environmental signals (SK and by a response regulator (RR that regulates the cellular response. This basic motif can be modified by the addition of a third protein that interacts either with the SK or the RR in a way that could change the dynamic response of the TCS module. In this work we aim at understanding the effect of such an additional protein (which we call "third component" on the functional properties of a prototypical TCS. To do so we build mathematical models of TCS with alternative designs for their interaction with that third component. These mathematical models are analyzed in order to identify the differences in dynamic behavior inherent to each design, with respect to functionally relevant properties such as sensitivity to changes in either the parameter values or the molecular concentrations, temporal responsiveness, possibility of multiple steady states, or stochastic fluctuations in the system. The differences are then correlated to the physiological requirements that impinge on the functioning of the TCS. This analysis sheds light on both, the dynamic behavior of synthetically designed TCS, and the conditions under which natural selection might favor each of the designs. We find that a third component that modulates SK activity increases the parameter space where a bistable response of the TCS module to signals is possible, if SK is monofunctional, but decreases it when the SK is bifunctional. The presence of a third component that modulates RR activity decreases the parameter space where a bistable response of the TCS module to signals is possible.

  5. Giardia mitosomal protein import machinery differentially recognizes mitochondrial targeting signals.

    Science.gov (United States)

    Nyindodo-Ogari, Lilian; Schwartzbach, Steven D; Estraño, Carlos E

    2014-01-01

    Giardia lamblia mitosomes are believed to be vestigial mitochondria which lack a genome. Similar to higher eukaryotes, mitosomal proteins possess either N-terminal or internal mitosomal targeting sequences. To date, some components of the higher eukaryote archetypal mitochondrial protein import apparatus have been identified and characterized in Giardia mitosomes; therefore, it is expected that mitochondrial signals will be recognized by the mitosomal protein import system. To further determine the level of conservation of the Giardia mitosome protein import apparatus, we expressed mitochondrial proteins from higher eukaryotes in Giardia. These recombinant proteins include Tom20 and Tom22; two components of the mitochondrial protein import machinery. Our results indicate that N-terminal mitochondrial targeting sequence is recognized by the mitosomal protein import machinery; however, interestingly the internal mitochondrial targeting sequences of higher eukaryotes are not recognized by the mitosome. Our results indicate that Giardia mitosome protein transport machinery shows differential recognition of higher eukaryotic mitochondria transfer signals, suggesting a divergence of the transport system in G. lamblia. Therefore, our data support the hypothesis that the protein import machinery in Giardia lamblia mitosome is an incomplete vestigial derivative of mitochondria components.

  6. Two-component Duality and Strings

    CERN Document Server

    Freund, Peter G O

    2007-01-01

    A phenomenologically successful two-component hadronic duality picture led to Veneziano's amplitude, the fundamental first step to string theory. This picture is briefly recalled and its two components are identified as the open strings (mesons and baryons) and closed strings (Pomeron).

  7. Predicting Secretory Proteins with SignalP

    DEFF Research Database (Denmark)

    Nielsen, Henrik

    2017-01-01

    SignalP is the currently most widely used program for prediction of signal peptides from amino acid sequences. Proteins with signal peptides are targeted to the secretory pathway, but are not necessarily secreted. After a brief introduction to the biology of signal peptides and the history...

  8. A novel two-component system involved in the transition to secondary metabolism in Streptomyces coelicolor.

    Directory of Open Access Journals (Sweden)

    Daniel Rozas

    Full Text Available BACKGROUND: Bacterial two-component signal transduction regulatory systems are the major set of signalling proteins frequently mediating responses to changes in the environment. They typically consist of a sensor, a membrane-associated histidine kinase and a cytoplasmic response regulator. The membrane-associated sensor detects the environmental signal or stress, whereas the cytoplasmic regulatory protein controls the cellular response usually by gene transcription modulation. METHODOLOGY/PRINCIPALFINDINGS: The Streptomyces coelicolor two genes operon SCO5784-SCO5785 encodes a two-component system, where SCO5784 encodes a histidine-kinase sensor and SCO5785 encodes a response regulator protein. When the expression level of the regulator gene decreases, the antibiotic synthesis and sporulation is delayed temporarily in addition to some ribosomal genes became up regulated, whereas the propagation of the regulatory gene in high copy number results in the earlier synthesis of antibiotics and sporulation, as well as the down regulation of some ribosomal genes and, moreover, in the overproduction of several extracellular proteins. Therefore, this two-component system in S. coelicolor seems to influence various processes characterised by the transition from primary to secondary metabolism, as determined by proteomic and transcriptomic analyses. CONCLUSIONS/SIGNIFICANCE: Propagation of SCO5785 in multicopy enhances the production of antibiotics as well as secretory proteins. In particular, the increase in the expression level of secretory protein encoding genes, either as an artefactual or real effect of the regulator, could be of potential usefulness when using Streptomyces strains as hosts for homologous or heterologous extracellular protein production.

  9. Sub classification and targeted characterization of prophage-encoded two-component cell lysis cassette

    Indian Academy of Sciences (India)

    K V Srividhya; S Krishnaswamy

    2007-08-01

    Bacteriophage induced lysis of host bacterial cell is mediated by a two component cell lysis cassette comprised of holin and lysozyme. Prophages are integrated forms of bacteriophages in bacterial genomes providing a repertoire for bacterial evolution. Analysis using the prophage database (http://bicmku.in:8082) constructed by us showed 47 prophages were associated with putative two component cell lysis genes. These proteins cluster into four different subgroups. In this process, a putative holin (essd) and endolysin (ybcS), encoded by the defective lambdoid prophage DLP12 was found to be similar to two component cell lysis genes in functional bacteriophages like p21 and P1. The holin essd was found to have a characteristic dual start motif with two transmembrane regions and C-terminal charged residues as in class II holins. Expression of a fusion construct of essd in Escherichia coli showed slow growth. However, under appropriate conditions, this protein could be over expressed and purified for structure function studies. The second component of the cell lysis cassette, ybcS, was found to have an N-terminal SAR (Signal Arrest Release) transmembrane domain. The construct of ybcS has been over expressed in E. coli and the purified protein was functional, exhibiting lytic activity against E. coli and Salmonella typhi cell wall substrate. Such targeted sequence-structure-function characterization of proteins encoded by cryptic prophages will help understand the contribution of prophage proteins to bacterial evolution.

  10. Identification and functional assay of the interaction motifs in the partner protein OsNAR2.1 of the two-component system for high-affinity nitrate transport.

    Science.gov (United States)

    Liu, Xiaoqin; Huang, Daimin; Tao, Jinyuan; Miller, Anthony J; Fan, Xiaorong; Xu, Guohua

    2014-10-01

    A partner protein, NAR2, is essential for high-affinity nitrate transport of the NRT2 protein in plants. However, the NAR2 motifs that interact with NRT2s for their plasma membrane (PM) localization and nitrate transporter activity have not been functionally characterized. In this study, OsNAR2.1 mutations with different carbon (C)-terminal deletions and nine different point mutations in the conserved regions of NAR2 homologs in plants were generated to explore the essential motifs involved in the interaction with OsNRT2.3a. Screening using the membrane yeast two-hybrid system and Xenopus oocytes for nitrogen-15 ((15)N) uptake demonstrated that either R100G or D109N point mutations impaired the OsNAR2.1 interaction with OsNRT2.3a. Western blotting and visualization using green fluorescent protein fused to either the N- or C-terminus of OsNAR2.1 indicated that OsNAR2.1 is expressed in both the PM and cytoplasm. The split-yellow fluorescent protein (YFP)/BiFC analyses indicated that OsNRT2.3a was targeted to the PM in the presence of OsNAR2.1, while either R100G or D109N mutation resulted in the loss of OsNRT2.3a-YFP signal in the PM. Based on these results, arginine 100 and aspartic acid 109 of the OsNAR2.1 protein are key amino acids in the interaction with OsNRT2.3a, and their interaction occurs in the PM but not cytoplasm.

  11. Two-component Abelian sandpile models.

    Science.gov (United States)

    Alcaraz, F C; Pyatov, P; Rittenberg, V

    2009-04-01

    In one-component Abelian sandpile models, the toppling probabilities are independent quantities. This is not the case in multicomponent models. The condition of associativity of the underlying Abelian algebras imposes nonlinear relations among the toppling probabilities. These relations are derived for the case of two-component quadratic Abelian algebras. We show that Abelian sandpile models with two conservation laws have only trivial avalanches.

  12. Regulation of Nuclear Localization of Signaling Proteins by Cytokinin

    Energy Technology Data Exchange (ETDEWEB)

    Kieber, J.J.

    2010-05-01

    Cytokinins are a class of mitogenic plant hormones that play an important role in most aspects of plant development, including shoot and root growth, vascular and photomorphogenic development and leaf senescence. A model for cytokinin perception and signaling has emerged that is similar to bacterial two-component phosphorelays. In this model, binding of cytokinin to the extracellular domain of the Arabidopsis histidine kinase (AHKs) receptors induces autophosphorylation within the intracellular histidine-kinase domain. The phosphoryl group is subsequently transferred to cytosolic Arabidopsis histidine phosphotransfer proteins (AHPs), which have been suggested to translocate to the nucleus in response to cytokinin treatment, where they then transfer the phosphoryl group to nuclear-localized response regulators (Type-A and Type-B ARRs). We examined the effects of cytokinin on AHP subcellular localization in Arabidopsis and, contrary to expectations, the AHPs maintained a constant nuclear/cytosolic distribution following cytokinin treatment. Furthermore, mutation of the conserved phosphoacceptor histidine residue of the AHP, as well as disruption of multiple cytokinin signaling elements, did not affect the subcellular localization of the AHP proteins. Finally, we present data indicating that AHPs maintain a nuclear/cytosolic distribution by balancing active transport into and out of the nucleus. Our findings suggest that the current models indicating relocalization of AHP protein into the nucleus in response to cytokinin are incorrect. Rather, AHPs actively maintain a consistent nuclear/cytosolic distribution regardless of the status of the cytokinin response pathway.

  13. Regulation of Nuclear Localization of Signaling Proteins by Cytokinin

    Energy Technology Data Exchange (ETDEWEB)

    Kieber, J.J.

    2010-05-01

    Cytokinins are a class of mitogenic plant hormones that play an important role in most aspects of plant development, including shoot and root growth, vascular and photomorphogenic development and leaf senescence. A model for cytokinin perception and signaling has emerged that is similar to bacterial two-component phosphorelays. In this model, binding of cytokinin to the extracellular domain of the Arabidopsis histidine kinase (AHKs) receptors induces autophosphorylation within the intracellular histidine-kinase domain. The phosphoryl group is subsequently transferred to cytosolic Arabidopsis histidine phosphotransfer proteins (AHPs), which have been suggested to translocate to the nucleus in response to cytokinin treatment, where they then transfer the phosphoryl group to nuclear-localized response regulators (Type-A and Type-B ARRs). We examined the effects of cytokinin on AHP subcellular localization in Arabidopsis and, contrary to expectations, the AHPs maintained a constant nuclear/cytosolic distribution following cytokinin treatment. Furthermore, mutation of the conserved phosphoacceptor histidine residue of the AHP, as well as disruption of multiple cytokinin signaling elements, did not affect the subcellular localization of the AHP proteins. Finally, we present data indicating that AHPs maintain a nuclear/cytosolic distribution by balancing active transport into and out of the nucleus. Our findings suggest that the current models indicating relocalization of AHP protein into the nucleus in response to cytokinin are incorrect. Rather, AHPs actively maintain a consistent nuclear/cytosolic distribution regardless of the status of the cytokinin response pathway.

  14. Signal peptides and protein localization prediction

    DEFF Research Database (Denmark)

    Nielsen, Henrik

    2005-01-01

    In 1999, the Nobel prize in Physiology or Medicine was awarded to Gunther Blobel “for the discovery that proteins have intrinsic signals that govern their transport and localization in the cell”. Since the subcellular localization of a protein is an important clue to its function...

  15. Two-component model of solar plages

    Institute of Scientific and Technical Information of China (English)

    LI; Jianping(李建平); DING; Mingde(丁明德); FANG; Cheng(方成)

    2002-01-01

    By use of the 2-m Mcmath-Pierce telescope at Kitt Peak, the high-quality spectra of a plage with moderate brightness near the center of solar disk were obtained. The data include seven spectral lines, which are Hα, Hβ, CaII H and K lines and the infrared triplet. With the consideration of fine structures of solar plages, a two-component atmospheric model is constructed by keeping the cool component to be the quiet atmosphere. Three cases of the hot component are given for different filling factors where the temperature and density distribution are adjusted in order to reproduce the seven observed spectral profiles. We also briefly discuss the influence of the column density at the base of the corona, m0, and the macro-turbulent velocity on the required filling factor and computed profiles. The two-component model is compared with precious one-component semi-empirical models. The limitation of the model is pointed out and further improvement is indicated.

  16. Protein cysteine oxidation in redox signaling

    DEFF Research Database (Denmark)

    Forman, Henry Jay; Davies, Michael J; Krämer, Anna C

    2017-01-01

    . Previous studies have claimed that RSOH can be detected as an adduct (e.g., with 5,5-dimethylcyclohexane-1,3-dione; dimedone). Here, kinetic data are discussed which indicate that few proteins can form RSOH under physiological signaling conditions. We also present experimental evidence that indicates......Oxidation of critical signaling protein cysteines regulated by H2O2 has been considered to involve sulfenic acid (RSOH) formation. RSOH may subsequently form either a sulfenyl amide (RSNHR') with a neighboring amide, or a mixed disulfide (RSSR') with another protein cysteine or glutathione...

  17. Protein Translation and Signaling in Human Eosinophils

    Directory of Open Access Journals (Sweden)

    Stephane Esnault

    2017-09-01

    Full Text Available We have recently reported that, unlike IL-5 and GM-CSF, IL-3 induces increased translation of a subset of mRNAs. In addition, we have demonstrated that Pin1 controls the activity of mRNA binding proteins, leading to enhanced mRNA stability, GM-CSF protein production and prolonged eosinophil (EOS survival. In this review, discussion will include an overview of cap-dependent protein translation and its regulation by intracellular signaling pathways. We will address the more general process of mRNA post-transcriptional regulation, especially regarding mRNA binding proteins, which are critical effectors of protein translation. Furthermore, we will focus on (1 the roles of IL-3-driven sustained signaling on enhanced protein translation in EOS, (2 the mechanisms regulating mRNA binding proteins activity in EOS, and (3 the potential targeting of IL-3 signaling and the signaling leading to mRNA binding activity changes to identify therapeutic targets to treat EOS-associated diseases.

  18. Arabidopsis ethylene-response gene ETR1: Similiarity of product to two-component regulators

    Energy Technology Data Exchange (ETDEWEB)

    Chang, C.; Kwok, S.F.; Bleecker, A.B.; Meyerowitz, E.M. (California Institute of Technology, Pasadena, CA (United States))

    1993-10-22

    Ethylene behaves as a hormone in plants, regulating such aspects of growth and development as fruit ripening, flower senescence, and abscission. Ethylene insensitivity is conferred by dominant mutations in the ETR1 gene early in the ethylene signal transduction pathway of Arabidopsis thaliana. The ETR1 gene was cloned by the method of chromosome walking. Each of the four known etr1 mutant alleles contains a missense mutation near the amino terminus of the predicted protein. Although the sequence of the amino-terminal half of the deduced ETR1 protein appears to be novel, the carboxyl-terminal half is similar in sequence to both components of the prokaryotic family of signal transducers known as the two-component systems. Thus, an early step in ethylene signal transduction in plants may involve transfer of phosphate as in prokaryotic two-component systems. The dominant etr1-1 mutant gene conferred ethylene insensitivity to wild-type Arabidopsis plants when introduced by transformation.

  19. Two-Component Description for Relativistic Fermions

    Institute of Scientific and Technical Information of China (English)

    CHEN Yu-Qi; SANG Wen-Long; YANG Lan-Fei

    2009-01-01

    We propose a two-component form to describe massive relativistic fermions in gauge theories. Relations between the Green's functions in this form and those in the conventional four-component form are derived. It is shown that the S-matrix elements in both forms are exactly the same. The description of the fermion in the new form simplifies significantly the γ-matrix algebra in the four-component form. In particular, in perturbative calculations the propagator of the fermion is a scalar function. As examples, we use this form to reproduce the relativistic spectrum of hydrodron atom, the S-matrix of e+ e-→μ+ μ- and QED one-loop vacuum polarization of photon.

  20. The SaeRS Two-Component System of Staphylococcus aureus

    Science.gov (United States)

    Liu, Qian; Yeo, Won-Sik; Bae, Taeok

    2016-01-01

    In the Gram-positive pathogenic bacterium Staphylococcus aureus, the SaeRS two-component system (TCS) plays a major role in controlling the production of over 20 virulence factors including hemolysins, leukocidins, superantigens, surface proteins, and proteases. The SaeRS TCS is composed of the sensor histidine kinase SaeS, response regulator SaeR, and two auxiliary proteins SaeP and SaeQ. Since its discovery in 1994, the sae locus has been studied extensively, and its contributions to staphylococcal virulence and pathogenesis have been well documented and understood; however, the molecular mechanism by which the SaeRS TCS receives and processes cognate signals is not. In this article, therefore, we review the literature focusing on the signaling mechanism and its interaction with other global regulators. PMID:27706107

  1. Chloroplast two-component systems: evolution of the link between photosynthesis and gene expression

    OpenAIRE

    Puthiyaveetil, Sujith; Allen, John F.

    2009-01-01

    Two-component signal transduction, consisting of sensor kinases and response regulators, is the predominant signalling mechanism in bacteria. This signalling system originated in prokaryotes and has spread throughout the eukaryotic domain of life through endosymbiotic, lateral gene transfer from the bacterial ancestors and early evolutionary precursors of eukaryotic, cytoplasmic, bioenergetic organelles—chloroplasts and mitochondria. Until recently, it was thought that two-component systems i...

  2. Tobacco two-component gene NTHK2

    Institute of Scientific and Technical Information of China (English)

    2001-01-01

    By using a previously isolated tobacco two- component gene NTHK1 as a probe, we screened a cDNA library and obtained a homologous gene designated NTHK2. Sequencing analysis revealed that NTHK2 encoded a putative ethylene receptor homolog and contained a histidine kinase domain and a receiver domain. In the histidine kinase domain, the histidine at the phosphorylation site was replaced by an asparagine. Southern analysis indicated that NTHK2 was present at low copies in tobacco genome. The expression of NTHK2 was studied using a competitive RT-PCR method. It was found that, in young flower buds, NTHK2 was expressed abundantly, while in other organs or tissues, it was expressed in a low level. When leaf was subjected to wounding (cutting) treatment, NTHK2 expression was increased. When tobacco seedlings were stressed with PEG and heat shock, NTHK2 transcription was also enhanced. Other treatments showed little effects. These results indicated that NTHK2 might be involved in the developmental processes and in plant responses to some environmental stresses.

  3. The evolution of two-component systems in bacteria reveals different strategies for niche adaptation.

    Directory of Open Access Journals (Sweden)

    Eric Alm

    2006-11-01

    Full Text Available Two-component systems including histidine protein kinases represent the primary signal transduction paradigm in prokaryotic organisms. To understand how these systems adapt to allow organisms to detect niche-specific signals, we analyzed the phylogenetic distribution of nearly 5,000 histidine protein kinases from 207 sequenced prokaryotic genomes. We found that many genomes carry a large repertoire of recently evolved signaling genes, which may reflect selective pressure to adapt to new environmental conditions. Both lineage-specific gene family expansion and horizontal gene transfer play major roles in the introduction of new histidine kinases into genomes; however, there are differences in how these two evolutionary forces act. Genes imported via horizontal transfer are more likely to retain their original functionality as inferred from a similar complement of signaling domains, while gene family expansion accompanied by domain shuffling appears to be a major source of novel genetic diversity. Family expansion is the dominant source of new histidine kinase genes in the genomes most enriched in signaling proteins, and detailed analysis reveals that divergence in domain structure and changes in expression patterns are hallmarks of recent expansions. Finally, while these two modes of gene acquisition are widespread across bacterial taxa, there are clear species-specific preferences for which mode is used.

  4. Ubiquitin in signaling and protein quality control

    DEFF Research Database (Denmark)

    Al-Saoudi, Sofie Vincents

    Protein ubiquitylation is an important post-translational modification that holds a variety of cellular functions. This Ph.D. thesis is comprised of two studies, of which one focused on ubiquitylation related to inflammatory signaling, and the other on the role of the ubiquitin-proteasome system ...

  5. Glycosphingolipid–Protein Interaction in Signal Transduction

    Directory of Open Access Journals (Sweden)

    Domenico Russo

    2016-10-01

    Full Text Available Glycosphingolipids (GSLs are a class of ceramide-based glycolipids essential for embryo development in mammals. The synthesis of specific GSLs depends on the expression of distinctive sets of GSL synthesizing enzymes that is tightly regulated during development. Several reports have described how cell surface receptors can be kept in a resting state or activate alternative signalling events as a consequence of their interaction with GSLs. Specific GSLs, indeed, interface with specific protein domains that are found in signalling molecules and which act as GSL sensors to modify signalling responses. The regulation exerted by GSLs on signal transduction is orthogonal to the ligand–receptor axis, as it usually does not directly interfere with the ligand binding to receptors. Due to their properties of adjustable production and orthogonal action on receptors, GSLs add a new dimension to the control of the signalling in development. GSLs can, indeed, dynamically influence progenitor cell response to morphogenetic stimuli, resulting in alternative differentiation fates. Here, we review the available literature on GSL–protein interactions and their effects on cell signalling and development.

  6. Glycosphingolipid–Protein Interaction in Signal Transduction

    Science.gov (United States)

    Russo, Domenico; Parashuraman, Seetharaman; D’Angelo, Giovanni

    2016-01-01

    Glycosphingolipids (GSLs) are a class of ceramide-based glycolipids essential for embryo development in mammals. The synthesis of specific GSLs depends on the expression of distinctive sets of GSL synthesizing enzymes that is tightly regulated during development. Several reports have described how cell surface receptors can be kept in a resting state or activate alternative signalling events as a consequence of their interaction with GSLs. Specific GSLs, indeed, interface with specific protein domains that are found in signalling molecules and which act as GSL sensors to modify signalling responses. The regulation exerted by GSLs on signal transduction is orthogonal to the ligand–receptor axis, as it usually does not directly interfere with the ligand binding to receptors. Due to their properties of adjustable production and orthogonal action on receptors, GSLs add a new dimension to the control of the signalling in development. GSLs can, indeed, dynamically influence progenitor cell response to morphogenetic stimuli, resulting in alternative differentiation fates. Here, we review the available literature on GSL–protein interactions and their effects on cell signalling and development. PMID:27754465

  7. Sentra : a database of signal transduction proteins for comparative genome analysis.

    Energy Technology Data Exchange (ETDEWEB)

    D' Souza, M.; Glass, E. M.; Syed, M. H.; Zhang, Y.; Rodriguez, A.; Maltsev, N.; Galerpin, M. Y.; Mathematics and Computer Science; Univ. of Chicago; NIH

    2007-01-01

    Sentra (http://compbio.mcs.anl.gov/sentra), a database of signal transduction proteins encoded in completely sequenced prokaryotic genomes, has been updated to reflect recent advances in understanding signal transduction events on a whole-genome scale. Sentra consists of two principal components, a manually curated list of signal transduction proteins in 202 completely sequenced prokaryotic genomes and an automatically generated listing of predicted signaling proteins in 235 sequenced genomes that are awaiting manual curation. In addition to two-component histidine kinases and response regulators, the database now lists manually curated Ser/Thr/Tyr protein kinases and protein phosphatases, as well as adenylate and diguanylate cyclases and c-di-GMP phosphodiesterases, as defined in several recent reviews. All entries in Sentra are extensively annotated with relevant information from public databases (e.g. UniProt, KEGG, PDB and NCBI). Sentra's infrastructure was redesigned to support interactive cross-genome comparisons of signal transduction capabilities of prokaryotic organisms from a taxonomic and phenotypic perspective and in the framework of signal transduction pathways from KEGG. Sentra leverages the PUMA2 system to support interactive analysis and annotation of signal transduction proteins by the users.

  8. Reciprocal Regulation as a Source of Ultrasensitivity in Two-Component Systems with a Bifunctional Sensor Kinase

    Science.gov (United States)

    Straube, Ronny

    2014-01-01

    Two-component signal transduction systems, where the phosphorylation state of a regulator protein is modulated by a sensor kinase, are common in bacteria and other microbes. In many of these systems, the sensor kinase is bifunctional catalyzing both, the phosphorylation and the dephosphorylation of the regulator protein in response to input signals. Previous studies have shown that systems with a bifunctional enzyme can adjust the phosphorylation level of the regulator protein independently of the total protein concentrations – a property known as concentration robustness. Here, I argue that two-component systems with a bifunctional enzyme may also exhibit ultrasensitivity if the input signal reciprocally affects multiple activities of the sensor kinase. To this end, I consider the case where an allosteric effector inhibits autophosphorylation and, concomitantly, activates the enzyme's phosphatase activity, as observed experimentally in the PhoQ/PhoP and NRII/NRI systems. A theoretical analysis reveals two operating regimes under steady state conditions depending on the effector affinity: If the affinity is low the system produces a graded response with respect to input signals and exhibits stimulus-dependent concentration robustness – consistent with previous experiments. In contrast, a high-affinity effector may generate ultrasensitivity by a similar mechanism as phosphorylation-dephosphorylation cycles with distinct converter enzymes. The occurrence of ultrasensitivity requires saturation of the sensor kinase's phosphatase activity, but is restricted to low effector concentrations, which suggests that this mode of operation might be employed for the detection and amplification of low abundant input signals. Interestingly, the same mechanism also applies to covalent modification cycles with a bifunctional converter enzyme, which suggests that reciprocal regulation, as a mechanism to generate ultrasensitivity, is not restricted to two-component systems, but

  9. Reciprocal regulation as a source of ultrasensitivity in two-component systems with a bifunctional sensor kinase.

    Directory of Open Access Journals (Sweden)

    Ronny Straube

    2014-05-01

    Full Text Available Two-component signal transduction systems, where the phosphorylation state of a regulator protein is modulated by a sensor kinase, are common in bacteria and other microbes. In many of these systems, the sensor kinase is bifunctional catalyzing both, the phosphorylation and the dephosphorylation of the regulator protein in response to input signals. Previous studies have shown that systems with a bifunctional enzyme can adjust the phosphorylation level of the regulator protein independently of the total protein concentrations--a property known as concentration robustness. Here, I argue that two-component systems with a bifunctional enzyme may also exhibit ultrasensitivity if the input signal reciprocally affects multiple activities of the sensor kinase. To this end, I consider the case where an allosteric effector inhibits autophosphorylation and, concomitantly, activates the enzyme's phosphatase activity, as observed experimentally in the PhoQ/PhoP and NRII/NRI systems. A theoretical analysis reveals two operating regimes under steady state conditions depending on the effector affinity: If the affinity is low the system produces a graded response with respect to input signals and exhibits stimulus-dependent concentration robustness--consistent with previous experiments. In contrast, a high-affinity effector may generate ultrasensitivity by a similar mechanism as phosphorylation-dephosphorylation cycles with distinct converter enzymes. The occurrence of ultrasensitivity requires saturation of the sensor kinase's phosphatase activity, but is restricted to low effector concentrations, which suggests that this mode of operation might be employed for the detection and amplification of low abundant input signals. Interestingly, the same mechanism also applies to covalent modification cycles with a bifunctional converter enzyme, which suggests that reciprocal regulation, as a mechanism to generate ultrasensitivity, is not restricted to two-component

  10. Reciprocal regulation as a source of ultrasensitivity in two-component systems with a bifunctional sensor kinase.

    Directory of Open Access Journals (Sweden)

    Ronny Straube

    2014-05-01

    Full Text Available Two-component signal transduction systems, where the phosphorylation state of a regulator protein is modulated by a sensor kinase, are common in bacteria and other microbes. In many of these systems, the sensor kinase is bifunctional catalyzing both, the phosphorylation and the dephosphorylation of the regulator protein in response to input signals. Previous studies have shown that systems with a bifunctional enzyme can adjust the phosphorylation level of the regulator protein independently of the total protein concentrations--a property known as concentration robustness. Here, I argue that two-component systems with a bifunctional enzyme may also exhibit ultrasensitivity if the input signal reciprocally affects multiple activities of the sensor kinase. To this end, I consider the case where an allosteric effector inhibits autophosphorylation and, concomitantly, activates the enzyme's phosphatase activity, as observed experimentally in the PhoQ/PhoP and NRII/NRI systems. A theoretical analysis reveals two operating regimes under steady state conditions depending on the effector affinity: If the affinity is low the system produces a graded response with respect to input signals and exhibits stimulus-dependent concentration robustness--consistent with previous experiments. In contrast, a high-affinity effector may generate ultrasensitivity by a similar mechanism as phosphorylation-dephosphorylation cycles with distinct converter enzymes. The occurrence of ultrasensitivity requires saturation of the sensor kinase's phosphatase activity, but is restricted to low effector concentrations, which suggests that this mode of operation might be employed for the detection and amplification of low abundant input signals. Interestingly, the same mechanism also applies to covalent modification cycles with a bifunctional converter enzyme, which suggests that reciprocal regulation, as a mechanism to generate ultrasensitivity, is not restricted to two-component

  11. A putative colR(XC1049)-colS(XC1050) two-component signal transduction system in Xanthomonas campestris positively regulates hrpC and hrpE operons and is involved in virulence, the hypersensitive response and tolerance to various stresses.

    Science.gov (United States)

    Zhang, Sui-Sheng; He, Yong-Qiang; Xu, Li-Ming; Chen, Bo-Wen; Jiang, Bo-Le; Liao, Jie; Cao, Jin-Rui; Liu, Dan; Huang, Yan-Qiang; Liang, Xiao-Xia; Tang, Dong-Jie; Lu, Guang-Tao; Tang, Ji-Liang

    2008-01-01

    The ColR-ColS two-component signal transduction system was originally characterized as a regulatory system involved in the capacity of root-colonizing biocontrol bacterium Pseudomonas fluorescens to colonize plant roots. There are three pairs of putative colR-colS two-component regulatory systems annotated in the phytopathogen Xanthomonas campestris pathovar campestris. Mutational studies revealed that one of them, named colR(XC1049) and colS(XC1050), is a global regulatory system involved in various cellular processes, including virulence, hypersensitive response and stress tolerance. Growth rate determination showed that, although the colR(XC1049) and colS(XC1050) mutants are not auxotrophic, colR(XC1049) and colS(XC1050) are required for the pathogen to proliferate well in standard media and host plants. Assays of beta-glucuronidase activities of plasmid-driven promoter-gusA reporters and/or semi-quantitative RT-PCR demonstrated that colR(XC1049) and colS(XC1050) positively regulate expression of hrpC and hrpE operons, and that expression of colR(XC1049) and colS(XC1050) is not controlled by key hrp regulators HrpG and HrpX.

  12. Dynamics of two-component membranes surrounded by viscoelastic media.

    Science.gov (United States)

    Komura, Shigeyuki; Yasuda, Kento; Okamoto, Ryuichi

    2015-11-01

    We discuss the dynamics of two-component fluid membranes which are surrounded by viscoelastic media. We assume that membrane-embedded proteins can diffuse laterally and induce a local membrane curvature. The mean squared displacement of a tagged membrane segment is obtained as a generalized Einstein relation. When the elasticity of the surrounding media obeys a power-law behavior in frequency, an anomalous diffusion of the membrane segment is predicted. We also consider the situation where the proteins generate active non-equilibrium forces. The generalized Einstein relation is further modified by an effective temperature that depends on the force dipole energy. The obtained generalized Einstein relations are useful for membrane microrheology experiments.

  13. The Evolution of Two-Component Systems in Bacteria RevealsDifferent Strategies for Niche Adaptation

    Energy Technology Data Exchange (ETDEWEB)

    Alm, Eric; Huang, Katherine; Arkin, Adam

    2006-09-13

    Two-component systems including histidine protein kinasesrepresent the primary signal transduction paradigm in prokaryoticorganisms. To understand how these systems adapt to allow organisms todetect niche-specific signals, we analyzed the phylogenetic distributionof nearly 5000 histidine protein kinases from 207 sequenced prokaryoticgenomes. We found that many genomes carry a large repertoire of recentlyevolved signaling genes, which may reflect selective pressure to adapt tonew environmental conditions. Both lineage-specific gene family expansionand horizontal gene transfer play major roles in the introduction of newhistidine kinases into genomes; however, there are differences in howthese two evolutionary forces act. Genes imported via horizontal transferare more likely to retain their original functionality as inferred from asimilar complement of signaling domains, while gene family expansionaccompanied by domain shuffling appears to be a major source of novelgenetic diversity. Family expansion is the dominantsource of newhistidine kinase genes in the genomes most enriched in signalingproteins, and detailed analysis reveals that divergence in domainstructure and changes in expression patterns are hallmarks of recentexpansions. Finally, while these two modes of gene acquisition arewidespread across bacterial taxa, there are clear species-specificpreferences for which mode is used.

  14. Evolutionary analysis and lateral gene transfer of two-component regulatory systems associated with heavy-metal tolerance in bacteria.

    Science.gov (United States)

    Bouzat, Juan L; Hoostal, Matthew J

    2013-05-01

    Microorganisms have adapted intricate signal transduction mechanisms to coordinate tolerance to toxic levels of metals, including two-component regulatory systems (TCRS). In particular, both cop and czc operons are regulated by TCRS; the cop operon plays a key role in bacterial tolerance to copper, whereas the czc operon is involved in the efflux of cadmium, zinc, and cobalt from the cell. Although the molecular physiology of heavy metal tolerance genes has been extensively studied, their evolutionary relationships are not well-understood. Phylogenetic relationships among heavy-metal efflux proteins and their corresponding two-component regulatory proteins revealed orthologous and paralogous relationships from species divergences and ancient gene duplications. The presence of heavy metal tolerance genes on bacterial plasmids suggests these genes may be prone to spread through horizontal gene transfer. Phylogenetic inferences revealed nine potential examples of lateral gene transfer associated with metal efflux proteins and two examples for regulatory proteins. Notably, four of the examples suggest lateral transfer across major evolutionary domains. In most cases, differences in GC content in metal tolerance genes and their corresponding host genomes confirmed lateral gene transfer events. Three-dimensional protein structures predicted for the response regulators encoded by cop and czc operons showed a high degree of structural similarity with other known proteins involved in TCRS signal transduction, which suggests common evolutionary origins of functional phenotypes and similar mechanisms of action for these response regulators.

  15. Mitogen-activated protein kinase signaling pathways of the tangerine pathotype of Alternaria alternata

    Directory of Open Access Journals (Sweden)

    Kuang-Ren Chung

    2013-06-01

    Full Text Available Mitogen-activated protein kinase (MAPK- mediated signaling pathways have been known to have important functions in eukaryotic organisms. The mechanisms by which the filamentous fungus Alternaria alternata senses and responds to environmental signals have begun to be elucidated. Available data indicate that A. alternata utilizes the Fus3, Hog1 and Slt2 MAPK-mediated signaling pathways, either separately or in a cooperative manner, for conidia formation, resistance to oxidative and osmotic stress, and pathogenesis to citrus. This review provides an overview of our current knowledge of MAPK signaling pathways, in conjunction with the two-component histidine kinase and the Skn7 response regulator, in the tangerine pathotype of A. alternata.

  16. Ubiquitin in signaling and protein quality control

    DEFF Research Database (Denmark)

    Al-Saoudi, Sofie Vincents

    Protein ubiquitylation is an important post-translational modification that holds a variety of cellular functions. This Ph.D. thesis is comprised of two studies, of which one focused on ubiquitylation related to inflammatory signaling, and the other on the role of the ubiquitin-proteasome system......-terminal methionine (M1), and recently, the deubiquitylating enzyme, OTULIN, was discovered to counter LUBAC activity by exclusively cleaving M1-linked ubiquitin chains. We provide the molecular detail of the interaction between the LUBAC subunit, HOIP, and OTULIN. The interaction was mapped to the PUB-domain of HOIP...

  17. Dynamical principles of two-component genetic oscillators.

    Directory of Open Access Journals (Sweden)

    Raúl Guantes

    2006-03-01

    Full Text Available Genetic oscillators based on the interaction of a small set of molecular components have been shown to be involved in the regulation of the cell cycle, the circadian rhythms, or the response of several signaling pathways. Uncovering the functional properties of such oscillators then becomes important for the understanding of these cellular processes and for the characterization of fundamental properties of more complex clocks. Here, we show how the dynamics of a minimal two-component oscillator is drastically affected by its genetic implementation. We consider a repressor and activator element combined in a simple logical motif. While activation is always exerted at the transcriptional level, repression is alternatively operating at the transcriptional (Design I or post-translational (Design II level. These designs display differences on basic oscillatory features and on their behavior with respect to molecular noise or entrainment by periodic signals. In particular, Design I induces oscillations with large activator amplitudes and arbitrarily small frequencies, and acts as an "integrator" of external stimuli, while Design II shows emergence of oscillations with finite, and less variable, frequencies and smaller amplitudes, and detects better frequency-encoded signals ("resonator". Similar types of stimulus response are observed in neurons, and thus this work enables us to connect very different biological contexts. These dynamical principles are relevant for the characterization of the physiological roles of simple oscillator motifs, the understanding of core machineries of complex clocks, and the bio-engineering of synthetic oscillatory circuits.

  18. SOCS proteins in regulation of receptor tyrosine kinase signaling

    DEFF Research Database (Denmark)

    Kazi, Julhash U.; Kabir, Nuzhat N.; Flores Morales, Amilcar;

    2014-01-01

    . The signaling mediated by RTKs must be tightly regulated by interacting proteins including protein-tyrosine phosphatases and ubiquitin ligases. The suppressors of cytokine signaling (SOCS) family proteins are well-known negative regulators of cytokine receptors signaling consisting of eight structurally similar...

  19. Two-component systems and toxinogenesis regulation in Clostridium botulinum.

    Science.gov (United States)

    Connan, Chloé; Popoff, Michel R

    2015-05-01

    Botulinum neurotoxins (BoNTs) are the most potent toxins ever known. They are mostly produced by Clostridium botulinum but also by other clostridia. BoNTs associate with non-toxic proteins (ANTPs) to form complexes of various sizes. Toxin production is highly regulated through complex networks of regulatory systems involving an alternative sigma factor, BotR, and at least 6 recently described two-component systems (TCSs). TCSs allow bacteria to sense environmental changes and to respond to various stimuli by regulating the expression of specific genes at a transcriptional level. Several environmental stimuli have been identified to positively or negatively regulate toxin synthesis; however, the link between environmental stimuli and TCSs is still elusive. This review aims to highlight the role of TCSs as a central point in the regulation of toxin production in C. botulinum.

  20. Protein phosphorylation in bcterial signaling and regulation

    KAUST Repository

    Mijakovic, Ivan

    2016-01-26

    In 2003, it was demonstrated for the first time that bacteria possess protein-tyrosine kinases (BY-kinases), capable of phosphorylating other cellular proteins and regulating their activity. It soon became apparent that these kinases phosphorylate a number of protein substrates, involved in different cellular processes. More recently, we found out that BY-kinases can be activated by several distinct protein interactants, and are capable of engaging in cross-phosphorylation with other kinases. Evolutionary studies based on genome comparison indicate that BY-kinases exist only in bacteria. They are non-essential (present in about 40% bacterial genomes), and their knockouts lead to pleiotropic phenotypes, since they phosphorylate many substrates. Surprisingly, BY-kinase genes accumulate mutations at an increased rate (non-synonymous substitution rate significantly higher than other bacterial genes). One direct consequence of this phenomenon is no detectable co-evolution between kinases and their substrates. Their promiscuity towards substrates thus seems to be “hard-wired”, but why would bacteria maintain such promiscuous regulatory devices? One explanation is the maintenance of BY-kinases as rapidly evolving regulators, which can readily adopt new substrates when environmental changes impose selective pressure for quick evolution of new regulatory modules. Their role is clearly not to act as master regulators, dedicated to triggering a single response, but they might rather be employed to contribute to fine-tuning and improving robustness of various cellular responses. This unique feature makes BY-kinases a potentially useful tool in synthetic biology. While other bacterial kinases are very specific and their signaling pathways insulated, BY-kinase can relatively easily be engineered to adopt new substrates and control new biosynthetic processes. Since they are absent in humans, and regulate some key functions in pathogenic bacteria, they are also very promising

  1. Bioactive motifs of agouti signal protein.

    Science.gov (United States)

    Virador, V M; Santis, C; Furumura, M; Kalbacher, H; Hearing, V J

    2000-08-25

    The switch between the synthesis of eu- and pheomelanins is modulated by the interaction of two paracrine signaling molecules, alpha-melanocyte stimulating hormone (MSH) and agouti signal protein (ASP), which interact with melanocytes via the MSH receptor (MC1R). Comparison of the primary sequence of ASP with the known MSH pharmacophore provides no suggestion about the putative bioactive domain(s) of ASP. To identify such bioactive motif(s), we synthesized 15-mer peptides that spanned the primary sequence of ASP and determined their effects on the melanogenic activities of murine melanocytes. Northern and Western blotting were used, together with chemical analysis of melanins and enzymatic assays, to identify three distinct bioactive regions of ASP that down-regulate eumelanogenesis. The decrease in eumelanin production was mediated by down-regulation of mRNA levels for tyrosinase and other melanogenic enzymes, as occurs in vivo, and these effects were comparable to those elicited by intact recombinant ASP. Shorter peptides in those motifs were synthesized and their effects on melanogenesis were further investigated. The amino acid arginine, which is present in the MSH peptide pharmacophore (HFRW), is also in the most active domain of ASP (KVARP). Our data suggest that lysines and an arginine (in motifs such as KxxxxKxxR or KxxRxxxxK) are important for the bioactivity of ASP. Identification of the specific ASP epitope that interacts with the MC1R has potential pharmacological applications in treating dysfunctions of skin pigmentation.

  2. G protein signaling in plants: minus times minus equals plus.

    Science.gov (United States)

    Stateczny, Dave; Oppenheimer, Jara; Bommert, Peter

    2016-12-01

    Heterotrimeric G proteins are key regulators in the transduction of extracellular signals both in animals and plants. In plants, heterotrimeric G protein signaling plays essential roles in development and in response to biotic and abiotic stress. However, over the last decade it has become clear that plants have unique mechanisms of G protein signaling. Although plants share most of the core components of heterotrimeric G proteins, some of them exhibit unusual properties compared to their animal counterparts. In addition, plants do not share functional GPCRs. Therefore the well-established paradigm of the animal G protein signaling cycle is not applicable in plants. In this review, we summarize recent insights into these unique mechanisms of G protein signaling in plants with special focus on the evident potential of G protein signaling as a target to modify developmental and physiological parameters important for yield increase. Copyright © 2016 Elsevier Ltd. All rights reserved.

  3. An Introductory Idea for Teaching Two-Component Phase Diagrams

    Science.gov (United States)

    Peckham, Gavin D.; McNaught, Ian J.

    2011-01-01

    The teaching of two-component phase diagrams has attracted little attention in this "Journal," and it is hoped that this article will make a useful contribution. Current physical chemistry textbooks describe two-component phase diagrams adequately, but do so in a piecemeal fashion one section at a time; first solid-liquid equilibria, then…

  4. Two-component micro injection moulding for hearing aid applications

    DEFF Research Database (Denmark)

    Islam, Aminul; Hansen, Hans Nørgaard; Marhöfer, David Maximilian

    2012-01-01

    Two-component (2k) injection moulding is an important process technique at the present state of technology, and it is growing rapidly in the field of precision micro moulding. Besides combining different material properties in the same product, two-component moulding can eliminate many assembly s...

  5. A minimal model for two-component dark matter

    Energy Technology Data Exchange (ETDEWEB)

    Esch, Sonja; Klasen, Michael; Yaguna, Carlos E. [Institut fuer theoretische Physik, Universitaet Muenster, Wilhelm-Klemm-Strasse 9,D-48149 Muenster (Germany)

    2015-07-01

    We propose and study a new minimal model for two-component dark matter. The model contains only three additional fields, one fermion and two scalars, all singlets under the Standard Model gauge group. Two of these fields, one fermion and one scalar, are odd under a Z{sub 2} symmetry that renders them simultaneously stable. Thus, both particles contribute to the observed dark matter density. This model resembles the union of the singlet scalar and the singlet fermionic models but it contains some new features of its own. We analyze in some detail its dark matter phenomenology. Regarding the relic density, the main novelty is the possible annihilation of one dark matter particle into the other, which can affect the predicted relic density in a significant way. Regarding dark matter detection, we identify a new contribution that can lead either to an enhancement or to a suppression of the spin-independent cross section for the scalar dark matter particle. Finally, we define a set of five benchmarks models compatible with all present bounds and examine their direct detection prospects at planned experiments. A generic feature of this model is that both particles give rise to observable signals in 1-ton direct detection experiments. In fact, such experiments will be able to probe even a subdominant dark matter component at the percent level.

  6. A minimal model for two-component dark matter

    Science.gov (United States)

    Esch, Sonja; Klasen, Michael; Yaguna, Carlos E.

    2014-09-01

    We propose and study a new minimal model for two-component dark matter. The model contains only three additional fields, one fermion and two scalars, all singlets under the Standard Model gauge group. Two of these fields, one fermion and one scalar, are odd under a Z 2 symmetry that renders them simultaneously stable. Thus, both particles contribute to the observed dark matter density. This model resembles the union of the singlet scalar and the singlet fermionic models but it contains some new features of its own. We analyze in some detail its dark matter phenomenology. Regarding the relic density, the main novelty is the possible annihilation of one dark matter particle into the other, which can affect the predicted relic density in a significant way. Regarding dark matter detection, we identify a new contribution that can lead either to an enhancement or to a suppression of the spin-independent cross section for the scalar dark matter particle. Finally, we define a set of five benchmarks models compatible with all present bounds and examine their direct detection prospects at planned experiments. A generic feature of this model is that both particles give rise to observable signals in 1-ton direct detection experiments. In fact, such experiments will be able to probe even a subdominant dark matter component at the percent level.

  7. A novel two-component system involved in secretion stress response in Streptomyces lividans.

    Directory of Open Access Journals (Sweden)

    Sonia Gullón

    Full Text Available BACKGROUND: Misfolded proteins accumulating outside the bacterial cytoplasmic membrane can interfere with the secretory machinery, hence the existence of quality factors to eliminate these misfolded proteins is of capital importance in bacteria that are efficient producers of secretory proteins. These bacteria normally use a specific two-component system to respond to the stress produced by the accumulation of the misfolded proteins, by activating the expression of HtrA-like proteases to specifically eliminate the incorrectly folded proteins. METHODOLOGY/PRINCIPAL FINDINGS: Overproduction of alpha-amylase in S. lividans causing secretion stress permitted the identification of a two-component system (SCO4156-SCO4155 that regulates three HtrA-like proteases which appear to be involved in secretion stress response. Mutants in each of the genes forming part of the two-genes operon that encodes the sensor and regulator protein components accumulated misfolded proteins outside the cell, strongly suggesting the involvement of this two-component system in the S. lividans secretion stress response. CONCLUSIONS/SIGNIFICANCE: To our knowledge this is the first time that a specific secretion stress response two-component system is found to control the expression of three HtrA-like protease genes in S. lividans, a bacterium that has been repeatedly used as a host for the synthesis of homologous and heterologous secretory proteins of industrial application.

  8. Identification of four nuclear transport signal-binding proteins that interact with diverse transport signals.

    Science.gov (United States)

    Yamasaki, L; Kanda, P; Lanford, R E

    1989-07-01

    The transport of proteins into the nucleus requires not only the presence of a nuclear transport signal on the targeted protein but also the signal recognition proteins and the nuclear pore translocation apparatus. Complicating the search for the signal recognition proteins is the fact that the nuclear transport signals identified share little obvious homology. In this study, synthetic peptides homologous to the nuclear transport signals from the simian virus 40 large T antigen, Xenopus oocyte nucleoplasmin, adenovirus E1A, and Saccharomyces cerevisiae MAT alpha 2 proteins were coupled to a UV-photoactivable cross-linker and iodinated for use in an in vitro cross-linking reaction with cellular lysates. Four proteins, p140, p100, p70, and p55, which specifically interacted with the nuclear transport signal peptides were identified. Unique patterns of reactivity were observed with closely related pairs of nuclear transport signal peptides. Competition experiments with labeled and unlabeled peptides demonstrated that heterologous signals were able to bind the same protein and suggested that diverse signals use a common transport pathway. The subcellular distribution of the four nuclear transport signal-binding proteins suggested that nuclear transport involves both cytoplasmic and nuclear receptors. The four proteins were not bound by wheat germ agglutinin and were not associated tightly with the nuclear pore complex.

  9. LocSigDB: a database of protein localization signals

    OpenAIRE

    Negi, Simarjeet; Pandey, Sanjit; Srinivasan, Satish M; Mohammed, Akram; Guda, Chittibabu

    2015-01-01

    LocSigDB (http://genome.unmc.edu/LocSigDB/) is a manually curated database of experimental protein localization signals for eight distinct subcellular locations; primarily in a eukaryotic cell with brief coverage of bacterial proteins. Proteins must be localized at their appropriate subcellular compartment to perform their desired function. Mislocalization of proteins to unintended locations is a causative factor for many human diseases; therefore, collection of known sorting signals will hel...

  10. Bacillus subtilis Two-Component System Sensory Kinase DegS Is Regulated by Serine Phosphorylation in Its Input Domain

    DEFF Research Database (Denmark)

    Jers, Carsten; Kobir, Ahasanul; Søndergaard, Elsebeth Oline;

    2011-01-01

    Bacillus subtilis two-component system DegS/U is well known for the complexity of its regulation. The cytosolic sensory kinase DegS does not receive a single predominant input signal like most two-component kinases, instead it integrates a wide array of metabolic inputs that modulate its activity...

  11. The essential YycFG two-component system controls cell wall metabolism in Bacillus subtilis

    DEFF Research Database (Denmark)

    Bisicchia, Paola; Noone, David; Lioliou, Efthimia

    2007-01-01

    Adaptation of bacteria to the prevailing environmental and nutritional conditions is often mediated by two-component signal transduction systems (TCS). The Bacillus subtilis YycFG TCS has attracted special attention as it is essential for viability and its regulon is poorly defined. Here we show...

  12. Biased and g protein-independent signaling of chemokine receptors

    DEFF Research Database (Denmark)

    Steen, Anne; Larsen, Olav; Thiele, Stefanie

    2014-01-01

    ), different receptors (with the same ligand), or different tissues or cells (for the same ligand-receptor pair). Most often biased signaling is differentiated into G protein-dependent and β-arrestin-dependent signaling. Yet, it may also cover signaling differences within these groups. Moreover, it may...

  13. Regulator of G-protein signaling - 5 (RGS5 is a novel repressor of hedgehog signaling.

    Directory of Open Access Journals (Sweden)

    William M Mahoney

    Full Text Available Hedgehog (Hh signaling plays fundamental roles in morphogenesis, tissue repair, and human disease. Initiation of Hh signaling is controlled by the interaction of two multipass membrane proteins, patched (Ptc and smoothened (Smo. Recent studies identify Smo as a G-protein coupled receptor (GPCR-like protein that signals through large G-protein complexes which contain the Gαi subunit. We hypothesize Regulator of G-Protein Signaling (RGS proteins, and specifically RGS5, are endogenous repressors of Hh signaling via their ability to act as GTPase activating proteins (GAPs for GTP-bound Gαi, downstream of Smo. In support of this hypothesis, we demonstrate that RGS5 over-expression inhibits sonic hedgehog (Shh-mediated signaling and osteogenesis in C3H10T1/2 cells. Conversely, signaling is potentiated by siRNA-mediated knock-down of RGS5 expression, but not RGS4 expression. Furthermore, using immuohistochemical analysis and co-immunoprecipitation (Co-IP, we demonstrate that RGS5 is present with Smo in primary cilia. This organelle is required for canonical Hh signaling in mammalian cells, and RGS5 is found in a physical complex with Smo in these cells. We therefore conclude that RGS5 is an endogenous regulator of Hh-mediated signaling and that RGS proteins are potential targets for novel therapeutics in Hh-mediated diseases.

  14. Structure and Function of Vps15 in the Endosomal G Protein Signaling Pathway

    Energy Technology Data Exchange (ETDEWEB)

    Heenan, Erin J.; Vanhooke, Janeen L.; Temple, Brenda R.; Betts, Laurie; Sondek, John E.; Dohlman, Henrik G.; (UNC)

    2009-09-11

    G protein-coupled receptors mediate cellular responses to a wide variety of stimuli, including taste, light, and neurotransmitters. In the yeast Saccharomyces cerevisiae, activation of the pheromone pathway triggers events leading to mating. The view had long been held that the G protein-mediated signal occurs principally at the plasma membrane. Recently, it has been shown that the G protein {alpha} subunit Gpa1 can promote signaling at endosomes and requires two components of the sole phosphatidylinositol-3-kinase in yeast, Vps15 and Vps34. Vps15 contains multiple WD repeats and also binds to Gpa1 preferentially in the GDP-bound state; these observations led us to hypothesize that Vps15 may function as a G protein {beta} subunit at the endosome. Here we show an X-ray crystal structure of the Vps15 WD domain that reveals a seven-bladed propeller resembling that of typical G{beta} subunits. We show further that the WD domain is sufficient to bind Gpa1 as well as to Atg14, a potential G{gamma} protein that exists in a complex with Vps15. The Vps15 kinase domain together with the intermediate domain (linking the kinase and WD domains) also contributes to Gpa1 binding and is necessary for Vps15 to sustain G protein signaling. These findings reveal that the Vps15 G{beta}-like domain serves as a scaffold to assemble Gpa1 and Atg14, whereas the kinase and intermediate domains are required for proper signaling at the endosome.

  15. LocSigDB: a database of protein localization signals.

    Science.gov (United States)

    Negi, Simarjeet; Pandey, Sanjit; Srinivasan, Satish M; Mohammed, Akram; Guda, Chittibabu

    2015-01-01

    LocSigDB (http://genome.unmc.edu/LocSigDB/) is a manually curated database of experimental protein localization signals for eight distinct subcellular locations; primarily in a eukaryotic cell with brief coverage of bacterial proteins. Proteins must be localized at their appropriate subcellular compartment to perform their desired function. Mislocalization of proteins to unintended locations is a causative factor for many human diseases; therefore, collection of known sorting signals will help support many important areas of biomedical research. By performing an extensive literature study, we compiled a collection of 533 experimentally determined localization signals, along with the proteins that harbor such signals. Each signal in the LocSigDB is annotated with its localization, source, PubMed references and is linked to the proteins in UniProt database along with the organism information that contain the same amino acid pattern as the given signal. From LocSigDB webserver, users can download the whole database or browse/search for data using an intuitive query interface. To date, LocSigDB is the most comprehensive compendium of protein localization signals for eight distinct subcellular locations. Database URL: http://genome.unmc.edu/LocSigDB/

  16. Heterotrimeric G protein-coupled signaling in plants.

    Science.gov (United States)

    Urano, Daisuke; Jones, Alan M

    2014-01-01

    Investigators studying G protein-coupled signaling--often called the best-understood pathway in the world owing to intense research in medical fields--have adopted plants as a new model to explore the plasticity and evolution of G signaling. Much research on plant G signaling has not disappointed. Although plant cells have most of the core elements found in animal G signaling, differences in network architecture and intrinsic properties of plant G protein elements make G signaling in plant cells distinct from the animal paradigm. In contrast to animal G proteins, plant G proteins are self-activating, and therefore regulation of G activation in plants occurs at the deactivation step. The self-activating property also means that plant G proteins do not need and therefore do not have typical animal G protein-coupled receptors. Targets of activated plant G proteins, also known as effectors, are unlike effectors in animal cells. The simpler repertoire of G signal elements in Arabidopsis makes G signaling easier to manipulate in a multicellular context.

  17. Prioritization of a plant polysaccharide over a mucus carbohydrate is enforced by a Bacteroides hybrid two-component system.

    Science.gov (United States)

    Lynch, Jonathan B; Sonnenburg, Justin L

    2012-08-01

    Bacteroides is a dominant genus within the intestinal microbiota of healthy humans. Key adaptations of the Bacteroides to the dynamic intestinal ecosystem include a diverse repertoire of genes involved in sensing and processing numerous diet- and host-derived polysaccharides. One such adaptation is the carbohydrate-sensing hybrid two-component system (HTCS) family of signalling sensors, which has been widely expanded within the Bacteroides. Using Bacteroides thetaiotaomicron as a model, we have created a chimeric HTCS consisting of the well-characterized sensing domain of one HTCS, BT1754, and the regulatory domain of another HTCS, BT0366, to explore the regulatory capabilities of these molecules. We found that the BT0366 regulatory region directly binds to and mediates induction of the adjacent polysaccharide utilization locus (PUL) using whole-genome transcriptional profiling after inducing signalling through our chimeric protein. We also found that BT0366 activation simultaneously leads to repression of distal PULs involved in mucus carbohydrate consumption. These results suggest a novel mechanism by which an HTCS enforces a nutrient hierarchy within the Bacteroides via induction and repression of multiple PULs. Thus, hybrid two-component systems provide a mechanism for prioritizing consumption of carbohydrates through simultaneous binding and regulation of multiple polysaccharide utilization loci. © 2012 Blackwell Publishing Ltd.

  18. Circulation Condition of Two-component Bose-Einstein Condensate

    Institute of Scientific and Technical Information of China (English)

    2001-01-01

    In the report we point out that there exists an intrinsic difference in the internal symmetry of the two components spin-1/2 Bose condensates from that of spinor Bose condensates of the atoms with hyperfine states of nonzero integer-spins,which gives rise to a new topological constrain on the circulation for this two-component spin-1/2 Bose condensates.It is shown that the SU(2) symmetry of the spin-1/2 Bose condensate implies a

  19. Itinerant Ferromagnetism in a Polarized Two-Component Fermi Gas

    DEFF Research Database (Denmark)

    Massignan, Pietro; Yu, Zhenhua; Bruun, Georg

    2013-01-01

    We analyze when a repulsively interacting two-component Fermi gas becomes thermodynamically unstable against phase separation. We focus on the strongly polarized limit, where the free energy of the homogeneous mixture can be calculated accurately in terms of well-defined quasiparticles, the repul......We analyze when a repulsively interacting two-component Fermi gas becomes thermodynamically unstable against phase separation. We focus on the strongly polarized limit, where the free energy of the homogeneous mixture can be calculated accurately in terms of well-defined quasiparticles...

  20. Regulation, Signaling, and Physiological Functions of G-Proteins.

    Science.gov (United States)

    Syrovatkina, Viktoriya; Alegre, Kamela O; Dey, Raja; Huang, Xin-Yun

    2016-09-25

    Heterotrimeric guanine-nucleotide-binding regulatory proteins (G-proteins) mainly relay the information from G-protein-coupled receptors (GPCRs) on the plasma membrane to the inside of cells to regulate various biochemical functions. Depending on the targeted cell types, tissues, and organs, these signals modulate diverse physiological functions. The basic schemes of heterotrimeric G-proteins have been outlined. In this review, we briefly summarize what is known about the regulation, signaling, and physiological functions of G-proteins. We then focus on a few less explored areas such as the regulation of G-proteins by non-GPCRs and the physiological functions of G-proteins that cannot be easily explained by the known G-protein signaling pathways. There are new signaling pathways and physiological functions for G-proteins to be discovered and further interrogated. With the advancements in structural and computational biological techniques, we are closer to having a better understanding of how G-proteins are regulated and of the specificity of G-protein interactions with their regulators.

  1. A Signal Processing Method to Explore Similarity in Protein Flexibility

    Directory of Open Access Journals (Sweden)

    Simina Vasilache

    2010-01-01

    Full Text Available Understanding mechanisms of protein flexibility is of great importance to structural biology. The ability to detect similarities between proteins and their patterns is vital in discovering new information about unknown protein functions. A Distance Constraint Model (DCM provides a means to generate a variety of flexibility measures based on a given protein structure. Although information about mechanical properties of flexibility is critical for understanding protein function for a given protein, the question of whether certain characteristics are shared across homologous proteins is difficult to assess. For a proper assessment, a quantified measure of similarity is necessary. This paper begins to explore image processing techniques to quantify similarities in signals and images that characterize protein flexibility. The dataset considered here consists of three different families of proteins, with three proteins in each family. The similarities and differences found within flexibility measures across homologous proteins do not align with sequence-based evolutionary methods.

  2. Post-translational modification of PII signal transduction proteins

    OpenAIRE

    Mike eMerrick

    2015-01-01

    The PII proteins constitute one of the most widely distributed families of signal transduction proteins in nature. They are pivotal players in the control of nitrogen metabolism in bacteria and archaea, and are also found in the plastids of plants. Quite remarkably PII proteins control the activities of a diverse range of enzymes, transcription factors and membrane transport proteins, and in all known cases they achieve their regulatory effect by direct interaction with their target. PII prot...

  3. Closing the loop: The PmrA/PmrB two-component system negatively controls expression of its posttranscriptional activator PmrD

    OpenAIRE

    Kato, Akinori; Latifi, Tammy; Groisman, Eduardo A.

    2003-01-01

    A fundamental question in biology is how an organism integrates multiple signals to mediate an appropriate cellular response. The PmrA/PmrB two-component system of Salmonella enterica can be activated independently by Fe3+, which is sensed by the PmrB protein, and in low Mg2+, which is sensed by the PhoQ protein. The low-Mg2+ activation requires pmrD, a PhoP/PhoQ-activated gene that activates the response regulator PmrA at a posttranscriptional level. We now report that pmrD expression is neg...

  4. Two component permeation through thin zeolite MFI membranes

    NARCIS (Netherlands)

    Keizer, K.; Burggraaf, A.J.; Vroon, Z.A.E.P.; Verweij, H.

    1998-01-01

    Two component permeation measurements have been performed by the Wicke-Kallenbach method on a thin (3 μm) zeolite MFI (Silicalite-1) membrane with molecules of different kinetic diameters, d(k). The membrane was supported by a flat porous α-Al2O3 substrate. The results obtained could be classified i

  5. two component permeation through thin zeolite MFI membranes

    NARCIS (Netherlands)

    Keizer, Klaas; Burggraaf, Anthonie; Burggraaf, A.J.; Vroon, Z.A.E.P.; Vroon, Z.A.E.P.; Verweij, H.

    1998-01-01

    Two component permeation measurements have been performed by the Wicke–Kallenbach method on a thin (3 μm) zeolite MFI (Silicalite-1) membrane with molecules of different kinetic diameters, dk. The membrane was supported by a flat porous -Al2O3 substrate. The results obtained could be classified in s

  6. TWO-COMPONENT JETS AND THE FANAROFF-RILEY DICHOTOMY

    NARCIS (Netherlands)

    Meliani, Z.; Keppens, R.; Sauty, C.

    2010-01-01

    Transversely stratified jets are observed in many classes of astrophysical objects, ranging from young stellar objects, mu-quasars, to active galactic nuclei and even in gamma-ray bursts. Theoretical arguments support this transverse stratification of jets with two components induced by intrinsic fe

  7. Two component injection moulding: Present and future perspectives

    DEFF Research Database (Denmark)

    Islam, Aminul; Hansen, Hans Nørgaard

    2009-01-01

    Two component injection moulding has widespread industrial applications. Still the technology is yet to gain its full potential in highly demanding and technically challenging applications areas. The smart use of this technology can open the doors for cost effective and convergent manufacturing...

  8. Entanglement Properties in Two-Component Bose-Einstein Condensate

    Science.gov (United States)

    Jiang, Di-You

    2016-10-01

    We investigate entanglement inseparability and bipartite entanglement of in two-component Bose-Einstein condensate in the presence of the nonlinear interatomic interaction, interspecies interaction. Entanglement inseparability and bipartite entanglement have the similar properties. More entanglement can be generated by adjusting the nonlinear interatomic interaction and control the time interval of the entanglement by adjusting interspecies interaction.

  9. Goal-Directed Aiming: Two Components but Multiple Processes

    Science.gov (United States)

    Elliott, Digby; Hansen, Steve; Grierson, Lawrence E. M.; Lyons, James; Bennett, Simon J.; Hayes, Spencer J.

    2010-01-01

    This article reviews the behavioral literature on the control of goal-directed aiming and presents a multiple-process model of limb control. The model builds on recent variants of Woodworth's (1899) two-component model of speed-accuracy relations in voluntary movement and incorporates ideas about dynamic online limb control based on prior…

  10. Stenotrophomonas maltophilia PhoP, a Two-Component Response Regulator, Involved in Antimicrobial Susceptibilities.

    Directory of Open Access Journals (Sweden)

    Ming-Che Liu

    Full Text Available Stenotrophomonas maltophilia, a gram-negative bacterium, has increasingly emerged as an important nosocomial pathogen. It is well-known for resistance to a variety of antimicrobial agents including cationic antimicrobial polypeptides (CAPs. Resistance to polymyxin B, a kind of CAPs, is known to be controlled by the two-component system PhoPQ. To unravel the role of PhoPQ in polymyxin B resistance of S. maltophilia, a phoP mutant was constructed. We found MICs of polymyxin B, chloramphenicol, ampicillin, gentamicin, kanamycin, streptomycin and spectinomycin decreased 2-64 fold in the phoP mutant. Complementation of the phoP mutant by the wild-type phoP gene restored all of the MICs to the wild type levels. Expression of PhoP was shown to be autoregulated and responsive to Mg2+ levels. The polymyxin B and gentamicin killing tests indicated that pretreatment of low Mg2+ can protect the wild-type S. maltophilia from killing but not phoP mutant. Interestingly, we found phoP mutant had a decrease in expression of SmeZ, an efflux transporter protein for aminoglycosides in S. maltophilia. Moreover, phoP mutant showed increased permeability in the cell membrane relative to the wild-type. In summary, we demonstrated the two-component regulator PhoP of S. maltophilia is involved in antimicrobial susceptibilities and low Mg2+ serves as a signal for triggering the pathway. Both the alteration in membrane permeability and downregulation of SmeZ efflux transporter in the phoP mutant contributed to the increased drug susceptibilities of S. maltophilia, in particular for aminoglycosides. This is the first report to describe the role of the Mg2+-sensing PhoP signaling pathway of S. maltophilia in regulation of the SmeZ efflux transporter and in antimicrobial susceptibilities. This study suggests PhoPQ TCS may serve as a target for development of antimicrobial agents against multidrug-resistant S. maltophilia.

  11. A two component system is involved in acid adaptation of Lactobacillus delbrueckii subsp. bulgaricus.

    Science.gov (United States)

    Cui, Yanhua; Liu, Wei; Qu, Xiaojun; Chen, Zhangting; Zhang, Xu; Liu, Tong; Zhang, Lanwei

    2012-05-20

    The Gram-positive bacterium Lactobacillus delbrueckii subsp. bulgaricus is of vital importance to the food industry, especially to the dairy industry. Two component systems (TCSs) are one of the most important mechanisms for environmental sensing and signal transduction in the majority of Gram-positive and Gram-negative bacteria. A typical TCS consists of a histidine protein kinase (HPK) and a cytoplasmic response regulator (RR). To investigate the functions of TCSs during acid adaptation in L. bulgaricus, we used quantitative PCR to reveal how TCSs expression changes during acid adaptation. Two TCSs (JN675228/JN675229 and JN675230/JN675231) and two HPKs (JN675236 and JN675240) were induced during acid adaptation. These TCSs were speculated to be related with the acid adaptation ability of L. bulgaricus. The mutants of JN675228/JN675229 were constructed in order to investigate the functions of JN675228/JN675229. The mutants showed reduced acid adaptation compared to that of wild type, and the complemented strains were similar to the wild-type strain. These observations suggested that JN675228 and JN675229 were involved in acid adaptation in L. bulgaricus. The interaction between JN675228 and JN675229 was identified by means of yeast two-hybrid system. The results indicated there is interaction between JN675228 and JN675229.

  12. Structural studies of the activation of the two component receiver domain NTRC by multidimensional heteronuclear NMR

    Energy Technology Data Exchange (ETDEWEB)

    Nohaile, M J [Univ. of California, Berkeley, CA (United States). Dept. of Chemistry

    1996-05-01

    Multidimensional heteronuclear NMR spectroscopy was used to investigate the N-terminal domain of the transcriptional enhancer NTRC (NiTrogen Regulatory protein C). This domain belongs to the family of receiver domains of two-component regulatory systems involved in signal transduction. Phosphorylation of NTRC at D54 leads to an activated form of the molecule which stimulates transcription of genes involved in nitrogen regulation. Three and four dimensional NMR techniques were used to determine an intermediate resolution structure of the unphosphorylated, inactive form of the N-terminal domain of NTRC. The structure is comprised of five {alpha}-helices and a five-stranded {beta}-sheet in a ({beta}/{alpha}){sub 5} topology. Analysis of the backbone dynamics of NTRC indicate that helix 4 and strand 5 are significantly more flexible than the rest of the secondary structure of the protein and that the loops making up the active site are flexible. The short lifetime of phospho-NTRC hampers the study of this form. However, conditions for determining the resonance assignments and, possibly, the three dimensional structure of phosphorylated NTRC have been obtained. Tentative assignments of the phosphorylated form indicate that the majority of the changes that NTRC experiences upon phosphorylation occur in helix 3, strand 4, helix 4, strand 5, and the loop between strand 5 and helix 5 (the 3445 face of NTRC) as well as near the site of phosphorylation. In order to examine a stable, activated form of the protein, constitutively active mutants of NTRC were investigated.

  13. Structural studies of the activation of the two component receiver domain NTRC by multidimensional heteronuclear NMR

    Energy Technology Data Exchange (ETDEWEB)

    Nohaile, Michael James [Univ. of California, Berkeley, CA (United States). Dept. of Chemistry

    1996-05-01

    Multidimensional heteronuclear NMR spectroscopy was used to investigate the N-terminal domain of the transcriptional enhancer NTRC (NiTrogen Regulatory protein C). This domain belongs to the family of receiver domains of two-component regulatory systems involved in signal transduction. Phosphorylation of NTRC at D54 leads to an activated form of the molecule which stimulates transcription of genes involved in nitrogen regulation. Three and four dimensional NMR techniques were used to determine an intermediate resolution structure of the unphosphorylated, inactive form of the N-terminal domain of NTRC. The structure is comprised of five α-helices and a five-stranded β-sheet in a (β/α)5 topology. Analysis of the backbone dynamics of NTRC indicate that helix 4 and strand 5 are significantly more flexible than the rest of the secondary structure of the protein and that the loops making up the active site are flexible. The short lifetime of phospho-NTRC hampers the study of this form. However, conditions for determining the resonance assignments and, possibly, the three dimensional structure of phosphorylated NTRC have been obtained. Tentative assignments of the phosphorylated form indicate that the majority of the changes that NTRC experiences upon phosphorylation occur in helix 3, strand 4, helix 4, strand 5, and the loop between strand 5 and helix 5 (the 3445 face of NTRC) as well as near the site of phosphorylation. In order to examine a stable, activated form of the protein, constitutively active mutants of NTRC were investigated.

  14. Activation of the Campylobacter jejuni FlgSR two-component system is linked to the flagellar export apparatus.

    Science.gov (United States)

    Joslin, Stephanie N; Hendrixson, David R

    2009-04-01

    Activation of sigma(54)-dependent gene expression essential for formation of flagella in Campylobacter jejuni requires the components of the inner membrane-localized flagellar export apparatus and the FlgSR two-component regulatory system. In this study, we characterized the FlgS sensor kinase and how activation of the protein is linked to the flagellar export apparatus. We found that FlgS is localized to the C. jejuni cytoplasm and that His141 of FlgS is essential for autophosphorylation, phosphorelay to the cognate FlgR response regulator, motility, and expression of sigma(54)-dependent flagellar genes. Mutants with incomplete flagellar export apparatuses produced wild-type levels of FlgS and FlgR, but they were defective for signaling through the FlgSR system. By using genetic approaches, we found that FlgSR activity is linked to and downstream of the flagellar export apparatus in a regulatory cascade that terminates in expression of sigma(54)-dependent flagellar genes. By analyzing defined flhB and fliI mutants of C. jejuni that form flagellar export apparatuses that are secretion incompetent, we determined that formation of the apparatus is required to contribute to the signal sensed by FlgS to terminate in activation of expression of sigma(54)-dependent flagellar genes. Considering that the flagellar export apparatuses of Escherichia coli and Salmonella species influence sigma(28)-dependent flagellar gene expression, our work expands the signaling activity of the apparatuses to include sigma(54)-dependent pathways of C. jejuni and possibly other motile bacteria. This study indicates that these apparatuses have broader functions beyond flagellar protein secretion, including activation of essential two-component regulatory systems required for expression of sigma(54)-dependent flagellar genes.

  15. Heart Failure Protein May Signal Early Brain Damage

    Science.gov (United States)

    ... page: https://medlineplus.gov/news/fullstory_162447.html Heart Failure Protein May Signal Early Brain Damage Higher levels ... stress. Blood levels of NT-proBNP rise when heart failure worsens and fall when it gets better. Previous ...

  16. Dynamic phospholipid signaling by G protein-coupled receptors

    NARCIS (Netherlands)

    Weernink, Paschal A. Oude; Han, Li; Jakobs, Karl H.; Schmidt, Martina

    2007-01-01

    G protein-coupled receptors (GPCRs) control a variety of fundamental cellular processes by regulating phospholipid signaling pathways. Essential for signaling by a large number of receptors is the hydrolysis of the membrane phosphoinositide PIP2 by phospholipase C (PLC) into the second messengers IP

  17. Fluorescopic evaluation of protein-lipid relations in cellular signalling.

    NARCIS (Netherlands)

    Pap, E.H.W.

    1994-01-01

    IntroductionCellular communication is partly mediated through the modulation of protein activity, structure and dynamics by lipids. In contrast to the biochemical aspects of lipid signalling, relatively little is known about the physical properties of the "signal" lipids (lipids involved in cellular

  18. PACRG, a protein linked to ciliary motility, mediates cellular signaling.

    Science.gov (United States)

    Loucks, Catrina M; Bialas, Nathan J; Dekkers, Martijn P J; Walker, Denise S; Grundy, Laura J; Li, Chunmei; Inglis, P Nick; Kida, Katarzyna; Schafer, William R; Blacque, Oliver E; Jansen, Gert; Leroux, Michel R

    2016-07-01

    Cilia are microtubule-based organelles that project from nearly all mammalian cell types. Motile cilia generate fluid flow, whereas nonmotile (primary) cilia are required for sensory physiology and modulate various signal transduction pathways. Here we investigate the nonmotile ciliary signaling roles of parkin coregulated gene (PACRG), a protein linked to ciliary motility. PACRG is associated with the protofilament ribbon, a structure believed to dictate the regular arrangement of motility-associated ciliary components. Roles for protofilament ribbon-associated proteins in nonmotile cilia and cellular signaling have not been investigated. We show that PACRG localizes to a small subset of nonmotile cilia in Caenorhabditis elegans, suggesting an evolutionary adaptation for mediating specific sensory/signaling functions. We find that it influences a learning behavior known as gustatory plasticity, in which it is functionally coupled to heterotrimeric G-protein signaling. We also demonstrate that PACRG promotes longevity in C. elegans by acting upstream of the lifespan-promoting FOXO transcription factor DAF-16 and likely upstream of insulin/IGF signaling. Our findings establish previously unrecognized sensory/signaling functions for PACRG and point to a role for this protein in promoting longevity. Furthermore, our work suggests additional ciliary motility-signaling connections, since EFHC1 (EF-hand containing 1), a potential PACRG interaction partner similarly associated with the protofilament ribbon and ciliary motility, also positively regulates lifespan.

  19. Mitogen-activated protein kinase signaling in plants

    DEFF Research Database (Denmark)

    Rodriguez, Maria Cristina Suarez; Petersen, Morten; Mundy, John

    2010-01-01

    Eukaryotic mitogen-activated protein kinase (MAPK) cascades have evolved to transduce environmental and developmental signals into adaptive and programmed responses. MAPK cascades relay and amplify signals via three types of reversibly phosphorylated kinases leading to the phosphorylation of subs...... the Arabidopsis thaliana MAPKs MPK3, 4, and 6 and MAP2Ks MKK1, 2, 4, and 5. Future work needs to focus on identifying substrates of MAPKs, and on understanding how specificity is achieved among MAPK signaling pathways....

  20. A two-component NZRI metamaterial based rectangular cloak

    Science.gov (United States)

    Islam, Sikder Sunbeam; Faruque, Mohammd Rashed Iqbal; Islam, Mohammad Tariqul

    2015-10-01

    A new two-component, near zero refractive index (NZRI) metamaterial is presented for electromagnetic rectangular cloaking operation in the microwave range. In the basic design a pi-shaped, metamaterial was developed and its characteristics were investigated for the two major axes (x and z-axis) wave propagation through the material. For the z-axis wave propagation, it shows more than 2 GHz bandwidth and for the x-axis wave propagation; it exhibits more than 1 GHz bandwidth of NZRI property. The metamaterial was then utilized in designing a rectangular cloak where a metal cylinder was cloaked perfectly in the C-band area of microwave regime. The experimental result was provided for the metamaterial and the cloak and these results were compared with the simulated results. This is a novel and promising design for its two-component NZRI characteristics and rectangular cloaking operation in the electromagnetic paradigm.

  1. On a periodic two-component Hunter-Saxton equation

    CERN Document Server

    Kohlmann, Martin

    2011-01-01

    We determine the solution of the geodesic equation associated with a periodic two-component Hunter-Saxton system on a semidirect product obtained from the diffeomorphism group of the circle, modulo rigid rotations, and a space of scalar functions. In particular, we compute the time of breakdown of the geodesic flow. As a further goal, we establish a local well-posedness result for the two-component Hunter-Saxton system in the smooth category. The paper gets in line with some recent results for the generalized Hunter-Saxton equation provided by Escher, Wu and Wunsch in [J. Escher, Preprint 2010] and [H. Wu, M. Wunsch, arXiv:1009.1688v1 [math.AP

  2. Two Component Injection Moulding for Moulded Interconnect Devices

    DEFF Research Database (Denmark)

    Islam, Aminul

    The moulded interconnect devices (MIDs) contain huge possibilities for many applications in micro electro-mechanical-systems because of their potential in reducing the number of components, process steps and finally in miniaturization of the product. Among the available MID process chains, two...... component (2k) injection moulding is one of the most industrially adaptive processes. However, the use of two component injection moulding for MID fabrication, with circuit patterns in sub-millimeter range, is still a big challenge. This book searches for the technical difficulties associated...... with the process and makes attempts to overcome those challenges. In search of suitable polymer materials for MID applications, potential materials are characterized in terms of polymer-polymer bond strength, polymer-polymer interface quality and selective metallization. The experimental results find the factors...

  3. Two-component microinjection moulding for MID fabrication

    DEFF Research Database (Denmark)

    Islam, Aminul; Hansen, Hans Nørgaard; Tang, Peter Torben

    2010-01-01

    Moulded interconnect devices (MIDs) are plastic substrates with electrical infrastructure. The fabrication of MIDs is usually based on injection moulding, and different process chains may be identified from this starting point. The use of MIDs has been driven primarily by the automotive sector......, but recently, the medical sector seems more and more interested. In particular, the possibility of miniaturisation of three-dimensional components with electrical infrastructure is attractive. The present paper describes possible manufacturing routes and challenges of miniaturised MIDs based on two......-component injection moulding and subsequent metallisation. This technology promises cost effective and convergent manufacturing approaches for both macro- and microapplications. This paper presents the results of industrial MID production based on two-component injection moulding and discusses the important issues...

  4. Cytoplasmic tail of coronavirus spike protein has intracellular targeting signals

    Indian Academy of Sciences (India)

    JIBIN SADASIVAN; MANMEET SINGH; JAYASRI DAS SARMA

    2017-06-01

    Intracellular trafficking and localization studies of spike protein from SARS and OC43 showed that SARS spikeprotein is localized in the ER or ERGIC compartment and OC43 spike protein is predominantly localized in thelysosome. Differential localization can be explained by signal sequence. The sequence alignment using Clustal Wshows that the signal sequence present at the cytoplasmic tail plays an important role in spike protein localization. Aunique GYQEL motif is identified at the cytoplasmic terminal of OC43 spike protein which helps in localization in thelysosome, and a novel KLHYT motif is identified in the cytoplasmic tail of SARS spike protein which helps in ER orERGIC localization. This study sheds some light on the role of cytoplasmic tail of spike protein in cell-to-cell fusion,coronavirus host cell fusion and subsequent pathogenicity.

  5. Heart failure-specific changes in protein kinase signalling.

    Science.gov (United States)

    Lorenz, Kristina; Stathopoulou, Konstantina; Schmid, Evelyn; Eder, Petra; Cuello, Friederike

    2014-06-01

    Among the myriad of molecular alterations occurring in heart failure development, aggravation of the disease is often attributed to global or local changes in protein kinase activity, thus making protein kinases attractive targets for therapeutic intervention. Since protein kinases do not only have maladaptive roles, but also contribute to the physiological integrity of cells, it is a challenging task to circumvent undesired inhibition of protein kinase activity. Identification of posttranslational modifications and/or protein-protein interactions that are exclusively apparent under pathophysiological conditions provides exciting information for alternative non-kinase inhibitory treatment strategies that eliminate maladaptive functions of a protein kinase, but preserve the beneficial ones. Here, we focus on the disease-specific regulation of a number of protein kinases, namely, Ca(2+)/calmodulin-dependent protein kinase II isoform δ (CaMKIIδ), G protein-coupled receptor kinase 2 (GRK2), extracellular signal-regulated kinase 1 and 2 (ERK1/2), protein kinase D (PKD) and protein kinase C isoform β2 (PKCβ2), which are embedded in complex signal transduction pathways implicated in heart failure development, and discuss potential avenues for novel treatment strategies to combat heart disease.

  6. Calprotectin Increases the Activity of the SaeRS Two Component System and Murine Mortality during Staphylococcus aureus Infections.

    Directory of Open Access Journals (Sweden)

    Hoonsik Cho

    2015-07-01

    Full Text Available Calprotectin, the most abundant cytoplasmic protein in neutrophils, suppresses the growth of Staphylococcus aureus by sequestering the nutrient metal ions Zn and Mn. Here we show that calprotectin can also enhance the activity of the SaeRS two component system (TCS, a signaling system essential for production of over 20 virulence factors in S. aureus. The activity of the SaeRS TCS is repressed by certain divalent ions found in blood or neutrophil granules; however, the Zn bound-form of calprotectin relieves this repression. During staphylococcal encounter with murine neutrophils or staphylococcal infection of the murine peritoneal cavity, calprotectin increases the activity of the SaeRS TCS as well as the production of proinflammatory cytokines such as IL-1β and TNF-α, resulting in higher murine mortality. These results suggest that, under certain conditions, calprotectin can be exploited by S. aureus to increase bacterial virulence and host mortality.

  7. Calprotectin Increases the Activity of the SaeRS Two Component System and Murine Mortality during Staphylococcus aureus Infections.

    Science.gov (United States)

    Cho, Hoonsik; Jeong, Do-Won; Liu, Qian; Yeo, Won-Sik; Vogl, Thomas; Skaar, Eric P; Chazin, Walter J; Bae, Taeok

    2015-07-01

    Calprotectin, the most abundant cytoplasmic protein in neutrophils, suppresses the growth of Staphylococcus aureus by sequestering the nutrient metal ions Zn and Mn. Here we show that calprotectin can also enhance the activity of the SaeRS two component system (TCS), a signaling system essential for production of over 20 virulence factors in S. aureus. The activity of the SaeRS TCS is repressed by certain divalent ions found in blood or neutrophil granules; however, the Zn bound-form of calprotectin relieves this repression. During staphylococcal encounter with murine neutrophils or staphylococcal infection of the murine peritoneal cavity, calprotectin increases the activity of the SaeRS TCS as well as the production of proinflammatory cytokines such as IL-1β and TNF-α, resulting in higher murine mortality. These results suggest that, under certain conditions, calprotectin can be exploited by S. aureus to increase bacterial virulence and host mortality.

  8. A novel two-component response regulator links rpf with biofilm formation and virulence of Xanthomonas axonopodis pv. citri.

    Directory of Open Access Journals (Sweden)

    Tzu-Pi Huang

    Full Text Available Citrus bacterial canker caused by Xanthomonas axonopodis pv. citri is a serious disease that impacts citrus production worldwide, and X. axonopodis pv. citri is listed as a quarantine pest in certain countries. Biofilm formation is important for the successful development of a pathogenic relationship between various bacteria and their host(s. To understand the mechanisms of biofilm formation by X. axonopodis pv. citri strain XW19, the strain was subjected to transposon mutagenesis. One mutant with a mutation in a two-component response regulator gene that was deficient in biofilm formation on a polystyrene microplate was selected for further study. The protein was designated as BfdR for biofilm formation defective regulator. BfdR from strain XW19 shares 100% amino acid sequence identity with XAC1284 of X. axonopodis pv. citri strain 306 and 30-100% identity with two-component response regulators in various pathogens and environmental microorganisms. The bfdR mutant strain exhibited significantly decreased biofilm formation on the leaf surfaces of Mexican lime compared with the wild type strain. The bfdR mutant was also compromised in its ability to cause canker lesions. The wild-type phenotype was restored by providing pbfdR in trans in the bfdR mutant. Our data indicated that BfdR did not regulate the production of virulence-related extracellular enzymes including amylase, lipase, protease, and lecithinase or the expression of hrpG, rfbC, and katE; however, BfdR controlled the expression of rpfF in XVM2 medium, which mimics cytoplasmic fluids in planta. In conclusion, biofilm formation on leaf surfaces of citrus is important for canker development in X. axonopodis pv. citri XW19. The process is controlled by the two-component response regulator BfdR via regulation of rpfF, which is required for the biosynthesis of a diffusible signal factor.

  9. How fast does a signal propagate through proteins?

    Directory of Open Access Journals (Sweden)

    Hui T Young

    Full Text Available As the molecular basis of signal propagation in the cell, proteins are regulated by perturbations, such as mechanical forces or ligand binding. The question arises how fast such a signal propagates through the protein molecular scaffold. As a first step, we have investigated numerically the dynamics of force propagation through a single (Ala[Formula: see text] protein following a sudden increase in the stretching forces applied to its end termini. The force propagates along the backbone into the center of the chain on the picosecond scale. Both conformational and tension dynamics are found in good agreement with a coarse-grained theory of force propagation through semiflexible polymers. The speed of force propagation of [Formula: see text]50Å ps(-1 derived from these simulations is likely to determine an upper speed limit of mechanical signal transfer in allosteric proteins or molecular machines.

  10. Protein import into plant mitochondria: signals, machinery, processing, and regulation.

    Science.gov (United States)

    Murcha, Monika W; Kmiec, Beata; Kubiszewski-Jakubiak, Szymon; Teixeira, Pedro F; Glaser, Elzbieta; Whelan, James

    2014-12-01

    The majority of more than 1000 proteins present in mitochondria are imported from nuclear-encoded, cytosolically synthesized precursor proteins. This impressive feat of transport and sorting is achieved by the combined action of targeting signals on mitochondrial proteins and the mitochondrial protein import apparatus. The mitochondrial protein import apparatus is composed of a number of multi-subunit protein complexes that recognize, translocate, and assemble mitochondrial proteins into functional complexes. While the core subunits involved in mitochondrial protein import are well conserved across wide phylogenetic gaps, the accessory subunits of these complexes differ in identity and/or function when plants are compared with Saccharomyces cerevisiae (yeast), the model system for mitochondrial protein import. These differences include distinct protein import receptors in plants, different mechanistic operation of the intermembrane protein import system, the location and activity of peptidases, the function of inner-membrane translocases in linking the outer and inner membrane, and the association/regulation of mitochondrial protein import complexes with components of the respiratory chain. Additionally, plant mitochondria share proteins with plastids, i.e. dual-targeted proteins. Also, the developmental and cell-specific nature of mitochondrial biogenesis is an aspect not observed in single-celled systems that is readily apparent in studies in plants. This means that plants provide a valuable model system to study the various regulatory processes associated with protein import and mitochondrial biogenesis.

  11. Comparative Expression Analysis of Two-Component System Members in Arabidopsis and Oryza sativa under Abiotic Stress

    Directory of Open Access Journals (Sweden)

    Anupama eSingh

    2015-09-01

    Full Text Available Two component system (TCS is one of the key signal sensing machinery which enables species to sense environmental stimuli. It essentially comprises of three major components, sensory histidine kinase proteins (HKs, histidine phosphotransfer proteins (Hpts and response regulator proteins (RRs. The members of the TCS family have already been identified in Arabidopsis and rice but the knowledge about their functional indulgence during various abiotic stress conditions remains meagre. Current study is an attempt to carry out comprehensive analysis of the expression of TCS members in response to various abiotic stress conditions and in various plant tissues in Arabidopsis and rice using MPSS and publicly available microarray data. The analysis suggests that despite having almost similar number of genes, rice expresses higher number of TCS members during various abiotic stress conditions than Arabidopsis. We found that the TCS machinery is regulated by not only various abiotic stresses, but also by the tissue specificity. Analysis of expression of some representative members of TCS gene family showed their regulation by the diurnal cycle in rice seedlings, thus bringing-in another level of their transcriptional control. Thus, we report a highly complex and tight regulatory network of TCS members, as influenced by the tissue, abiotic stress signal and diurnal rhythm. The insights on the comparative expression analysis presented in this study may provide crucial leads towards dissection of diverse role(s of the various TCS family members in Arabidopsis and rice.

  12. Rap G protein signal in normal and disordered lymphohematopoiesis

    Energy Technology Data Exchange (ETDEWEB)

    Minato, Nagahiro, E-mail: minato@imm.med.kyoto-u.ac.jp

    2013-09-10

    Rap proteins (Rap1, Rap2a, b, c) are small molecular weight GTPases of the Ras family. Rap G proteins mediate diverse cellular events such as cell adhesion, proliferation, and gene activation through various signaling pathways. Activation of Rap signal is regulated tightly by several specific regulatory proteins including guanine nucleotide exchange factors and GTPase-activating proteins. Beyond cell biological studies, increasing attempts have been made in the past decade to define the roles of Rap signal in specific functions of normal tissue systems as well as in cancer. In the immune and hematopoietic systems, Rap signal plays crucial roles in the development and function of essentially all lineages of lymphocytes and hematopoietic cells, and importantly, deregulated Rap signal may lead to unique pathological conditions depending on the affected cell types, including various types of leukemia and autoimmunity. The phenotypical studies have unveiled novel, even unexpected functional aspects of Rap signal in cells from a variety of tissues, providing potentially important clues for controlling human diseases, including malignancy.

  13. Rap G protein signal in normal and disordered lymphohematopoiesis.

    Science.gov (United States)

    Minato, Nagahiro

    2013-09-10

    Rap proteins (Rap1, Rap2a, b, c) are small molecular weight GTPases of the Ras family. Rap G proteins mediate diverse cellular events such as cell adhesion, proliferation, and gene activation through various signaling pathways. Activation of Rap signal is regulated tightly by several specific regulatory proteins including guanine nucleotide exchange factors and GTPase-activating proteins. Beyond cell biological studies, increasing attempts have been made in the past decade to define the roles of Rap signal in specific functions of normal tissue systems as well as in cancer. In the immune and hematopoietic systems, Rap signal plays crucial roles in the development and function of essentially all lineages of lymphocytes and hematopoietic cells, and importantly, deregulated Rap signal may lead to unique pathological conditions depending on the affected cell types, including various types of leukemia and autoimmunity. The phenotypical studies have unveiled novel, even unexpected functional aspects of Rap signal in cells from a variety of tissues, providing potentially important clues for controlling human diseases, including malignancy. © 2013 Elsevier Inc. All rights reserved.

  14. Linking proteins to signaling pathways for experiment design and evaluation.

    Directory of Open Access Journals (Sweden)

    Illés J Farkas

    Full Text Available Biomedical experimental work often focuses on altering the functions of selected proteins. These changes can hit signaling pathways, and can therefore unexpectedly and non-specifically affect cellular processes. We propose PathwayLinker, an online tool that can provide a first estimate of the possible signaling effects of such changes, e.g., drug or microRNA treatments. PathwayLinker minimizes the users' efforts by integrating protein-protein interaction and signaling pathway data from several sources with statistical significance tests and clear visualization. We demonstrate through three case studies that the developed tool can point out unexpected signaling bias in normal laboratory experiments and identify likely novel signaling proteins among the interactors of known drug targets. In our first case study we show that knockdown of the Caenorhabditis elegans gene cdc-25.1 (meant to avoid progeny may globally affect the signaling system and unexpectedly bias experiments. In the second case study we evaluate the loss-of-function phenotypes of a less known C. elegans gene to predict its function. In the third case study we analyze GJA1, an anti-cancer drug target protein in human, and predict for this protein novel signaling pathway memberships, which may be sources of side effects. Compared to similar services, a major advantage of PathwayLinker is that it drastically reduces the necessary amount of manual literature searches and can be used without a computational background. PathwayLinker is available at http://PathwayLinker.org. Detailed documentation and source code are available at the website.

  15. Revisiting Apoplastic Auxin Signaling Mediated by AUXIN BINDING PROTEIN 1.

    Science.gov (United States)

    Feng, Mingxiao; Kim, Jae-Yean

    2015-10-01

    It has been suggested that AUXIN BINDING PROTEIN 1 (ABP1) functions as an apoplastic auxin receptor, and is known to be involved in the post-transcriptional process, and largely independent of the already well-known SKP-cullin-F-box-transport inhibitor response (TIR1) /auxin signaling F-box (AFB) (SCF(TIR1/AFB)) pathway. In the past 10 years, several key components downstream of ABP1 have been reported. After perceiving the auxin signal, ABP1 interacts, directly or indirectly, with plasma membrane (PM)-localized transmembrane proteins, transmembrane kinase (TMK) or SPIKE1 (SPK1), or other unidentified proteins, which transfer the signal into the cell to the Rho of plants (ROP). ROPs interact with their effectors, such as the ROP interactive CRIB motif-containing protein (RIC), to regulate the endocytosis/exocytosis of the auxin efflux carrier PIN-FORMED (PIN) proteins to mediate polar auxin transport across the PM. Additionally, ABP1 is a negative regulator of the traditional SCF(TIR1/AFB) auxin signaling pathway. However, Gao et al. (2015) very recently reported that ABP1 is not a key component in auxin signaling, and the famous abp1-1 and abp1-5 mutant Arabidopsis lines are being called into question because of possible additional mutantion sites, making it necessary to reevaluate ABP1. In this review, we will provide a brief overview of the history of ABP1 research.

  16. Heterotrimeric G protein signalling in the plant kingdom

    Science.gov (United States)

    Urano, Daisuke; Chen, Jin-Gui; Botella, José Ramón; Jones, Alan M.

    2013-01-01

    In animals, heterotrimeric G proteins, comprising α-, β-and γ-subunits, perceive extracellular stimuli through cell surface receptors, and transmit signals to ion channels, enzymes and other effector proteins to affect numerous cellular behaviours. In plants, G proteins have structural similarities to the corresponding molecules in animals but transmit signals by atypical mechanisms and effector proteins to control growth, cell proliferation, defence, stomate movements, channel regulation, sugar sensing and some hormonal responses. In this review, we summarize the current knowledge on the molecular regulation of plant G proteins, their effectors and the physiological functions studied mainly in two model organisms: Arabidopsis thaliana and rice (Oryza sativa). We also look at recent progress on structural analyses, systems biology and evolutionary studies. PMID:23536550

  17. Heterotrimeric G protein signalling in the plant kingdom.

    Science.gov (United States)

    Urano, Daisuke; Chen, Jin-Gui; Botella, José Ramón; Jones, Alan M

    2013-03-27

    In animals, heterotrimeric G proteins, comprising α-, β-and γ-subunits, perceive extracellular stimuli through cell surface receptors, and transmit signals to ion channels, enzymes and other effector proteins to affect numerous cellular behaviours. In plants, G proteins have structural similarities to the corresponding molecules in animals but transmit signals by atypical mechanisms and effector proteins to control growth, cell proliferation, defence, stomate movements, channel regulation, sugar sensing and some hormonal responses. In this review, we summarize the current knowledge on the molecular regulation of plant G proteins, their effectors and the physiological functions studied mainly in two model organisms: Arabidopsis thaliana and rice (Oryza sativa). We also look at recent progress on structural analyses, systems biology and evolutionary studies.

  18. PathFinder: mining signal transduction pathway segments from protein-protein interaction networks

    Directory of Open Access Journals (Sweden)

    Yang Jiong

    2007-09-01

    Full Text Available Abstract Background A Signal transduction pathway is the chain of processes by which a cell converts an extracellular signal into a response. In most unicellular organisms, the number of signal transduction pathways influences the number of ways the cell can react and respond to the environment. Discovering signal transduction pathways is an arduous problem, even with the use of systematic genomic, proteomic and metabolomic technologies. These techniques lead to an enormous amount of data and how to interpret and process this data becomes a challenging computational problem. Results In this study we present a new framework for identifying signaling pathways in protein-protein interaction networks. Our goal is to find biologically significant pathway segments in a given interaction network. Currently, protein-protein interaction data has excessive amount of noise, e.g., false positive and false negative interactions. First, we eliminate false positives in the protein-protein interaction network by integrating the network with microarray expression profiles, protein subcellular localization and sequence information. In addition, protein families are used to repair false negative interactions. Then the characteristics of known signal transduction pathways and their functional annotations are extracted in the form of association rules. Conclusion Given a pair of starting and ending proteins, our methodology returns candidate pathway segments between these two proteins with possible missing links (recovered false negatives. In our study, S. cerevisiae (yeast data is used to demonstrate the effectiveness of our method.

  19. The multi-step phosphorelay mechanism of unorthodox two-component systems in E. coli realizes ultrasensitivity to stimuli while maintaining robustness to noises.

    Science.gov (United States)

    Kim, Jeong-Rae; Cho, Kwang-Hyun

    2006-12-01

    E. coli has two-component systems composed of histidine kinase proteins and response regulator proteins. For a given extracellular stimulus, a histidine kinase senses the stimulus, autophosphorylates and then passes the phosphates to the cognate response regulators. The histidine kinase in an orthodox two-component system has only one histidine domain where the autophosphorylation occurs, but a histidine kinase in some unusual two-component systems (unorthodox two-component systems) has two histidine domains and one aspartate domain. So, the unorthodox two-component systems have more complex phosphorelay mechanisms than orthodox two-component systems. In general, the two-component systems are required to promptly respond to external stimuli for survival of E. coli. In this respect, the complex multi-step phosphorelay mechanism seems to be disadvantageous, but there are several unorthodox two-component systems in E. coli. In this paper, we investigate the reason why such unorthodox two-component systems are present in E. coli. For this purpose, we have developed simplified mathematical models of both orthodox and unorthodox two-component systems and analyzed their dynamical characteristics through extensive computer simulations. We have finally revealed that the unorthodox two-component systems realize ultrasensitive responses to external stimuli and also more robust responses to noises than the orthodox two-component systems.

  20. The CpxRA two-component system is essential for Citrobacter rodentium virulence.

    Science.gov (United States)

    Thomassin, Jenny-Lee; Giannakopoulou, Natalia; Zhu, Lei; Gross, Jeremy; Salmon, Kristiana; Leclerc, Jean-Mathieu; Daigle, France; Le Moual, Hervé; Gruenheid, Samantha

    2015-05-01

    Citrobacter rodentium is a murine intestinal pathogen used as a model for the foodborne human pathogens enterohemorrhagic Escherichia coli and enteropathogenic E. coli. During infection, these pathogens use two-component signal transduction systems to detect and adapt to changing environmental conditions. In E. coli, the CpxRA two-component signal transduction system responds to envelope stress by modulating the expression of a myriad of genes. Quantitative real-time PCR showed that cpxRA was expressed in the colon of C57BL/6J mice infected with C. rodentium. To determine whether CpxRA plays a role during C. rodentium infection, a cpxRA deletion strain was generated and found to have a colonization defect during infection. This defect was independent of an altered growth rate or a defective type III secretion system, and single-copy chromosomal complementation of cpxRA restored virulence. The C. rodentium strains were then tested in C3H/HeJ mice, a lethal intestinal infection model. Mice infected with the ΔcpxRA strain survived infection, whereas mice infected with the wild-type or complemented strains succumbed to infection. Furthermore, we found that the cpxRA expression level was higher during early infection than at a later time point. Taken together, these data demonstrate that the CpxRA two-component signal transduction system is essential for the in vivo virulence of C. rodentium. In addition, these data suggest that fine-tuned cpxRA expression is important for infection. This is the first study that identifies a C. rodentium two-component transduction system required for pathogenesis. This study further indicates that CpxRA is an interesting target for therapeutics against enteric pathogens.

  1. Two-Component Multi-Parameter Time-Frequency Electromagnetics

    Institute of Scientific and Technical Information of China (English)

    HuangZhou; DongWeibin; HeTiezhi

    2003-01-01

    The two-component multi-parameter time-frequency electromagnetic method, used for the development of oilfields,makes use of both the traditional individual conductivity parameters of oil-producing layers and the dispersion information of the conductivity, i.e., the induced polarization parameter. The frequency-domain dispersion data is used to delineate the contacts between oil and water and the time domain dBz/dt component is used to estimate the depths to the un-known reservoirs so as to offer significant data in many aspects for oil exploration and detection.

  2. A polaritonic two-component Bose-Hubbard model

    Energy Technology Data Exchange (ETDEWEB)

    Hartmann, M J; Brandao, F G S L; Plenio, M B [Institute for Mathematical Sciences, Imperial College London, 53 Exhibition Road, SW7 2PE (United Kingdom)], E-mail: m.hartmann@imperial.ac.uk

    2008-03-15

    We demonstrate that polaritons in an array of interacting micro-cavities with strong atom-photon coupling can form a two-component Bose-Hubbard model in which both polariton species are protected against spontaneous emission as their atomic part is stored in two ground states of the atoms. The parameters of the effective model can be tuned via the driving strength of external lasers and include attractive and repulsive polariton interactions. We also describe a method to measure the number statistics in one cavity for each polariton species independently.

  3. Two component micro injection moulding for moulded interconnect devices

    DEFF Research Database (Denmark)

    Islam, Aminul

    2008-01-01

    Moulded interconnect devices (MIDs) contain huge possibilities for many applications in micro electro-mechanical-systems because of their capability of reducing the number of components, process steps and finally in miniaturization of the product. Among the available MID process chains, two...... and a reasonable adhesion between them. • Selective metallization of the two component plastic part (coating one polymer with metal and leaving the other one uncoated) To overcome these two main issues in MID fabrication for micro applications, the current Ph.D. project explores the technical difficulties...

  4. Two-component Fermi gas in a Harmonic Trap

    CERN Document Server

    Yi, X X; Cui, H T; Zhang, C M

    2002-01-01

    We consider a mixture of two-component Fermi gases at low temperature. The density profile of this degenerate Fermi gas is calculated under the semiclassical approximation. The results show that the fermion-fermion interactions make a large correction to the density profile at low temperature. The phase separation of such a mixture is also discussed for both attractive and repulsive interatomic interactions, and the numerical calculations demonstrate the exist of a stable temperature region $T_{c1}

  5. Interaction potentials and thermodynamic properties of two component semiclassical plasma

    Energy Technology Data Exchange (ETDEWEB)

    Ramazanov, T. S.; Moldabekov, Zh. A.; Ismagambetova, T. N. [Al-Farabi Kazakh National University, IETP, 71 al-Farabi Av., Almaty 050040 (Kazakhstan); Gabdullin, M. T. [Al-Farabi Kazakh National University, NNLOT, 71 al-Farabi Av., Almaty 050040 (Kazakhstan)

    2014-01-15

    In this paper, the effective interaction potential in two component semiclassical plasma, taking into account the long-range screening and the quantum-mechanical diffraction effects at short distances, is obtained on the basis of dielectric response function method. The structural properties of the semiclassical plasma are considered. The thermodynamic characteristics (the internal energy and the equation of state) are calculated using two methods: the method of effective potentials and the method of micropotentials with screening effect taken into account by the Ornstein-Zernike equation in the HNC approximation.

  6. Itinerant ferromagnetism in a polarized two-component Fermi gas.

    Science.gov (United States)

    Massignan, Pietro; Yu, Zhenhua; Bruun, Georg M

    2013-06-07

    We analyze when a repulsively interacting two-component Fermi gas becomes thermodynamically unstable against phase separation. We focus on the strongly polarized limit, where the free energy of the homogeneous mixture can be calculated accurately in terms of well-defined quasiparticles, the repulsive polarons. Phase diagrams as a function of polarization, temperature, mass imbalance, and repulsive polaron energy, as well as scattering length and range parameter, are provided. We show that the lifetime of the repulsive polaron increases significantly with the interaction range and the mass of the minority atoms, raising the prospects of detecting the transition to the elusive itinerant ferromagnetic state with ultracold atoms.

  7. Two component micro injection molding for MID fabrication

    DEFF Research Database (Denmark)

    Islam, Mohammad Aminul; Hansen, Hans Nørgaard; Tang, Peter Torben

    2009-01-01

    Molded Interconnect Devices (MIDs) are plastic substrates with electrical infrastructure. The fabrication of MIDs is usually based on injection molding and different process chains may be identified from this starting point. The use of MIDs has been driven primarily by the automotive sector......, but recently the medical sector seems more and more interested. In particular the possibility of miniaturization of 3D components with electrical infrastructure is attractive. The paper describes possible manufacturing routes and challenges of miniaturized MIDs based on two component micro injection molding...

  8. Heterotrimeric G protein signaling in polycystic kidney disease.

    Science.gov (United States)

    Hama, Taketsugu; Park, Frank

    2016-07-01

    Autosomal dominant polycystic kidney disease (ADPKD) is a signalopathy of renal tubular epithelial cells caused by naturally occurring mutations in two distinct genes, polycystic kidney disease 1 (PKD1) and 2 (PKD2). Genetic variants in PKD1, which encodes the polycystin-1 (PC-1) protein, remain the predominant factor associated with the pathogenesis of nearly two-thirds of all patients diagnosed with PKD. Although the relationship between defective PC-1 with renal cystic disease initiation and progression remains to be fully elucidated, there are numerous clinical studies that have focused upon the control of effector systems involving heterotrimeric G protein regulation. A major regulator in the activation state of heterotrimeric G proteins are G protein-coupled receptors (GPCRs), which are defined by their seven transmembrane-spanning regions. PC-1 has been considered to function as an unconventional GPCR, but the mechanisms by which PC-1 controls signal processing, magnitude, or trafficking through heterotrimeric G proteins remains to be fully known. The diversity of heterotrimeric G protein signaling in PKD is further complicated by the presence of non-GPCR proteins in the membrane or cytoplasm that also modulate the functional state of heterotrimeric G proteins within the cell. Moreover, PC-1 abnormalities promote changes in hormonal systems that ultimately interact with distinct GPCRs in the kidney to potentially amplify or antagonize signaling output from PC-1. This review will focus upon the canonical and noncanonical signaling pathways that have been described in PKD with specific emphasis on which heterotrimeric G proteins are involved in the pathological reorganization of the tubular epithelial cell architecture to exacerbate renal cystogenic pathways.

  9. Protein and signaling networks in vertebrate photoreceptor cells

    Directory of Open Access Journals (Sweden)

    Karl-Wilhelm eKoch

    2015-11-01

    Full Text Available Vertebrate photoreceptor cells are exquisite light detectors operating under very dim and bright illumination. The photoexcitation and adaptation machinery in photoreceptor cells consists of protein complexes that can form highly ordered supramolecular structures and control the homeostasis and mutual dependence of the secondary messengers cGMP and Ca2+. The visual pigment in rod photoreceptors, the G protein-coupled receptor rhodopsin is organized in tracks of dimers thereby providing a signaling platform for the dynamic scaffolding of the G protein transducin. Illuminated rhodopsin is turned off by phosphorylation catalyzed by rhodopsin kinase GRK1 under control of Ca2+-recoverin. The GRK1 protein complex partly assembles in lipid raft structures, where shutting off rhodopsin seems to be more effective. Re-synthesis of cGMP is another crucial step in the recovery of the photoresponse after illumination. It is catalyzed by membrane bound sensory guanylate cyclases and is regulated by specific neuronal Ca2+-sensor proteins called GCAPs. At least one guanylate cyclase (ROS-GC1 was shown to be part of a multiprotein complex having strong interactions with the cytoskeleton and being controlled in a multimodal Ca2+-dependent fashion. The final target of the cGMP signaling cascade is a cyclic nucleotide-gated channel that is a hetero-oligomeric protein located in the plasma membrane and interacting with accessory proteins in highly organized microdomains. We summarize results and interpretations of findings related to the inhomogeneous organization of signaling units in photoreceptor outer segments.

  10. Role of signal peptides in targeting of proteins in cyanobacteria.

    OpenAIRE

    Mackle, M M; Zilinskas, B A

    1994-01-01

    Proteins of cyanobacteria may be transported across one of two membrane systems: the typical eubacterial cell envelope (consisting of an inner membrane, periplasmic space, and an outer membrane) and the photosynthetic thylakoids. To investigate the role of signal peptides in targeting in cyanobacteria, Synechococcus sp. strain PCC 7942 was transformed with vectors carrying the chloramphenicol acetyltransferase reporter gene fused to coding sequences for one of four different signal peptides. ...

  11. PACRG, a protein linked to ciliary motility, mediates cellular signaling.

    OpenAIRE

    Loucks, Catrina M.; Bialas, Nathan J.; Dekkers, Martijn; Walker, Denise S.; Grundy, Laura J.; Li, Chunmei; Inglis, P. Nick; Kida, Katarzyna; Schafer, William R; Blacque, Oliver E; Jansen, Gert; Michel R Leroux

    2016-01-01

    Cilia are microtubule-based organelles that project from nearly all mammalian cell types. Motile cilia generate fluid flow, whereas nonmotile (primary) cilia are required for sensory physiology and modulate various signal transduction pathways. Here we investigate the nonmotile ciliary signaling roles of parkin coregulated gene (PACRG), a protein linked to ciliary motility. PACRG is associated with the protofilament ribbon, a structure believed to dictate the regular arrangement of motility-a...

  12. PACRG, a protein linked to ciliary motility, mediates cellular signaling

    OpenAIRE

    Loucks, Catrina M.; Bialas, Nathan J.; Dekkers, Martijn P. J.; Walker, Denise S.; Grundy, Laura J.; Li, Chunmei; Inglis, P. Nick; Kida, Katarzyna; Schafer, William R; Blacque, Oliver E; Jansen, Gert; Michel R Leroux

    2016-01-01

    Cilia are microtubule-based organelles that project from nearly all mammalian cell types. Motile cilia generate fluid flow, whereas nonmotile (primary) cilia are required for sensory physiology and modulate various signal transduction pathways. Here we investigate the nonmotile ciliary signaling roles of parkin coregulated gene (PACRG), a protein linked to ciliary motility. PACRG is associated with the protofilament ribbon, a structure believed to dictate the regular arrangement of motility-a...

  13. Graphene Oxide: A One- versus Two-Component Material.

    Science.gov (United States)

    Naumov, Anton; Grote, Fabian; Overgaard, Marc; Roth, Alexandra; Halbig, Christian E; Nørgaard, Kasper; Guldi, Dirk M; Eigler, Siegfried

    2016-09-14

    The structure of graphene oxide (GO) is a matter of discussion. While established GO models are based on functional groups attached to the carbon framework, another frequently used model claims that GO consists of two components, a slightly oxidized graphene core and highly oxidized molecular species, oxidative debris (OD), adsorbed on it. Those adsorbents are claimed to be the origin for optical properties of GO. Here, we examine this model by preparing GO with a low degree of functionalization, combining it with OD and studying the optical properties of both components and their combination in an artificial two-component system. The analyses of absorption and emission spectra as well as lifetime measurements reveal that properties of the combined system are distinctly different from those of GO. That confirms structural models of GO as a separate oxygenated hexagonal carbon framework with optical properties governed by its internal structure rather than the presence of OD. Understanding the structure of GO allows further reliable interpretation of its optical and electronic properties and enables controlled processing of GO.

  14. Science Signaling Podcast for 12 July 2016: Adaptor proteins limit signaling.

    Science.gov (United States)

    Wiley, H Steven; VanHook, Annalisa M

    2016-07-12

    This Podcast features an interview with Steven Wiley, senior author of a Research Article that appears in the 12 July 2016 issue of Science Signaling, about how the abundance of adaptor proteins and feedback regulators affect the flow of information downstream of the epidermal growth factor receptor (EGFR). Information flows through a signaling pathway by sequential interactions between core components of the pathway, many of which have enzymatic activity. Adaptor proteins do not directly participate in relaying the signal and do not have enzymatic activity, but are important for signaling because they facilitate interactions between the core components. Using quantitative methods, Shi et al demonstrated that core components of the EGFR pathway were highly abundant in both normal cells and cancer cells. However, adaptor proteins were present in much lower abundance in both cell types, indicating that it is the abundance of these proteins that limit signaling downstream of EGFR. The authors also found that differences in EGFR signaling between different cell types likely resulted from the variable abundance of feedback regulators.Listen to Podcast. Copyright © 2016, American Association for the Advancement of Science.

  15. Stable evolutionary signal in a Yeast protein interaction network

    Directory of Open Access Journals (Sweden)

    Ferdig Michael T

    2006-01-01

    Full Text Available Abstract Background The recently emerged protein interaction network paradigm can provide novel and important insights into the innerworkings of a cell. Yet, the heavy burden of both false positive and false negative protein-protein interaction data casts doubt on the broader usefulness of these interaction sets. Approaches focusing on one-protein-at-a-time have been powerfully employed to demonstrate the high degree of conservation of proteins participating in numerous interactions; here, we expand his 'node' focused paradigm to investigate the relative persistence of 'link' based evolutionary signals in a protein interaction network of S. cerevisiae and point out the value of this relatively untapped source of information. Results The trend for highly connected proteins to be preferably conserved in evolution is stable, even in the context of tremendous noise in the underlying protein interactions as well as in the assignment of orthology among five higher eukaryotes. We find that local clustering around interactions correlates with preferred evolutionary conservation of the participating proteins; furthermore the correlation between high local clustering and evolutionary conservation is accompanied by a stable elevated degree of coexpression of the interacting proteins. We use this conserved interaction data, combined with P. falciparum /Yeast orthologs, as proof-of-principle that high-order network topology can be used comparatively to deduce local network structure in non-model organisms. Conclusion High local clustering is a criterion for the reliability of an interaction and coincides with preferred evolutionary conservation and significant coexpression. These strong and stable correlations indicate that evolutionary units go beyond a single protein to include the interactions among them. In particular, the stability of these signals in the face of extreme noise suggests that empirical protein interaction data can be integrated with

  16. G protein signaling in the parasite Entamoeba histolytica

    Science.gov (United States)

    Bosch, Dustin E; Siderovski, David P

    2013-01-01

    The parasite Entamoeba histolytica causes amebic colitis and systemic amebiasis. Among the known amebic factors contributing to pathogenesis are signaling pathways involving heterotrimeric and Ras superfamily G proteins. Here, we review the current knowledge of the roles of heterotrimeric G protein subunits, Ras, Rho and Rab GTPase families in E. histolytica pathogenesis, as well as of their downstream signaling effectors and nucleotide cycle regulators. Heterotrimeric G protein signaling likely modulates amebic motility and attachment to and killing of host cells, in part through activation of an RGS-RhoGEF (regulator of G protein signaling–Rho guanine nucleotide exchange factor) effector. Rho family GTPases, as well as RhoGEFs and Rho effectors (formins and p21-activated kinases) regulate the dynamic actin cytoskeleton of E. histolytica and associated pathogenesis-related cellular processes, such as migration, invasion, phagocytosis and evasion of the host immune response by surface receptor capping. A remarkably large family of 91 Rab GTPases has multiple roles in a complex amebic vesicular trafficking system required for phagocytosis and pinocytosis and secretion of known virulence factors, such as amebapores and cysteine proteases. Although much remains to be discovered, recent studies of G protein signaling in E. histolytica have enhanced our understanding of parasitic pathogenesis and have also highlighted possible targets for pharmacological manipulation. PMID:23519208

  17. Receptor component protein (RCP): a member of a multi-protein complex required for G-protein-coupled signal transduction.

    Science.gov (United States)

    Prado, M A; Evans-Bain, B; Dickerson, I M

    2002-08-01

    The calcitonin-gene-related peptide (CGRP) receptor component protein (RCP) is a 148-amino-acid intracellular protein that is required for G-protein-coupled signal transduction at receptors for the neuropeptide CGRP. RCP works in conjunction with two other proteins to constitute a functional CGRP receptor: calcitonin-receptor-like receptor (CRLR) and receptor-activity-modifying protein 1 (RAMP1). CRLR has the stereotypical seven-transmembrane topology of a G-protein-coupled receptor; it requires RAMP1 for trafficking to the cell surface and for ligand specificity, and requires RCP for coupling to the cellular signal transduction pathway. We have made cell lines that expressed an antisense construct of RCP and determined that CGRP-mediated signal transduction was reduced, while CGRP binding was unaffected. Furthermore, signalling at two other endogenous G-protein-coupled receptors was unaffected, suggesting that RCP was specific for a limited subset of receptors.

  18. Dietary proteins and food-related reward signals

    Directory of Open Access Journals (Sweden)

    Katri Peuhkuri

    2011-06-01

    Full Text Available Proteins play a crucial role in almost all biological processes. Dietary proteins are generally considered as energy yielding nutrients and as a source of amino acids for various purposes. In addition, they may have a role in food-related reward signals. The purpose of this review was to give an overview of the role of dietary proteins in food-related reward and possible mechanisms behind such effects. Dietary proteins may elicit food-related reward by several different postprandial mechanisms, including neural and humoral signals from the gastrointestinal tract to the brain. In order to exert rewarding effects, protein have to be absorbed from the intestine and reach the target cells in sufficient concentrations, or act via receptors ad cell signalling in the gut without absorption. Complex interactions between different possible mechanisms make it very difficult to gain a clear view on the role and intesity of each mechanism. It is concluded that, in principle, dietary proteins may have a role in food-related reward. However, the evidence is based mostly on experiments with animal models and one should be careful in drawing conclusions of clinical relevance.

  19. Exact two-component relativistic energy band theory and application

    Energy Technology Data Exchange (ETDEWEB)

    Zhao, Rundong; Zhang, Yong; Xiao, Yunlong; Liu, Wenjian, E-mail: liuwj@pku.edu.cn [Beijing National Laboratory for Molecular Sciences, Institute of Theoretical and Computational Chemistry, State Key Laboratory of Rare Earth Materials Chemistry and Applications, College of Chemistry and Molecular Engineering, and Center for Computational Science and Engineering, Peking University, Beijing 100871 (China)

    2016-01-28

    An exact two-component (X2C) relativistic density functional theory in terms of atom-centered basis functions is proposed for relativistic calculations of band structures and structural properties of periodic systems containing heavy elements. Due to finite radial extensions of the local basis functions, the periodic calculation is very much the same as a molecular calculation, except only for an Ewald summation for the Coulomb potential of fluctuating periodic monopoles. For comparison, the nonrelativistic and spin-free X2C counterparts are also implemented in parallel. As a first and pilot application, the band gaps, lattice constants, cohesive energies, and bulk moduli of AgX (X = Cl, Br, I) are calculated to compare with other theoretical results.

  20. Two-component jet simulations: Combining analytical and numerical approaches

    CERN Document Server

    Matsakos, T; Trussoni, E; Tsinganos, K; Vlahakis, N; Sauty, C; Mignone, A

    2009-01-01

    Recent observations as well as theoretical studies of YSO jets suggest the presence of two steady components: a disk wind type outflow needed to explain the observed high mass loss rates and a stellar wind type outflow probably accounting for the observed stellar spin down. In this framework, we construct numerical two-component jet models by properly mixing an analytical disk wind solution with a complementary analytically derived stellar outflow. Their combination is controlled by both spatial and temporal parameters, in order to address different physical conditions and time variable features. We study the temporal evolution and the interaction of the two jet components on both small and large scales. The simulations reach steady state configurations close to the initial solutions. Although time variability is not found to considerably affect the dynamics, flow fluctuations generate condensations, whose large scale structures have a strong resemblance to observed YSO jet knots.

  1. Exact two-component relativistic energy band theory and application.

    Science.gov (United States)

    Zhao, Rundong; Zhang, Yong; Xiao, Yunlong; Liu, Wenjian

    2016-01-28

    An exact two-component (X2C) relativistic density functional theory in terms of atom-centered basis functions is proposed for relativistic calculations of band structures and structural properties of periodic systems containing heavy elements. Due to finite radial extensions of the local basis functions, the periodic calculation is very much the same as a molecular calculation, except only for an Ewald summation for the Coulomb potential of fluctuating periodic monopoles. For comparison, the nonrelativistic and spin-free X2C counterparts are also implemented in parallel. As a first and pilot application, the band gaps, lattice constants, cohesive energies, and bulk moduli of AgX (X = Cl, Br, I) are calculated to compare with other theoretical results.

  2. Recent advances in description of few two-component fermions

    CERN Document Server

    Kartavtsev, O I

    2012-01-01

    Overview of the recent advances in description of the few two-component fermions is presented. The zero-range interaction limit is generally considered to discuss the principal aspects of the few-body dynamics. Significant attention is paid to detailed description of two identical fermions of mass $m$ and a distinct particle of mass $m_1$; two universal $L^P = 1^-$ bound states arise for mass ratio $m/m_1$ increasing up to the critical value $\\mu_c \\approx 13.607$, beyond which the Efimov effect takes place. The topics considered include rigorous treatment of the few-fermion problem in the zero-range interaction limit, low-dimensional results, the four-body energy spectrum, crossover of the energy spectra for $m/m_1$ near the critical value $\\mu_c $, and properties of potential-dependent states. At last, enlisted are the problems, whose solution is in due course.

  3. Bond strength of two component injection moulded MID

    DEFF Research Database (Denmark)

    Islam, Mohammad Aminul; Hansen, Hans Nørgaard; Tang, Peter Torben

    2006-01-01

    Most products of the future will require industrially adapted, cost effective production processes and on this issue two-component (2K) injection moulding is a potential candidate for MID manufacturing. MID based on 2k injection moulded plastic part with selectively metallised circuit tracks allows...... the integration of electrical and mechanical functionalities in a real 3D structure. If 2k injection moulding is applied with two polymers, of which one is plateable and the other is not, it will be possible to make 3D electrical structures directly on the component. To be applicable in the real engineering field...... the two different plastic materials in the MID structure require good bonding between them. This paper finds suitable combinations of materials for MIDs from both bond strength and metallisation view-point. Plastic parts were made by two-shot injection moulding and the effects of some important process...

  4. Efficient two-component relativistic method for large systems

    Energy Technology Data Exchange (ETDEWEB)

    Nakai, Hiromi [Department of Chemitsry and Biochemistry, School of Advanced Science and Engineering, Waseda University, Tokyo 169-8555 (Japan); Research Institute for Science and Engineering, Waseda University, Tokyo 169-8555 (Japan); CREST, Japan Science and Technology Agency, 4-1-8 Honcho, Kawaguchi, Saitama 332-0012 (Japan); Elements Strategy Initiative for Catalysts and Batteries (ESICB), Kyoto University, Katsura, Kyoto 615-8520 (Japan)

    2015-12-31

    This paper reviews a series of theoretical studies to develop efficient two-component (2c) relativistic method for large systems by the author’s group. The basic theory is the infinite-order Douglas-Kroll-Hess (IODKH) method for many-electron Dirac-Coulomb Hamiltonian. The local unitary transformation (LUT) scheme can effectively produce the 2c relativistic Hamiltonian, and the divide-and-conquer (DC) method can achieve linear-scaling of Hartree-Fock and electron correlation methods. The frozen core potential (FCP) theoretically connects model potential calculations with the all-electron ones. The accompanying coordinate expansion with a transfer recurrence relation (ACE-TRR) scheme accelerates the computations of electron repulsion integrals with high angular momenta and long contractions.

  5. No electrostatic supersolitons in two-component plasmas

    Energy Technology Data Exchange (ETDEWEB)

    Verheest, Frank, E-mail: frank.verheest@ugent.be [Sterrenkundig Observatorium, Universiteit Gent, Krijgslaan 281, B–9000 Gent (Belgium); School of Chemistry and Physics, University of KwaZulu-Natal, Durban 4000 (South Africa); Lakhina, Gurbax S., E-mail: lakhina@iigm.iigs.res.in [Indian Institute of Geomagnetism, New Panvel (W), Navi Mumbai (India); Hellberg, Manfred A., E-mail: hellberg@ukzn.ac.za [School of Chemistry and Physics, University of KwaZulu-Natal, Durban 4000 (South Africa)

    2014-06-15

    The concept of acoustic supersolitons was introduced for a very specific plasma with five constituents, and discussed only for a single set of plasma parameters. Supersolitons are characterized by having subsidiary extrema on the sides of a typical bipolar electric field signature, or by association with a root beyond double layers in the fully nonlinear Sagdeev pseudopotential description. It was subsequently found that supersolitons could exist in several plasma models having three constituent species, rather than four or five. In the present paper, it is proved that standard two-component plasma models cannot generate supersolitons, by recalling and extending results already in the literature, and by establishing the necessary properties of a more recent model.

  6. Budding Transition of Asymmetric Two-component Lipid Domains

    CERN Document Server

    Wolff, Jean; Andelman, David

    2016-01-01

    We propose a model that accounts for the budding transition of asymmetric two-component lipid domains, where the two monolayers (leaflets) have different average compositions controlled by independent chemical potentials. Assuming a coupling between the local curvature and local lipid composition in each of the leaflets, we discuss the morphology and thermodynamic behavior of asymmetric lipid domains. The membrane free-energy contains three contributions: the bending energy, the line tension, and a Landau free-energy for a lateral phase separation. Within a mean-field treatment, we obtain various phase diagrams containing fully budded, dimpled, and flat states as a function of the two leaflet compositions. The global phase behavior is analyzed, and depending on system parameters, the phase diagrams include one-phase, two-phase and three-phase regions. In particular, we predict various phase coexistence regions between different morphologies of domains, which may be observed in multi-component membranes or ves...

  7. How insects overcome two-component plant chemical defence

    DEFF Research Database (Denmark)

    Pentzold, Stefan; Zagrobelny, Mika; Rook, Frederik;

    2014-01-01

    Insect herbivory is often restricted by glucosylated plant chemical defence compounds that are activated by plant β-glucosidases to release toxic aglucones upon plant tissue damage. Such two-component plant defences are widespread in the plant kingdom and examples of these classes of compounds...... are alkaloid, benzoxazinoid, cyanogenic and iridoid glucosides as well as glucosinolates and salicinoids. Conversely, many insects have evolved a diversity of counteradaptations to overcome this type of constitutive chemical defence. Here we discuss that such counter-adaptations occur at different time points......-component chemical defence. These adaptations include host plant choice, non-disruptive feeding guilds and various physiological adaptations as well as metabolic enzymatic strategies of the insect’s digestive system. Furthermore, insect adaptations often act in combination, may exist in both generalists...

  8. Parallel TREE code for two-component ultracold plasma analysis

    Science.gov (United States)

    Jeon, Byoungseon; Kress, Joel D.; Collins, Lee A.; Grønbech-Jensen, Niels

    2008-02-01

    The TREE method has been widely used for long-range interaction N-body problems. We have developed a parallel TREE code for two-component classical plasmas with open boundary conditions and highly non-uniform charge distributions. The program efficiently handles millions of particles evolved over long relaxation times requiring millions of time steps. Appropriate domain decomposition and dynamic data management were employed, and large-scale parallel processing was achieved using an intermediate level of granularity of domain decomposition and ghost TREE communication. Even though the computational load is not fully distributed in fine grains, high parallel efficiency was achieved for ultracold plasma systems of charged particles. As an application, we performed simulations of an ultracold neutral plasma with a half million particles and a half million time steps. For the long temporal trajectories of relaxation between heavy ions and light electrons, large configurations of ultracold plasmas can now be investigated, which was not possible in past studies.

  9. Prediction of N-terminal protein sorting signals

    DEFF Research Database (Denmark)

    Claros, Manuel G.; Brunak, Søren; von Heijne, Gunnar

    1997-01-01

    Recently, neural networks have been applied to a widening range of problems in molecular biology. An area particularly suited to neural-network methods is the identification of protein sorting signals and the prediction of their cleavage sites, as these functional units are encoded by local, linear...

  10. The protein phosphatase 7 regulates phytochrome signaling in Arabidopsis.

    Directory of Open Access Journals (Sweden)

    Thierry Genoud

    Full Text Available The psi2 mutant of Arabidopsis displays amplification of the responses controlled by the red/far red light photoreceptors phytochrome A (phyA and phytochrome B (phyB but no apparent defect in blue light perception. We found that loss-of-function alleles of the protein phosphatase 7 (AtPP7 are responsible for the light hypersensitivity in psi2 demonstrating that AtPP7 controls the levels of phytochrome signaling. Plants expressing reduced levels of AtPP7 mRNA display reduced blue-light induced cryptochrome signaling but no noticeable deficiency in phytochrome signaling. Our genetic analysis suggests that phytochrome signaling is enhanced in the AtPP7 loss of function alleles, including in blue light, which masks the reduced cryptochrome signaling. AtPP7 has been found to interact both in yeast and in planta assays with nucleotide-diphosphate kinase 2 (NDPK2, a positive regulator of phytochrome signals. Analysis of ndpk2-psi2 double mutants suggests that NDPK2 plays a critical role in the AtPP7 regulation of the phytochrome pathway and identifies NDPK2 as an upstream element involved in the modulation of the salicylic acid (SA-dependent defense pathway by light. Thus, cryptochrome- and phytochrome-specific light signals synchronously control their relative contribution to the regulation of plant development. Interestingly, PP7 and NDPK are also components of animal light signaling systems.

  11. Analysis of signaling networks distributed over intracellular compartments based on protein-protein interactions

    OpenAIRE

    2014-01-01

    BackgroundBiological processes are usually distributed over various intracellular compartments. Proteins from diverse cellular compartments are often involved in similar signaling networks. However, the difference in the reaction rates between similar proteins among different compartments is usually quite high. We suggest that the estimation of frequency of intracompartmental as well as intercompartmental protein-protein interactions is an appropriate approach to predict the efficiency of a p...

  12. The signaling helix: a common functional theme in diverse signaling proteins

    Directory of Open Access Journals (Sweden)

    Aravind L

    2006-09-01

    Full Text Available Abstract Background The mechanism by which the signals are transmitted between receptor and effector domains in multi-domain signaling proteins is poorly understood. Results Using sensitive sequence analysis methods we identify a conserved helical segment of around 40 residues in a wide range of signaling proteins, including numerous sensor histidine kinases such as Sln1p, and receptor guanylyl cyclases such as the atrial natriuretic peptide receptor and nitric oxide receptors. We term this helical segment the signaling (S-helix and present evidence that it forms a novel parallel coiled-coil element, distinct from previously known helical segments in signaling proteins, such as the Dimerization-Histidine phosphotransfer module of histidine kinases, the intra-cellular domains of the chemotaxis receptors, inter-GAF domain helical linkers and the α-helical HAMP module. Analysis of domain architectures allowed us to reconstruct the domain-neighborhood graph for the S-helix, which showed that the S-helix almost always occurs between two signaling domains. Several striking patterns in the domain neighborhood of the S-helix also became evident from the graph. It most often separates diverse N-terminal sensory domains from various C-terminal catalytic signaling domains such as histidine kinases, cNMP cyclase, PP2C phosphatases, NtrC-like AAA+ ATPases and diguanylate cyclases. It might also occur between two sensory domains such as PAS domains and occasionally between a DNA-binding HTH domain and a sensory domain. The sequence conservation pattern of the S-helix revealed the presence of a unique constellation of polar residues in the dimer-interface positions within the central heptad of the coiled-coil formed by the S-helix. Conclusion Combining these observations with previously reported mutagenesis studies on different S-helix-containing proteins we suggest that it functions as a switch that prevents constitutive activation of linked downstream

  13. The Transmembrane Adaptor Protein SIT Inhibits TCR-Mediated Signaling

    Science.gov (United States)

    Arndt, Börge; Krieger, Tina; Kalinski, Thomas; Thielitz, Anja; Reinhold, Dirk; Roessner, Albert; Schraven, Burkhart; Simeoni, Luca

    2011-01-01

    Transmembrane adaptor proteins (TRAPs) organize signaling complexes at the plasma membrane, and thus function as critical linkers and integrators of signaling cascades downstream of antigen receptors. We have previously shown that the transmembrane adaptor protein SIT regulates the threshold for thymocyte selection. Moreover, T cells from SIT-deficient mice are hyperresponsive to CD3 stimulation and undergo enhanced lymphopenia-induced homeostatic proliferation, thus indicating that SIT inhibits TCR-mediated signaling. Here, we have further addressed how SIT regulates signaling cascades in T cells. We demonstrate that the loss of SIT enhances TCR-mediated Akt activation and increased phosphorylation/inactivation of Foxo1, a transcription factor of the Forkhead family that inhibits cell cycle progression and regulates T-cell homeostasis. We have also shown that CD4+ T cells from SIT-deficient mice display increased CD69 and CD40L expression indicating an altered activation status. Additional biochemical analyses further revealed that suppression of SIT expression by RNAi in human T cells resulted in an enhanced proximal TCR signaling. In summary, the data identify SIT as an important modulator of TCR-mediated signaling that regulates T-cell activation, homeostasis and tolerance. PMID:21957439

  14. Predicting Protein Subcellular Location Using Digital Signal Processing

    Institute of Scientific and Technical Information of China (English)

    Yu-Xi PAN; Da-Wei LI; Yun DUAN; Zhi-Zhou ZHANG; Ming-Qing XU; Guo-Yin FENG; Lin HE

    2005-01-01

    The biological functions of a protein are closely related to its attributes in a cell. With the rapid accumulation of newly found protein sequence data in databanks, it is highly desirable to develop an automated method for predicting the subcellular location of proteins. The establishment of such a predictor will expedite the functional determination of newly found proteins and the process of prioritizing genes and proteins identified by genomic efforts as potential molecular targets for drug design. The traditional algorithms for predicting these attributes were based solely on amino acid composition in which no sequence order effect was taken into account. To improve the prediction quality, it is necessary to incorporate such an effect. However, the number of possible patterns in protein sequences is extremely large, posing a formidable difficulty for realizing this goal. To deal with such difficulty, a well-developed tool in digital signal processing named digital Fourier transform (DFT) [1] was introduced. After being translated to a digital signal according to the hydrophobicity of each amino acid, a protein was analyzed by DFT within the frequency domain. A set of frequency spectrum parameters, thus obtained, were regarded as the factors to represent the sequence order effect. A significant improvement in prediction quality was observed by incorporating the frequency spectrum parameters with the conventional amino acid composition. One of the crucial merits of this approach is that many existing tools in mathematics and engineering can be easily applied in the predicting process. It is anticipated that digital signal processing may serve as a useful vehicle for many other protein science areas.

  15. Predicting protein subcellular location using digital signal processing.

    Science.gov (United States)

    Pan, Yu-Xi; Li, Da-Wei; Duan, Yun; Zhang, Zhi-Zhou; Xu, Ming-Qing; Feng, Guo-Yin; He, Lin

    2005-02-01

    The biological functions of a protein are closely related to its attributes in a cell. With the rapid accumulation of newly found protein sequence data in databanks, it is highly desirable to develop an automated method for predicting the subcellular location of proteins. The establishment of such a predictor will expedite the functional determination of newly found proteins and the process of prioritizing genes and proteins identified by genomic efforts as potential molecular targets for drug design. The traditional algorithms for predicting these attributes were based solely on amino acid composition in which no sequence order effect was taken into account. To improve the prediction quality, it is necessary to incorporate such an effect. However, the number of possible patterns in protein sequences is extremely large, posing a formidable difficulty for realizing this goal. To deal with such difficulty, a well-developed tool in digital signal processing named digital Fourier transform (DFT) [1] was introduced. After being translated to a digital signal according to the hydrophobicity of each amino acid, a protein was analyzed by DFT within the frequency domain. A set of frequency spectrum parameters, thus obtained, were regarded as the factors to represent the sequence order effect. A significant improvement in prediction quality was observed by incorporating the frequency spectrum parameters with the conventional amino acid composition. One of the crucial merits of this approach is that many existing tools in mathematics and engineering can be easily applied in the predicting process. It is anticipated that digital signal processing may serve as a useful vehicle for many other protein science areas.

  16. Membrane recruitment of scaffold proteins drives specific signaling.

    Directory of Open Access Journals (Sweden)

    Frédéric Pincet

    Full Text Available Cells must give the right response to each stimulus they receive. Scaffolding, a signaling process mediated by scaffold proteins, participates in the decoding of the cues by specifically directing signal transduction. The aim of this paper is to describe the molecular mechanisms of scaffolding, i.e. the principles by which scaffold proteins drive a specific response of the cell. Since similar scaffold proteins are found in many species, they evolved according to the purpose of each organism. This means they require adaptability. In the usual description of the mechanisms of scaffolding, scaffold proteins are considered as reactors where molecules involved in a cascade of reactions are simultaneously bound with the right orientation to meet and interact. This description is not realistic: (i it is not verified by experiments and (ii timing and orientation constraints make it complex which seems to contradict the required adaptability. A scaffold protein, Ste5, is used in the MAPK pathway of Saccharomyces cerevisiae for the cell to provide a specific response to stimuli. The massive amount of data available for this pathway makes it ideal to investigate the actual mechanisms of scaffolding. Here, a complete treatment of the chemical reactions allows the computation of the distributions of all the proteins involved in the MAPK pathway when the cell receives various cues. These distributions are compared to several experimental results. It turns out that the molecular mechanisms of scaffolding are much simpler and more adaptable than previously thought in the reactor model. Scaffold proteins bind only one molecule at a time. Then, their membrane recruitment automatically drives specific, amplified and localized signal transductions. The mechanisms presented here, which explain how the membrane recruitment of a protein can produce a drastic change in the activity of cells, are generic and may be commonly used in many biological processes.

  17. Biomolecular Simulation of Base Excision Repair and Protein Signaling

    Energy Technology Data Exchange (ETDEWEB)

    Straatsma, TP; McCammon, J A; Miller, John H; Smith, Paul E; Vorpagel, Erich R; Wong, Chung F; Zacharias, Martin W

    2006-03-03

    The goal of the Biomolecular Simulation of Base Excision Repair and Protein Signaling project is to enhance our understanding of the mechanism of human polymerase-β, one of the key enzymes in base excision repair (BER) and the cell-signaling enzymes cyclic-AMP-dependent protein kinase. This work used molecular modeling and simulation studies to specifically focus on the • dynamics of DNA and damaged DNA • dynamics and energetics of base flipping in DNA • mechanism and fidelity of nucleotide insertion by BER enzyme human polymerase-β • mechanism and inhibitor design for cyclic-AMP-dependent protein kinase. Molecular dynamics simulations and electronic structure calculations have been performed using the computer resources at the Molecular Science Computing Facility at the Environmental Molecular Sciences Laboratory.

  18. Characterization of signalling pathways by reverse phase protein arrays.

    Science.gov (United States)

    Malinowsky, Katharina; Wolff, Claudia; Schott, Christina; Becker, Karl-Friedrich

    2013-01-01

    Reverse phase protein array (RPPA) is a very suitable technique to analyze large numbers of proteins in small samples like for example tumor biopsies. Beside their small size another major hindrance for the analysis of proteins from biopsies is the extraction of proteins from formalin-fixed and paraffin-embedded (FFPE) tissues. Here we describe a protocol, allowing quantitative extraction of large numbers of proteins from FFPE tissues and their subsequent analysis by RPPA. To elucidate the role of epidermal growth factor receptor (EGFR) signalling in ovarian cancer, we analyzed 23 primary tumors and corresponding metastases for the expression of 25 proteins involved in EGFR signalling with special emphasis on epithelial-mesenchymal transition (EMT). We found a significant correlation of Snail with EGFR((Tyr1086)) and p38 MAPK((Thr180/Tyr182)) in primary ovarian carcinoma and with EGFR((Tyr1086)) in their corresponding metastases. Additionally, we showed that high expression levels of the E-cadherin repressor Snail in primary tumors combined with high expression levels of the pp38 MAPK((Thr180/Tyr182)) in metastasis lead to an increased risk for death in ovarian carcinoma patients.

  19. Structural features of the two-component system LisR/LisK suggests multiple responses for the adaptation and survival of Listeria monocytogenes

    Directory of Open Access Journals (Sweden)

    Nelson Enrique Arenas Suarez

    2013-08-01

    Full Text Available Here, we characterized the structure of the two-component regulatory system, LisR/LisK, in Listeria monocytogenes. To predict the structure of both proteins and the relationship between them, we employed several bioinformatic tools and databases. Based on our results, LisK protein is embedded in the cell membrane and its modular composition (HAMP, histidine kinase and ATPase domains is associated with its autophosphorylation (His-266. A stimulus-response likely determines the sequential signal propagation from the bacterial cell surface to its cytoplasmic components. According to our results, LisR is a cytoplasmic protein with a receptor domain (homologous to CheY that comprises a phosphoacceptor residue (Asp-52 and a DNA-binding domain, which may allow the transmission of a specific transcriptional response. LisR/LisK has been experimentally characterized both biochemically andfunctionally in other Bacilli pathophysiology; our structure-function approach may facilitate the design of suitable inhibitors.

  20. XB130: A novel adaptor protein in cancer signal transduction

    Science.gov (United States)

    ZHANG, RUIYAO; ZHANG, JINGYAO; WU, QIFEI; MENG, FANDI; LIU, CHANG

    2016-01-01

    Adaptor proteins are functional proteins that contain two or more protein-binding modules to link signaling proteins together, which affect cell growth and shape and have no enzymatic activity. The actin filament-associated protein (AFAP) family is an important member of the adaptor proteins, including AFAP1, AFAP1L1 and AFAP1L2/XB130. AFAP1 and AFAP1L1 share certain common characteristics and function as an actin-binding protein and a cSrc-activating protein. XB130 exhibits certain unique features in structure and function. The mRNA of XB130 is expressed in human spleen, thyroid, kidney, brain, lung, pancreas, liver, colon and stomach, and the most prominent disease associated with XB130 is cancer. XB130 has a controversial effect on cancer. Studies have shown that XB130 can promote cancer progression and downregulation of XB130-reduced growth of tumors derived from certain cell lines. A higher mRNA level of XB130 was shown to be associated with a better survival in non-small cell lung cancer. Previous studies have shown that XB130 can regulate cell growth, migration and invasion and possibly has the effect through the cAMP-cSrc-phosphoinositide 3-kinase/Akt pathway. Except for cancer, XB130 is also associated with other pathological or physiological procedures, such as airway repair and regeneration. PMID:26998266

  1. Post-translational modification of PII signal transduction proteins

    Directory of Open Access Journals (Sweden)

    Mike eMerrick

    2015-01-01

    Full Text Available The PII proteins constitute one of the most widely distributed families of signal transduction proteins in nature. They are pivotal players in the control of nitrogen metabolism in bacteria and archaea, and are also found in the plastids of plants. Quite remarkably PII proteins control the activities of a diverse range of enzymes, transcription factors and membrane transport proteins, and in all known cases they achieve their regulatory effect by direct interaction with their target. PII proteins in the Proteobacteria and the Actinobacteria are subject to post-translational modification by uridylylation or adenylylation respectively, whilst in some Cyanobacteria they can be modified by phosphorylation. In all these cases the protein’s modification state is influenced by the cellular nitrogen status and is thought to regulate its activity. However in many organisms there is no evidence for modification of PII proteins and indeed the ability of these proteins to respond to the cellular nitrogen status is fundamentally independent of post-translational modification. In this review we explore the role of post-translational modification in PII proteins in the light of recent studies.

  2. Implementation of Two Component Advective Flow Solution in XSPEC

    CERN Document Server

    Debnath, Dipak; Mondal, Santanu

    2014-01-01

    Spectral and Temporal properties of black hole candidates can be explained reasonably well using Chakrabarti-Titarchuk solution of two component advective flow (TCAF). This model requires two accretion rates, namely, the Keplerian disk accretion rate and the halo accretion rate, the latter being composed of a sub-Keplerian, low angular momentum flow which may or may not develop a shock. In this solution, the relevant parameter is the relative importance of the halo (which creates the Compton cloud region) rate with respect to the Keplerian disk rate (soft photon source). Though this model has been used earlier to manually fit data of several black hole candidates quite satisfactorily, for the first time, we made it user friendly by implementing it into XSPEC software of GSFC/NASA. This enables any user to extract physical parameters of the accretion flows, such as two accretion rates, the shock location, the shock strength etc. for any black hole candidate. We provide some examples of fitting a few cases usin...

  3. Hamiltonian of a homogeneous two-component plasma.

    Science.gov (United States)

    Essén, Hanno; Nordmark, A

    2004-03-01

    The Hamiltonian of one- and two-component plasmas is calculated in the negligible radiation Darwin approximation. Since the Hamiltonian is the phase space energy of the system its form indicates, according to statistical mechanics, the nature of the thermal equilibrium that plasmas strive to attain. The main issue is the length scale of the magnetic interaction energy. In the past a screening length lambda=1/square root of r(e)n], with n number density and r(e) classical electron radius, has been derived. We address the question whether the corresponding longer screening range obtained from the classical proton radius is physically relevant and the answer is affirmative. Starting from the Darwin Lagrangian it is nontrivial to find the Darwin Hamiltonian of a macroscopic system. For a homogeneous system we resolve the difficulty by temporarily approximating the particle number density by a smooth constant density. This leads to Yukawa-type screened vector potential. The nontrivial problem of finding the corresponding, divergence free, Coulomb gauge version is solved.

  4. A minimal model for two-component dark matter

    CERN Document Server

    Esch, Sonja; Yaguna, Carlos E

    2014-01-01

    We propose and study a new minimal model for two-component dark matter. The model contains only three additional fields, one fermion and two scalars, all singlets under the Standard Model gauge group. Two of these fields, one fermion and one scalar, are odd under a $Z_2$ symmetry that renders them simultaneously stable. Thus, both particles contribute to the observed dark matter density. This model resembles the union of the singlet scalar and the singlet fermionic models but it contains some new features of its own. We analyze in some detail its dark matter phenomenology. Regarding the relic density, the main novelty is the possible annihilation of one dark matter particle into the other, which can affect the predicted relic density in a significant way. Regarding dark matter detection, we identify a new contribution that can lead either to an enhancement or to a suppression of the spin-independent cross section for the scalar dark matter particle. Finally, we define a set of five benchmarks models compatibl...

  5. Two-component perfect fluid in FRW universe

    CERN Document Server

    ,

    2012-01-01

    We propose the cosmological model which allows to describe on equal footing the evolution of matter in the universe on the time interval from the inflation till the domination of dark energy. The matter is considered as a two-component perfect fluid imitated by homogeneous scalar fields between which there is energy exchange. Dark energy is represented by the cosmological constant, which is supposed invariable during the whole evolution of the universe. The matter changes its equation of state with time, so that the era of radiation domination in the early universe smoothly passes into the era of a pressureless gas, which then passes into the late-time epoch, when the matter is represented by a gas of low-velocity cosmic strings. The inflationary phase is described as an analytic continuation of the energy density in the very early universe into the region of small negative values of the parameter which characterizes typical time of energy transfer from one matter component to another. The Hubble expansion ra...

  6. Biased and G protein-independent signaling of chemokine receptors

    Directory of Open Access Journals (Sweden)

    Anne eSteen

    2014-06-01

    Full Text Available Biased signaling or functional selectivity occurs when a 7TM receptor preferentially activates one of several available pathways. It can be divided into three distinct forms: ligand bias, receptor bias, and tissue or cell bias, where it is mediated by different ligands (on the same receptor, different receptors (with the same ligand or different tissues or cells (for the same ligand-receptor pair. Most often biased signaling is differentiated into G protein-dependent and β-arrestin-dependent signaling. Yet, it may also cover signaling differences within these groups. Moreover, it may not be absolute, i.e. full versus no activation. Here we discuss biased signaling in the chemokine system, including the structural basis for biased signaling in chemokine receptors, as well as in class A 7TM receptors in general. This includes overall helical movements and the contributions of micro-switches based on recently published 7TM crystals and molecular dynamics studies. All three forms of biased signaling are abundant in the chemokine system. This challenges our understanding of classic redundancy inevitably ascribed to this system, where multiple chemokines bind to the same receptor and where a single chemokine may bind to several receptors – in both cases with the same functional outcome. The ubiquitous biased signaling confer a hitherto unknown specificity to the chemokine system with a complex interaction pattern that is better described as promiscuous with context-defined roles and different functional outcomes in a ligand-, receptor- or cell/tissue-defined manner. As the low number of successful drug development plans implies, there are great difficulties in targeting chemokine receptors; in particular with regard to receptor antagonists as anti-inflammatory drugs. Un-defined and putative non-selective targeting of the complete cellular signaling system could be the underlying cause of lack of success. Therefore, biased ligands could be the

  7. Identification and quantitation of signal molecule-dependent protein phosphorylation

    KAUST Repository

    Groen, Arnoud J.

    2013-09-03

    Phosphoproteomics is a fast-growing field that aims at characterizing phosphorylated proteins in a cell or a tissue at a given time. Phosphorylation of proteins is an important regulatory mechanism in many cellular processes. Gel-free phosphoproteome technique involving enrichment of phosphopeptide coupled with mass spectrometry has proven to be invaluable to detect and characterize phosphorylated proteins. In this chapter, a gel-free quantitative approach involving 15N metabolic labelling in combination with phosphopeptide enrichment by titanium dioxide (TiO2) and their identification by MS is described. This workflow can be used to gain insights into the role of signalling molecules such as cyclic nucleotides on regulatory networks through the identification and quantification of responsive phospho(proteins). © Springer Science+Business Media New York 2013.

  8. Two-component systems in microbial communities: approaches and resources for generating and analyzing metagenomic data sets.

    Science.gov (United States)

    Podar, Mircea

    2007-01-01

    Two-component signal transduction represents the main mechanism by which bacterial cells interact with their environment. The functional diversity of two-component systems and their relative importance in the different taxonomic groups and ecotypes of bacteria has become evident with the availability of several hundred genomic sequences. The vast majority of bacteria, including many high rank taxonomic units, while being components of complex microbial communities remain uncultured (i.e., have not been isolated or grown in the laboratory). Environmental genomic data from such communities are becoming available, and in addition to its profound impact on microbial ecology it will propel molecular biological disciplines beyond the traditional model organisms. This chapter describes the general approaches used in generating environmental genomic data and how that data can be used to advance the study of two component-systems and signal transduction in general.

  9. Bioinformatics analysis of two-component regulatory systems in Staphylococcus epidermidis

    Institute of Scientific and Technical Information of China (English)

    QIN Zhiqiang; ZHONG Yang; ZHANG Jian; HE Youyu; WU Yang; JIANG Juan; CHEN Jiemin; LUO Xiaomin; QU Di

    2004-01-01

    Sixteen pairs of two-component regulatory systems are identified in the genome of Staphylococcus epidermidis ATCC12228 strain, which is newly sequenced by our laboratory for Medical Molecular Virology and Chinese National Human Genome Center at Shanghai, by using bioinformatics analysis. Comparative analysis of the twocomponent regulatory systems in S. epidermidis and that of S.aureus and Bacillus subtilis shows that these systems may regulate some important biological functions, e.g. growth,biofilm formation, and expression of virulence factors in S.epidermidis. Two conserved domains, i.e. HATPase_c and REC domains, are found in all 16 pairs of two-component proteins.Homologous modelling analysis indicates that there are 4similar HATPase_c domain structures of histidine kinases and 13 similar REC domain structures of response regulators,and there is one AMP-PNP binding pocket in the HATPase_c domain and three active aspartate residues in the REC domain. Preliminary experiment reveals that the bioinformatics analysis of the conserved domain structures in the two-component regulatory systems in S. epidermidis may provide useful information for discovery of potential drug target.

  10. Inhibition of Protein-Protein Interactions and Signaling by Small Molecules

    Science.gov (United States)

    Freire, Ernesto

    2010-03-01

    Protein-protein interactions are at the core of cell signaling pathways as well as many bacterial and viral infection processes. As such, they define critical targets for drug development against diseases such as cancer, arthritis, obesity, AIDS and many others. Until now, the clinical inhibition of protein-protein interactions and signaling has been accomplished with the use of antibodies or soluble versions of receptor molecules. Small molecule replacements of these therapeutic agents have been extremely difficult to develop; either the necessary potency has been hard to achieve or the expected biological effect has not been obtained. In this presentation, we show that a rigorous thermodynamic approach that combines differential scanning calorimetry (DSC) and isothermal titration calorimetry (ITC) provides a unique platform for the identification and optimization of small molecular weight inhibitors of protein-protein interactions. Recent advances in the development of cell entry inhibitors of HIV-1 using this approach will be discussed.

  11. Protein tyrosine nitration in cellular signal transduction pathways

    Science.gov (United States)

    Yakovlev, Vasily A.; Mikkelsen, Ross B.

    2015-01-01

    How specificity and reversibility in tyrosine nitration are defined biologically in cellular systems is poorly understood. As more investigations identify proteins involved in cell regulatory pathways in which only a small fraction of that protein pool is modified by nitration to affect cell function, the mechanisms of biological specificity and reversal should come into focus. In this review experimental evidence has been summarized to suggest that tyrosine nitration is a highly selective modification and under certain physiological conditions fulfills the criteria of a physiologically relevant signal. It can be specific, reversible, occurs on a physiological time scale, and, depending on a target, can result in either activation or inhibition. PMID:20843272

  12. One-component systems dominate signal transduction in prokaryotes

    OpenAIRE

    Ulrich, Luke E; Koonin, Eugene V.; Zhulin, Igor B.

    2005-01-01

    Two-component systems that link environmental signals to cellular responses are viewed as the primary mode of signal transduction in prokaryotes. By analyz-ing information encoded by 145 prokaryotic genomes, we found that the majority of signal transduction systems consist of a single protein that contains input and output domains but lacks phosphotransfer domains typical of two-component systems. One-component systems are evolutionarily older, more widely distributed among bacteria and archa...

  13. Conventional and novel Gγ protein families constitute the heterotrimeric G-protein signaling network in soybean.

    Directory of Open Access Journals (Sweden)

    Swarup Roy Choudhury

    Full Text Available Heterotrimeric G-proteins comprised of Gα, Gβ and Gγ proteins are important signal transducers in all eukaryotes. The Gγ protein of the G-protein heterotrimer is crucial for its proper targeting at the plasma membrane and correct functioning. Gγ proteins are significantly smaller and more diverse than the Gα and Gβ proteins. In model plants Arabidopsis and rice that have a single Gα and Gβ protein, the presence of two canonical Gγ proteins provide some diversity to the possible heterotrimeric combinations. Our recent analysis of the latest version of the soybean genome has identified ten Gγ proteins which belong to three distinct families based on their C-termini. We amplified the full length cDNAs, analyzed their detailed expression profile by quantitative PCR, assessed their localization and performed yeast-based interaction analysis to evaluate interaction specificity with different Gβ proteins. Our results show that ten Gγ genes are retained in the soybean genome and have interesting expression profiles across different developmental stages. Six of the newly identified proteins belong to two plant-specific Gγ protein families. Yeast-based interaction analyses predict some degree of interaction specificity between different Gβ and Gγ proteins. This research thus identifies a highly diverse G-protein network from a plant species. Homologs of these novel proteins have been previously identified as QTLs for grain size and yield in rice.

  14. The hedgehog signal induced modulation of bone morphogenetic protein signaling: an essential signaling relay for urinary tract morphogenesis.

    Directory of Open Access Journals (Sweden)

    Ryuma Haraguchi

    Full Text Available BACKGROUND: Congenital diseases of the urinary tract are frequently observed in infants. Such diseases present a number of developmental anomalies such as hydroureter and hydronephrosis. Although some genetically-modified mouse models of growth factor signaling genes reproduce urinary phenotypes, the pathogenic mechanisms remain obscure. Previous studies suggest that a portion of the cells in the external genitalia and bladder are derived from peri-cloacal mesenchymal cells that receive Hedgehog (Hh signaling in the early developmental stages. We hypothesized that defects in such progenitor cells, which give rise to urinary tract tissues, may be a cause of such diseases. METHODOLOGY/PRINCIPAL FINDINGS: To elucidate the pathogenic mechanisms of upper urinary tract malformations, we analyzed a series of Sonic hedgehog (Shh deficient mice. Shh(-/- displayed hydroureter and hydronephrosis phenotypes and reduced expression of several developmental markers. In addition, we suggested that Shh modulation at an early embryonic stage is responsible for such phenotypes by analyzing the Shh conditional mutants. Tissue contribution assays of Hh-responsive cells revealed that peri-cloacal mesenchymal cells, which received Hh signal secreted from cloacal epithelium, could contribute to the ureteral mesenchyme. Gain- and loss-of-functional mutants for Hh signaling revealed a correlation between Hh signaling and Bone morphogenetic protein (Bmp signaling. Finally, a conditional ablation of Bmp receptor type IA (BmprIA gene was examined in Hh-responsive cell lineages. This system thus made it possible to analyze the primary functions of the growth factor signaling relay. The defective Hh-to-Bmp signaling relay resulted in severe urinary tract phenotypes with a decrease in the number of Hh-responsive cells. CONCLUSIONS/SIGNIFICANCE: This study identified the essential embryonic stages for the pathogenesis of urinary tract phenotypes. These results suggested that Hh

  15. S-Glutathionylation and Redox Protein Signaling in Drug Addiction.

    Science.gov (United States)

    Womersley, Jacqueline S; Uys, Joachim D

    2016-01-01

    Drug addiction is a chronic relapsing disorder that comes at a high cost to individuals and society. Therefore understanding the mechanisms by which drugs exert their effects is of prime importance. Drugs of abuse increase the production of reactive oxygen and nitrogen species resulting in oxidative stress. This change in redox homeostasis increases the conjugation of glutathione to protein cysteine residues; a process called S-glutathionylation. Although traditionally regarded as a protective mechanism against irreversible protein oxidation, accumulated evidence suggests a more nuanced role for S-glutathionylation, namely as a mediator in redox-sensitive protein signaling. The reversible modification of protein thiols leading to alteration in function under different physiologic/pathologic conditions provides a mechanism whereby change in redox status can be translated into a functional response. As such, S-glutathionylation represents an understudied means of post-translational protein modification that may be important in the mechanisms underlying drug addiction. This review will discuss the evidence for S-glutathionylation as a redox-sensing mechanism and how this may be involved in the response to drug-induced oxidative stress. The function of S-glutathionylated proteins involved in neurotransmission, dendritic spine structure, and drug-induced behavioral outputs will be reviewed with specific reference to alcohol, cocaine, and heroin. Copyright © 2016. Published by Elsevier Inc.

  16. APP Protein Family Signaling at the Synapse: Insights from Intracellular APP-Binding Proteins.

    Science.gov (United States)

    Guénette, Suzanne; Strecker, Paul; Kins, Stefan

    2017-01-01

    Understanding the molecular mechanisms underlying amyloid precursor protein family (APP/APP-like proteins, APLP) function in the nervous system can be achieved by studying the APP/APLP interactome. In this review article, we focused on intracellular APP interacting proteins that bind the YENPTY internalization motif located in the last 15 amino acids of the C-terminal region. These proteins, which include X11/Munc-18-interacting proteins (Mints) and FE65/FE65Ls, represent APP cytosolic binding partners exhibiting different neuronal functions. A comparison of FE65 and APP family member mutant mice revealed a shared function for APP/FE65 protein family members in neurogenesis and neuronal positioning. Accumulating evidence also supports a role for membrane-associated APP/APLP proteins in synapse formation and function. Therefore, it is tempting to speculate that APP/APLP C-terminal interacting proteins transmit APP/APLP-dependent signals at the synapse. Herein, we compare our current knowledge of the synaptic phenotypes of APP/APLP mutant mice with those of mice lacking different APP/APLP interaction partners and discuss the possible downstream effects of APP-dependent FE65/FE65L or X11/Mint signaling on synaptic vesicle release, synaptic morphology and function. Given that the role of X11/Mint proteins at the synapse is well-established, we propose a model highlighting the role of FE65 protein family members for transduction of APP/APLP physiological function at the synapse.

  17. The Fractional Virial Potential Energy in Two-Component Systems

    Directory of Open Access Journals (Sweden)

    Caimmi, R.

    2008-12-01

    Full Text Available Two-component systems are conceived as macrogases, and the related equation of state is expressed using the virial theorem for subsystems, under the restriction of homeoidally striated density profiles. Explicit calculations are performed for a useful reference case and a few cases of astrophysical interest, both with and without truncation radius. Shallower density profiles are found to yield an equation of state, $phi=phi(y,m$, characterized (for assigned values of the fractional mass, $m=M_j/ M_i$ by the occurrence of two extremum points, a minimum and a maximum, as found in an earlier attempt. Steeper density profiles produce a similar equation of state, which implies that a special value of $m$ is related to a critical curve where the above mentioned extremum points reduce to a single horizontal inflexion point, and curves below the critical one show no extremum points. The similarity of the isofractional mass curves to van der Waals' isothermal curves, suggests the possibility of a phase transition in a bell-shaped region of the $({sf O}yphi$ plane, where the fractional truncation radius along a selected direction is $y=R_j/R_i$, and the fractional virial potential energy is $phi=(E_{ji}_mathrm{vir}/(E_{ij}_mathrm{vir}$. Further investigation is devoted to mass distributions described by Hernquist (1990 density profiles, for which an additional relation can be used to represent a sample of $N=16$ elliptical galaxies (EGs on the $({sf O}yphi$ plane. Even if the evolution of elliptical galaxies and their hosting dark matter (DM haloes, in the light of the model, has been characterized by equal fractional mass, $m$, and equal scaled truncation radius, or concentration, $Xi_u=R_u/r_u^dagger$, $u=i,j$, still it cannot be considered as strictly homologous, due to different values of fractional truncation radii, $y$, or fractional scaling radii, $y^dagger=r_j^dagger/r_i^dagger$, deduced from sample objects.

  18. Output Rate of Atomic Four-Wave Mixing in Two-Component Bose-Einstein Condensate

    Institute of Scientific and Technical Information of China (English)

    LI Jia-Hua; LI Wei-Bing; PENG Ju-Cun

    2004-01-01

    In this letter, following the proposal of Heurich et al. [Phys. Rev. A63 (2001) 033605], we analyze and discuss output rate of atomic four-wave mixing in the two-component Bose-Einstein condensate under the condition of the steady state. The results show that the magnitude of the signal beam increases with the increase of the intensity of the probe beam, up to a saturated value, then it decreases as the probe beam increases. The influence of the interaction range on the signal beam is also predicted. In particular, it is worth while pointing out that in contrast to the previous solutions, our obtained analytical solutions are of very simple and explicit forms, which open the door for further investigating the related physical mechanisms.

  19. A two-component system regulates hemin acquisition in Porphyromonas gingivalis.

    Directory of Open Access Journals (Sweden)

    Jodie C Scott

    Full Text Available Porphyromonas gingivalis is a Gram-negative oral anaerobe associated with infection of the periodontia. The organism has a small number of two-component signal transduction systems, and after comparing genome sequences of strains W83 and ATCC 33277 we discovered that the latter was mutant in histidine kinase (PGN_0752, while the cognate response regulator (PGN_0753 remained intact. Microarray-based transcriptional profiling and ChIP-seq assays were carried out with an ATCC 33277 transconjugant containing the functional histidine kinase from strain W83 (PG0719. The data showed that the regulon of this signal transduction system contained genes that were involved in hemin acquisition, including gingipains, at least three transport systems, as well as being self-regulated. Direct regulation by the response regulator was confirmed by electrophoretic mobility shift assays. In addition, the system appears to be activated by hemin and the regulator acts as both an activator and repressor.

  20. A proteomics strategy to elucidate functional protein-protein interactions applied to EGF signaling

    DEFF Research Database (Denmark)

    Blagoev, B.; Kratchmarova, I.; Ong, S.E.

    2003-01-01

    Mass spectrometry-based proteomics can reveal protein-protein interactions on a large scale, but it has been difficult to separate background binding from functionally important interactions and still preserve weak binders. To investigate the epidermal growth factor receptor (EGFR) pathway, we...... and Src homologous and collagen (Shc) protein. We identified 228 proteins, of which 28 were selectively enriched upon stimulation. EGFR and Shc, which interact directly with the bait, had large differential ratios. Many signaling molecules specifically formed complexes with the activated EGFR-Shc, as did...... plectin, epiplakin, cytokeratin networks, histone H3, the glycosylphosphatidylinositol (GPI)-anchored molecule CD59, and two novel proteins. SILAC combined with modification-based affinity purification is a useful approach to detect specific and functional protein-protein interactions....

  1. A Two-Component Regulatory System Integrates Redox State and Population Density Sensing in Pseudomonas putida▿ †

    OpenAIRE

    Fernández-Piñar, Regina; Ramos, Juan Luis; Rodríguez-Herva, José Juan; Espinosa-Urgel, Manuel

    2008-01-01

    A two-component system formed by a sensor histidine kinase and a response regulator has been identified as an element participating in cell density signal transduction in Pseudomonas putida KT2440. It is a homolog of the Pseudomonas aeruginosa RoxS/RoxR system, which in turn belongs to the RegA/RegB family, described in photosynthetic bacteria as a key regulatory element. In KT2440, the two components are encoded by PP_0887 (roxS) and PP_0888 (roxR), which are transcribed in a single unit. Ch...

  2. Dynamically Tunable Memory in Two-Component Gene Circuit

    Energy Technology Data Exchange (ETDEWEB)

    Ghim, C; Almaas, E

    2008-09-05

    Cell has the potential to remember the environmental conditions for many (10{sup 7}) generations but stochastic fluctuations set a fundamental limit on the stability of this memory. Here we explicitly take the binding-unbinding of macromolecules into account to propose a novel rationale for the protein-protein interaction in cell physiology. Based on the first-exit time and the corresponding deterministic characterization of various genetic circuits, we show that the reversible binding dynamics may stabilize non-genetically inherited cell states, providing a practical strategy for designing robust epigenetic memory.

  3. Systematic Prediction of Scaffold Proteins Reveals New Design Principles in Scaffold-Mediated Signal Transduction

    Science.gov (United States)

    Hu, Jianfei; Neiswinger, Johnathan; Zhang, Jin; Zhu, Heng; Qian, Jiang

    2015-01-01

    Scaffold proteins play a crucial role in facilitating signal transduction in eukaryotes by bringing together multiple signaling components. In this study, we performed a systematic analysis of scaffold proteins in signal transduction by integrating protein-protein interaction and kinase-substrate relationship networks. We predicted 212 scaffold proteins that are involved in 605 distinct signaling pathways. The computational prediction was validated using a protein microarray-based approach. The predicted scaffold proteins showed several interesting characteristics, as we expected from the functionality of scaffold proteins. We found that the scaffold proteins are likely to interact with each other, which is consistent with previous finding that scaffold proteins tend to form homodimers and heterodimers. Interestingly, a single scaffold protein can be involved in multiple signaling pathways by interacting with other scaffold protein partners. Furthermore, we propose two possible regulatory mechanisms by which the activity of scaffold proteins is coordinated with their associated pathways through phosphorylation process. PMID:26393507

  4. Systematic Prediction of Scaffold Proteins Reveals New Design Principles in Scaffold-Mediated Signal Transduction.

    Directory of Open Access Journals (Sweden)

    Jianfei Hu

    Full Text Available Scaffold proteins play a crucial role in facilitating signal transduction in eukaryotes by bringing together multiple signaling components. In this study, we performed a systematic analysis of scaffold proteins in signal transduction by integrating protein-protein interaction and kinase-substrate relationship networks. We predicted 212 scaffold proteins that are involved in 605 distinct signaling pathways. The computational prediction was validated using a protein microarray-based approach. The predicted scaffold proteins showed several interesting characteristics, as we expected from the functionality of scaffold proteins. We found that the scaffold proteins are likely to interact with each other, which is consistent with previous finding that scaffold proteins tend to form homodimers and heterodimers. Interestingly, a single scaffold protein can be involved in multiple signaling pathways by interacting with other scaffold protein partners. Furthermore, we propose two possible regulatory mechanisms by which the activity of scaffold proteins is coordinated with their associated pathways through phosphorylation process.

  5. DHHC protein-dependent palmitoylation protects regulator of G-protein signaling 4 from proteasome degradation

    OpenAIRE

    2010-01-01

    Regulator of G-protein signaling 4 (RGS4), an intracellular modulator of G-protein coupled receptor (GPCR)-mediated signaling, is regulated by multiple processes including palmitoylation and proteasome degradation. We found that co-expression of DHHC acyltransferases (DHHC3 or DHHC7), but not their acyltransferase-inactive mutants, increased expression levels of RGS4 but not its Cys2 to Ser mutant (RGS4C2S). DHHC3 interacts with and palmitoylates RGS4 but not RGS4C2S in vivo. Palmitoylation p...

  6. Prolactin receptor and signal transduction to milk protein genes

    Energy Technology Data Exchange (ETDEWEB)

    Djiane, J.; Daniel, N.; Bignon, C. [Unite d`Endocrinologie Moleculaire, Jouy en Josas (France)] [and others

    1994-06-01

    After cloning of the mammary gland prolactin (PRL) receptor cDNA, a functional assay was established using co-transfection of PRL receptor cDNA together with a milk protein promoter/chloramphenicol acetyl transferase (CAT) construct in Chinese hamster ovary (CHO) cells. Different mutants of the PRL receptor were tested in this CAT assay to delimit the domains in the receptor necessary for signal transduction to milk protein genes. In CHO cells stably transfected with PRL receptor cDNA, high numbers of PRL receptor are expressed. By metabolic labeling and immunoprecipitation, expressed PRL receptor was identified as a single species of 100 kDa. Using these cells, we analyzed the effects of PRL on intracellular free Ca{sup ++} concentration. PRL stimulates Ca{sup ++} entry and induces secondary Ca{sup ++} mobilization. The entry of Ca{sup ++} is a result of an increase in K{sup +} conductance that hyperpolarizes the membranes. We have also analyzed tyrosine phosphorylation induced by PRL. In CHO cells stably transfected with PRL receptor cDNA, PRL induced a very rapid and transient tyrosine phosphorylation of a 100-kDa protein which is most probably the PRL receptor. The same finding was obtained in mammary membranes after PRL injection to lactating rabbits. Whereas tyrosine kinase inhibitors genistein and lavendustin were without effect, PRL stimulation of milk protein gene promoters was partially inhibited by 2 {mu}M herbimycin in CHO cells co-transfected with PRL receptor cDNA and the {Beta} lactoglobulin CAT construct. Taken together these observations indicate that the cytoplasmic domain of the PRL receptor interacts with one or several tyrosine kinases, which may represent early postreceptor events necessary for PRL signal transduction to milk protein genes. 14 refs., 4 figs.

  7. Protein Kinase C and Toll-Like Receptor Signaling

    Directory of Open Access Journals (Sweden)

    Daniel J. Loegering

    2011-01-01

    Full Text Available Protein kinase C (PKC is a family of kinases that are implicated in a plethora of diseases, including cancer and cardiovascular disease. PKC isoforms can have different, and sometimes opposing, effects in these disease states. Toll-like receptors (TLRs are a family of pattern recognition receptors that bind pathogens and stimulate the secretion of cytokines. It has long been known that PKC inhibitors reduce LPS-stimulated cytokine secretion by macrophages, linking PKC activation to TLR signaling. Recent studies have shown that PKC-α, -δ, -ε, and -ζ are directly involved in multiple steps in TLR pathways. They associate with the TLR or proximal components of the receptor complex. These isoforms are also involved in the downstream activation of MAPK, RhoA, TAK1, and NF-κB. Thus, PKC activation is intimately involved in TLR signaling and the innate immune response.

  8. Continuous monitoring of membrane protein micro-domain association during cell signaling

    CERN Document Server

    Huang, Heng

    2011-01-01

    Central to understanding membrane bound cell signaling is to quantify how the membrane ultra-structure consisting of transient spatial domains modulates signaling and how the signaling influences this ultra-structure. Yet, measuring the association of membrane proteins with domains in living, intact cells poses considerable challenges. Here, we describe a non-destructive method to quantify protein-lipid domain and protein cytoskeleton interactions in single, intact cells enabling continuous monitoring of the protein domains interaction over time during signaling.

  9. Regulation of longevity by regulator of G-protein signaling protein, Loco.

    Science.gov (United States)

    Lin, Yuh-Ru; Kim, Keetae; Yang, Yanfei; Ivessa, Andreas; Sadoshima, Junichi; Park, Yongkyu

    2011-06-01

    Regulator of G-protein signaling (RGS) proteins contribute to G-protein signaling pathways as activators or repressors with GTPase-activating protein (GAP) activity. To characterize whether regulation of RGS proteins influences longevity in several species, we measured stress responses and lifespan of RGS-overexpressing and RGS-lacking mutants. Reduced expression of Loco, a RGS protein of Drosophila melanogaster, resulted in a longer lifespan for both male and female flies, also exhibiting stronger resistance to three different stressors (starvation, oxidation, and heat) and higher manganese-containing superoxide dismutase (MnSOD) activity. In addition, this reduction in Loco expression increased fat content and diminished cAMP levels. In contrast, overexpression of both genomic and cDNA loco gene significantly shortened the lifespan with weaker stress resistance and lower fat content. Deletion analysis of the Loco demonstrated that its RGS domain is required for the regulation of longevity. Consistently, when expression of RGS14, mammalian homologue of Loco, was reduced in rat fibroblast cells, the resistance to oxidative stress increased with higher MnSOD expression. The changes of yeast Rgs2 expression, which shares a conserved RGS domain with the fly Loco protein, also altered lifespan and stress resistance in Saccharomyces cerevisiae. Here, we provide the first evidence that RGS proteins with GAP activity affect both stress resistance and longevity in several species.

  10. The origins of the evolutionary signal used to predict protein-protein interactions

    Directory of Open Access Journals (Sweden)

    Swapna Lakshmipuram S

    2012-12-01

    Full Text Available Abstract Background The correlation of genetic distances between pairs of protein sequence alignments has been used to infer protein-protein interactions. It has been suggested that these correlations are based on the signal of co-evolution between interacting proteins. However, although mutations in different proteins associated with maintaining an interaction clearly occur (particularly in binding interfaces and neighbourhoods, many other factors contribute to correlated rates of sequence evolution. Proteins in the same genome are usually linked by shared evolutionary history and so it would be expected that there would be topological similarities in their phylogenetic trees, whether they are interacting or not. For this reason the underlying species tree is often corrected for. Moreover processes such as expression level, are known to effect evolutionary rates. However, it has been argued that the correlated rates of evolution used to predict protein interaction explicitly includes shared evolutionary history; here we test this hypothesis. Results In order to identify the evolutionary mechanisms giving rise to the correlations between interaction proteins, we use phylogenetic methods to distinguish similarities in tree topologies from similarities in genetic distances. We use a range of datasets of interacting and non-interacting proteins from Saccharomyces cerevisiae. We find that the signal of correlated evolution between interacting proteins is predominantly a result of shared evolutionary rates, rather than similarities in tree topology, independent of evolutionary divergence. Conclusions Since interacting proteins do not have tree topologies that are more similar than the control group of non-interacting proteins, it is likely that coevolution does not contribute much to, if any, of the observed correlations.

  11. One-component systems dominate signal transduction in prokaryotes

    Science.gov (United States)

    Ulrich, Luke E.; Koonin, Eugene V.; Zhulin, Igor B.

    2009-01-01

    Two-component systems that link environmental signals to cellular responses are viewed as the primary mode of signal transduction in prokaryotes. By analyz-ing information encoded by 145 prokaryotic genomes, we found that the majority of signal transduction systems consist of a single protein that contains input and output domains but lacks phosphotransfer domains typical of two-component systems. One-component systems are evolutionarily older, more widely distributed among bacteria and archaea, and display a greater diversity of domains than two-component systems. PMID:15680762

  12. Mitogen Activated Protein kinase signal transduction pathways in the prostate

    Directory of Open Access Journals (Sweden)

    Koul Sweaty

    2004-06-01

    Full Text Available Abstract The biochemistry of the mitogen activated protein kinases ERK, JNK, and p38 have been studied in prostate physiology in an attempt to elucidate novel mechanisms and pathways for the treatment of prostatic disease. We reviewed articles examining mitogen-activated protein kinases using prostate tissue or cell lines. As with other tissue types, these signaling modules are links/transmitters for important pathways in prostate cells that can result in cellular survival or apoptosis. While the activation of the ERK pathway appears to primarily result in survival, the roles of JNK and p38 are less clear. Manipulation of these pathways could have important implications for the treatment of prostate cancer and benign prostatic hypertrophy.

  13. Heparan sulfate proteoglycans: structure, protein interactions and cell signaling

    Directory of Open Access Journals (Sweden)

    Juliana L. Dreyfuss

    2009-09-01

    Full Text Available Heparan sulfate proteoglycans are ubiquitously found at the cell surface and extracellular matrix in all the animal species. This review will focus on the structural characteristics of the heparan sulfate proteoglycans related to protein interactions leading to cell signaling. The heparan sulfate chains due to their vast structural diversity are able to bind and interact with a wide variety of proteins, such as growth factors, chemokines, morphogens, extracellular matrix components, enzymes, among others. There is a specificity directing the interactions of heparan sulfates and target proteins, regarding both the fine structure of the polysaccharide chain as well precise protein motifs. Heparan sulfates play a role in cellular signaling either as receptor or co-receptor for different ligands, and the activation of downstream pathways is related to phosphorylation of different cytosolic proteins either directly or involving cytoskeleton interactions leading to gene regulation. The role of the heparan sulfate proteoglycans in cellular signaling and endocytic uptake pathways is also discussed.Proteoglicanos de heparam sulfato são encontrados tanto superfície celular quanto na matriz extracelular em todas as espécies animais. Esta revisão tem enfoque nas características estruturais dos proteoglicanos de heparam sulfato e nas interações destes proteoglicanos com proteínas que levam à sinalização celular. As cadeias de heparam sulfato, devido a sua variedade estrutural, são capazes de se ligar e interagir com ampla gama de proteínas, como fatores de crescimento, quimiocinas, morfógenos, componentes da matriz extracelular, enzimas, entreoutros. Existe uma especificidade estrutural que direciona as interações dos heparam sulfatos e proteínas alvo. Esta especificidade está relacionada com a estrutura da cadeia do polissacarídeo e os motivos conservados da cadeia polipeptídica das proteínas envolvidas nesta interação. Os heparam

  14. Cherry Valley ducks mitochondrial antiviral-signaling protein (MAVS mediated signaling pathway and antiviral activity research

    Directory of Open Access Journals (Sweden)

    Ning Li

    2016-09-01

    Full Text Available Mitochondrial antiviral-signaling protein (MAVS, an adaptor protein of retinoic acid-inducible gene I (RIG-I like receptors (RLRs-mediated signal pathway, is involved in innate immunity. In this study, Cherry Valley duck MAVS (duMAVS was cloned from the spleen and analyzed. duMAVS was determined to have a caspase activation and recruitment domain at N-terminal, followed by a proline rich domain and a transmembrane domain at C-terminal. Quantitative real time PCR indicated that duMAVS was expressed in all tissues tested across a broad expression spectrum. The expression of duMAVS was significantly up-regulated after infection with duck Tembusu virus. Overexpression of duMAVS could drive the activation of interferon-β, nuclear factor-κB, interferon regulatory factor 7, and many downstream factors (such as Mx, PKR, OAS, and IL-8 in duck embryo fibroblast cells. What’s more, RNA interference further confirmed that duMAVS was an important adaptor for IFN-β activation. The antiviral assay showed that duMAVS could suppress the various viral replications (duck Tembusu virus, novel reovirus, and duck plague virus at early stages of infection. Overall, these results showed that the main signal pathway mediated by duMAVS and it had a broad-spectrum antiviral ability. This research will be helpful to better understanding the innate immune system of ducks.

  15. A membrane protein/signaling protein interaction network for Arabidopsis version AMPv2.

    Science.gov (United States)

    Lalonde, Sylvie; Sero, Antoinette; Pratelli, Réjane; Pilot, Guillaume; Chen, Jin; Sardi, Maria I; Parsa, Saman A; Kim, Do-Young; Acharya, Biswa R; Stein, Erica V; Hu, Heng-Chen; Villiers, Florent; Takeda, Kouji; Yang, Yingzhen; Han, Yong S; Schwacke, Rainer; Chiang, William; Kato, Naohiro; Loqué, Dominique; Assmann, Sarah M; Kwak, June M; Schroeder, Julian I; Rhee, Seung Y; Frommer, Wolf B

    2010-01-01

    Interactions between membrane proteins and the soluble fraction are essential for signal transduction and for regulating nutrient transport. To gain insights into the membrane-based interactome, 3,852 open reading frames (ORFs) out of a target list of 8,383 representing membrane and signaling proteins from Arabidopsis thaliana were cloned into a Gateway-compatible vector. The mating-based split ubiquitin system was used to screen for potential protein-protein interactions (pPPIs) among 490 Arabidopsis ORFs. A binary robotic screen between 142 receptor-like kinases (RLKs), 72 transporters, 57 soluble protein kinases and phosphatases, 40 glycosyltransferases, 95 proteins of various functions, and 89 proteins with unknown function detected 387 out of 90,370 possible PPIs. A secondary screen confirmed 343 (of 386) pPPIs between 179 proteins, yielding a scale-free network (r(2) = 0.863). Eighty of 142 transmembrane RLKs tested positive, identifying 3 homomers, 63 heteromers, and 80 pPPIs with other proteins. Thirty-one out of 142 RLK interactors (including RLKs) had previously been found to be phosphorylated; thus interactors may be substrates for respective RLKs. None of the pPPIs described here had been reported in the major interactome databases, including potential interactors of G-protein-coupled receptors, phospholipase C, and AMT ammonium transporters. Two RLKs found as putative interactors of AMT1;1 were independently confirmed using a split luciferase assay in Arabidopsis protoplasts. These RLKs may be involved in ammonium-dependent phosphorylation of the C-terminus and regulation of ammonium uptake activity. The robotic screening method established here will enable a systematic analysis of membrane protein interactions in fungi, plants and metazoa.

  16. A membrane protein / signaling protein interaction network for Arabidopsis version AMPv2

    Directory of Open Access Journals (Sweden)

    Sylvie Lalonde

    2010-09-01

    Full Text Available Interactions between membrane proteins and the soluble fraction are essential for signal transduction and for regulating nutrient transport. To gain insights into the membrane-based interactome, 3,852 open reading frames (ORFs out of a target list of 8,383 representing membrane and signaling proteins from Arabidopsis thaliana were cloned into a Gateway compatible vector. The mating-based split-ubiquitin system was used to screen for potential protein-protein interactions (pPPIs among 490 Arabidopsis ORFs. A binary robotic screen between 142 receptor-like kinases, 72 transporters, 57 soluble protein kinases and phosphatases, 40 glycosyltransferases, 95 proteins of various functions and 89 proteins with unknown function detected 387 out of 90,370 possible PPIs. A secondary screen confirmed 343 (of 387 pPPIs between 179 proteins, yielding a scale-free network (r2=0.863. Eighty of 142 transmembrane receptor-like kinases (RLK tested positive, identifying three homomers, 63 heteromers and 80 pPPIs with other proteins. Thirty-one out of 142 RLK interactors (including RLKs had previously been found to be phosphorylated; thus interactors may be substrates for respective RLKs. None of the pPPIs described here had been reported in the major interactome databases, including potential interactors of G protein-coupled receptors, phospholipase C, and AMT ammonium transporters. Two RLKs found as putative interactors of AMT1;1 were independently confirmed using a split luciferase assay in Arabidopsis protoplasts. These RLKs may be involved in ammonium-dependent phosphorylation of the C-terminus and regulation of ammonium uptake activity. The robotic screening method established here will enable a systematic analysis of membrane protein interactions in fungi, plants and metazoa.

  17. Desired alteration of protein affinities: competitive selection of protein variants using yeast signal transduction machinery.

    Directory of Open Access Journals (Sweden)

    Misato Kaishima

    Full Text Available Molecules that can control protein-protein interactions (PPIs have recently drawn attention as new drug pipeline compounds. Here, we report a technique to screen desirable affinity-altered (affinity-enhanced and affinity-attenuated protein variants. We previously constructed a screening system based on a target protein fused to a mutated G-protein γ subunit (Gγcyto lacking membrane localization ability. This ability, required for signal transmission, is restored by recruiting Gγcyto into the membrane only when the target protein interacts with an artificially membrane-anchored candidate protein, thereby allowing interacting partners (Gγ recruitment system to be searched and identified. In the present study, the Gγ recruitment system was altered by integrating the cytosolic expression of a third protein as a competitor to set a desirable affinity threshold. This enabled the reliable selection of both affinity-enhanced and affinity-attenuated protein variants. The presented approach may facilitate the development of therapeutic proteins that allow the control of PPIs.

  18. R4 Regulator of G Protein Signaling (RGS) Proteins in Inflammation and Immunity.

    Science.gov (United States)

    Xie, Zhihui; Chan, Eunice C; Druey, Kirk M

    2016-03-01

    G protein-coupled receptors (GPCRs) have important functions in both innate and adaptive immunity, with the capacity to bridge interactions between the two arms of the host responses to pathogens through direct recognition of secreted microbial products or the by-products of host cells damaged by pathogen exposure. In the mid-1990s, a large group of intracellular proteins was discovered, the regulator of G protein signaling (RGS) family, whose main, but not exclusive, function appears to be to constrain the intensity and duration of GPCR signaling. The R4/B subfamily--the focus of this review--includes RGS1-5, 8, 13, 16, 18, and 21, which are the smallest RGS proteins in size, with the exception of RGS3. Prominent roles in the trafficking of B and T lymphocytes and macrophages have been described for RGS1, RGS13, and RGS16, while RGS18 appears to control platelet and osteoclast functions. Additional G protein independent functions of RGS13 have been uncovered in gene expression in B lymphocytes and mast cell-mediated allergic reactions. In this review, we discuss potential physiological roles of this RGS protein subfamily, primarily in leukocytes having central roles in immune and inflammatory responses. We also discuss approaches to target RGS proteins therapeutically, which represents a virtually untapped strategy to combat exaggerated immune responses leading to inflammation.

  19. The YvqE two-component system controls biofilm formation and acid production in Streptococcus pyogenes.

    Science.gov (United States)

    Isaka, Masanori; Tatsuno, Ichiro; Maeyama, Jun-Ichi; Matsui, Hideyuki; Zhang, Yan; Hasegawa, Tadao

    2016-07-01

    In Streptococcus pyogenes, proteins involved in determining virulence are controlled by stand-alone response regulators and by two-component regulatory systems. Previous studies reported that, compared to the parental strain, the yvqE sensor knockout strain showed significantly reduced growth and lower virulence. To determine the function of YvqE, we performed biofilm analysis and pH assays on yvqE mutants, and site-directed mutagenesis of YvqE. The yvqE deletion mutant showed a slower acid production rate, indicating that YvqE regulates acid production from sugar fermentation. The mutant strain, in which the Asp(26) residue in YvqE was replaced with Asn, affected biofilm formation, suggesting that this amino acid senses hydrogen ions produced by fermentative sugar metabolism. Signals received by YvqE were directly or indirectly responsible for inducing pilus expression. This study shows that at low environmental pH, biofilm formation in S. pyogenes is mediated by YvqE and suggests that regulation of pilus expression by environmental acidification could be directly under the control of YvqE.

  20. A two-component histidine kinase Shk1 controls stress response, sclerotial formation and fungicide resistance in Sclerotinia sclerotiorum.

    Science.gov (United States)

    Duan, Yabing; Ge, Changyan; Liu, Shengming; Wang, Jianxin; Zhou, Mingguo

    2013-09-01

    Fungal histidine kinases (HKs) are involved in osmotic and oxidative stress responses, hyphal development, fungicide sensitivity and virulence. Members of HK class III are known to signal through the high-osmolarity glycerol mitogen-activated protein kinase (HOG MAPK). In this study, we characterized the Shk1 gene (SS1G_12694.3), which encodes a putative class III HK, from the plant pathogen Sclerotinia sclerotiorum. Disruption of Shk1 resulted in resistance to phenylpyrrole and dicarboximide fungicides and increased sensitivity to hyperosmotic stress and H2 O2 -induced oxidative stress. The Shk1 mutant showed a significant reduction in vegetative hyphal growth and was unable to produce sclerotia. Quantitative real-time polymerase chain reaction (qRT-PCR and glycerol determination assays showed that the expression of SsHOG1 (the last kinase of the Hog pathway) and glycerol accumulation were regulated by the Shk1 gene, but PAK (p21-activated kinase) was not. In addition, the Shk1 mutant showed no change in virulence. All the defects were restored by genetic complementation of the Shk1 deletion mutant with the wild-type Shk1 gene. These findings indicate that Shk1 is involved in vegetative differentiation, sclerotial formation, glycerol accumulation and adaption to hyperosmotic and oxidative stresses, and to fungicides, in S. sclerotiorum. Taken together, our results demonstrate, for the first time, the role of two-component HKs in Sclerotinia.

  1. Engineering spatial gradients of signaling proteins using magnetic nanoparticles.

    Science.gov (United States)

    Bonnemay, L; Hostachy, S; Hoffmann, C; Gautier, J; Gueroui, Z

    2013-11-13

    Intracellular biochemical reactions are often localized in space and time, inducing gradients of enzymatic activity that may play decisive roles in determining cell's fate and functions. However, the techniques available to examine such enzymatic gradients of activity remain limited. Here, we propose a new method to engineer a spatial gradient of signaling protein concentration within Xenopus egg extracts using superparamagnetic nanoparticles. We show that, upon the application of a magnetic field, a concentration gradient of nanoparticles with a tunable length extension is established within confined egg extracts. We then conjugate the nanoparticles to RanGTP, a small G-protein controlling microtubule assembly. We found that the generation of an artificial gradient of Ran-nanoparticles modifies the spatial positioning of microtubule assemblies. Furthermore, the spatial control of the level of Ran concentration allows us to correlate the local fold increase in Ran-nanoparticle concentration with the spatial positioning of the microtubule-asters. Our assay provides a bottom-up approach to examine the minimum ingredients generating polarization and symmetry breaking within cells. More generally, these results show how magnetic nanoparticles and magnetogenetic tools can be used to control the spatiotemporal dynamics of signaling pathways.

  2. Computational Analysis of Signal Peptide-Dependent Secreted Proteins in Saccharomyces cerevisiae

    Institute of Scientific and Technical Information of China (English)

    2006-01-01

    Computer based software such as the SignalP v3.0, TargetP vl.01, big-PI predictor and TMHMM v2.0 were combined to predict the signal peptides, and the signal peptide-dependent secreted proteins among the 6 700 ORFs in genome of Saccharomyces cerevisiae. The results showed that 163 proteins were the secreted ones containing signal peptides, and they were secreted via Sec pathway. Among the 163 predicted secreted proteins, the signal peptides of 47 secreted proteins included only the H-domain and C-domain, without N-domain, but the signal peptides of other 116 secreted proteins included all the three domains. There were differences in the constitution of signal peptides between the secreted proteins of S. cerevisiae and of Candida albicans, but the length and amino acids types of their signal peptides were similar in general. Few of the same signal peptides occurred in the secreted proteins of S. cerevisiae genome, and the homology could be compared among the secreted proteins with the same signal peptides. The BLAST 2 SEQUENECES and CLUSTAL W were used to align the two protein sequences and multi-protein sequences, respectively. The alignment result indicated that homology of these sequences with the same signal peptide was very highly conservative in amino acid of complete gene. The effect of the signal peptides in S. cerevisia on expression of foreign eukaryotic secreted proteins is discussed in this paper.

  3. G protein modulation of recombinant P/Q-type calcium channels by regulators of G protein signalling proteins.

    Science.gov (United States)

    Mark, M D; Wittemann, S; Herlitze, S

    2000-10-01

    1. Fast synaptic transmission is triggered by the activation of presynaptic Ca2+ channels which can be inhibited by Gbetagamma subunits via G protein-coupled receptors (GPCR). Regulators of G protein signalling (RGS) proteins are GTPase-accelerating proteins (GAPs), which are responsible for >100-fold increases in the GTPase activity of G proteins and might be involved in the regulation of presynaptic Ca2+ channels. In this study we investigated the effects of RGS2 on G protein modulation of recombinant P/Q-type channels expressed in a human embryonic kidney (HEK293) cell line using whole-cell recordings. 2. RGS2 markedly accelerates transmitter-mediated inhibition and recovery from inhibition of Ba2+ currents (IBa) through P/Q-type channels heterologously expressed with the muscarinic acetylcholine receptor M2 (mAChR M2). 3. Both RGS2 and RGS4 modulate the prepulse facilitation properties of P/Q-type Ca2+ channels. G protein reinhibition is accelerated, while release from inhibition is slowed. These kinetics depend on the availability of G protein alpha and betagamma subunits which is altered by RGS proteins. 4. RGS proteins unmask the Ca2+ channel beta subunit modulation of Ca2+ channel G protein inhibition. In the presence of RGS2, P/Q-type channels containing the beta2a and beta3 subunits reveal significantly altered kinetics of G protein modulation and increased facilitation compared to Ca2+ channels coexpressed with the beta1b or beta4 subunit.

  4. Two-component system YvqEC-dependent bacterial resistance against vancomycin in Bacillus thuringiensis.

    Science.gov (United States)

    Zhang, Shumeng; Hu, Yimin; Fan, Qingyun; Wang, Xun; He, Jin

    2015-08-01

    YvqEC is one of the two-component signal transduction systems that may respond to cell envelope stress and enable cells to adjust multiple cellular functions. It consists of a histidine kinase YvqE and a response regulator YvqC. In this study, we separately constructed a single gene mutant ΔyvqE and a double gene mutant ΔyvqEC in Bacillus thuringiensis BMB171 through a homing endonucleases I-SceI mediated markerless gene deletion method. We found that the deletion of either yvqE or yvqEC weakened the resistance of B. thuringiensis against vancomycin. We also identified nine operons that may be involved in the cellular metabolism regulated by YvqC. This study not only enriches our understanding of bacterial resistance mechanisms against vancomycin, but also helps investigate the functions of YvqEC.

  5. Calcium binding proteins and calcium signaling in prokaryotes.

    Science.gov (United States)

    Domínguez, Delfina C; Guragain, Manita; Patrauchan, Marianna

    2015-03-01

    With the continued increase of genomic information and computational analyses during the recent years, the number of newly discovered calcium binding proteins (CaBPs) in prokaryotic organisms has increased dramatically. These proteins contain sequences that closely resemble a variety of eukaryotic calcium (Ca(2+)) binding motifs including the canonical and pseudo EF-hand motifs, Ca(2+)-binding β-roll, Greek key motif and a novel putative Ca(2+)-binding domain, called the Big domain. Prokaryotic CaBPs have been implicated in diverse cellular activities such as division, development, motility, homeostasis, stress response, secretion, transport, signaling and host-pathogen interactions. However, the majority of these proteins are hypothetical, and only few of them have been studied functionally. The finding of many diverse CaBPs in prokaryotic genomes opens an exciting area of research to explore and define the role of Ca(2+) in organisms other than eukaryotes. This review presents the most recent developments in the field of CaBPs and novel advancements in the role of Ca(2+) in prokaryotes.

  6. PINCH proteins regulate cardiac contractility by modulating integrin-linked kinase-protein kinase B signaling.

    Science.gov (United States)

    Meder, Benjamin; Huttner, Inken G; Sedaghat-Hamedani, Farbod; Just, Steffen; Dahme, Tillman; Frese, Karen S; Vogel, Britta; Köhler, Doreen; Kloos, Wanda; Rudloff, Jessica; Marquart, Sabine; Katus, Hugo A; Rottbauer, Wolfgang

    2011-08-01

    Integrin-linked kinase (ILK) is an essential component of the cardiac mechanical stretch sensor and is bound in a protein complex with parvin and PINCH proteins, the so-called ILK-PINCH-parvin (IPP) complex. We have recently shown that inactivation of ILK or β-parvin activity leads to heart failure in zebrafish via reduced protein kinase B (PKB/Akt) activation. Here, we show that PINCH proteins localize at sarcomeric Z disks and costameres in the zebrafish heart and skeletal muscle. To investigate the in vivo role of PINCH proteins for IPP complex stability and PKB signaling within the vertebrate heart, we inactivated PINCH1 and PINCH2 in zebrafish. Inactivation of either PINCH isoform independently leads to instability of ILK, loss of stretch-responsive anf and vegf expression, and progressive heart failure. The predominant cause of heart failure in PINCH morphants seems to be loss of PKB activity, since PKB phosphorylation at serine 473 is significantly reduced in PINCH-deficient hearts and overexpression of constitutively active PKB reconstitutes cardiac function in PINCH morphants. These findings highlight the essential function of PINCH proteins in controlling cardiac contractility by granting IPP/PKB-mediated signaling.

  7. Two overlapping two-component systems in Xanthomonas oryzae pv. oryzae contribute to full fitness in rice by regulating virulence factors expression

    Science.gov (United States)

    Zheng, Dehong; Yao, Xiaoyan; Duan, Meng; Luo, Yufeng; Liu, Biao; Qi, Pengyuan; Sun, Ming; Ruan, Lifang

    2016-01-01

    Two-component signal transduction systems (TCSs) are widely used by bacteria to adapt to the environment. In the present study, StoS (stress tolerance-related oxygen sensor) and SreKRS (salt response kinase, regulator, and sensor) were found to positively regulate extracellular polysaccharide (EPS) production and swarming in the rice pathogen Xanthomonas oryzae pv. oryzae (Xoo). Surprisingly, the absence of stoS or sreKRS did not attenuate virulence. To better understand the intrinsic functions of StoS and SreKRS, quantitative proteomics isobaric tags for relative and absolute quantitation (iTRAQ) was employed. Consistent with stoS and sreK mutants exhibiting a similar phenotype, the signalling circuits of StoS and SreKRS overlapped. Carbohydrate metabolism proteins and chemotaxis proteins, which could be responsible for EPS and swarming regulation, respectively, were reprogrammed in stoS and sreK mutants. Moreover, StoS and SreKRS demonstrated moderate expression of the major virulence factor, hypersensitive response and pathogenicity (Hrp) proteins through the HrpG-HrpX circuit. Most importantly, Xoo equipped with StoS and SreKRS outcompetes strains without StoS or SreKRS in co-infected rice and grows outside the host. Therefore, we propose that StoS and SreKRS adopt a novel strategy involving the moderation of Hrp protein expression and the promotion of EPS and motility to adapt to the environment. PMID:26957113

  8. G Protein-Coupled Receptor Signaling in Stem Cells and Cancer

    OpenAIRE

    Lynch, Jennifer R.; Jenny Yingzi Wang

    2016-01-01

    G protein-coupled receptors (GPCRs) are a large superfamily of cell-surface signaling proteins that bind extracellular ligands and transduce signals into cells via heterotrimeric G proteins. GPCRs are highly tractable drug targets. Aberrant expression of GPCRs and G proteins has been observed in various cancers and their importance in cancer stem cells has begun to be appreciated. We have recently reported essential roles for G protein-coupled receptor 84 (GPR84) and G protein subunit Gαq in ...

  9. Reconstruction of Protein-Protein Interaction Network of Insulin Signaling in Homo Sapiens

    Directory of Open Access Journals (Sweden)

    Saliha Durmuş Tekir

    2010-01-01

    Full Text Available Diabetes is one of the most prevalent diseases in the world. Type 1 diabetes is characterized by the failure of synthesizing and secreting of insulin because of destroyed pancreatic β-cells. Type 2 diabetes, on the other hand, is described by the decreased synthesis and secretion of insulin because of the defect in pancreatic β-cells as well as by the failure of responding to insulin because of malfunctioning of insulin signaling. In order to understand the signaling mechanisms of responding to insulin, it is necessary to identify all components in the insulin signaling network. Here, an interaction network consisting of proteins that have statistically high probability of being biologically related to insulin signaling in Homo sapiens was reconstructed by integrating Gene Ontology (GO annotations and interactome data. Furthermore, within this reconstructed network, interacting proteins which mediate the signal from insulin hormone to glucose transportation were identified using linear paths. The identification of key components functioning in insulin action on glucose metabolism is crucial for the efforts of preventing and treating type 2 diabetes mellitus.

  10. Activated protein synthesis and suppressed protein breakdown signaling in skeletal muscle of critically ill patients

    DEFF Research Database (Denmark)

    Jespersen, Jakob G; Nedergaard, Anders; Reitelseder, Søren

    2011-01-01

    involved in muscle mass regulation, we investigated the phosphorylation and expression of key factors in these protein synthesis and breakdown signaling pathways in thigh skeletal muscle of critically ill intensive care unit (ICU) patients compared with healthy controls.......Skeletal muscle mass is controlled by myostatin and Akt-dependent signaling on mammalian target of rapamycin (mTOR), glycogen synthase kinase 3β (GSK3β) and forkhead box O (FoxO) pathways, but it is unknown how these pathways are regulated in critically ill human muscle. To describe factors...

  11. Activated protein synthesis and suppressed protein breakdown signaling in skeletal muscle of critically ill patients

    DEFF Research Database (Denmark)

    Jespersen, Jakob G; Nedergaard, Anders; Reitelseder, Søren

    2011-01-01

    involved in muscle mass regulation, we investigated the phosphorylation and expression of key factors in these protein synthesis and breakdown signaling pathways in thigh skeletal muscle of critically ill intensive care unit (ICU) patients compared with healthy controls.......Skeletal muscle mass is controlled by myostatin and Akt-dependent signaling on mammalian target of rapamycin (mTOR), glycogen synthase kinase 3ß (GSK3ß) and forkhead box O (FoxO) pathways, but it is unknown how these pathways are regulated in critically ill human muscle. To describe factors...

  12. SpaK/SpaR two-component system characterized by a structure-driven domain-fusion method and in vitro phosphorylation studies.

    Directory of Open Access Journals (Sweden)

    Anu Chakicherla

    2009-06-01

    Full Text Available Here we introduce a quantitative structure-driven computational domain-fusion method, which we used to predict the structures of proteins believed to be involved in regulation of the subtilin pathway in Bacillus subtilis, and used to predict a protein-protein complex formed by interaction between the proteins. Homology modeling of SpaK and SpaR yielded preliminary structural models based on a best template for SpaK comprising a dimer of a histidine kinase, and for SpaR a response regulator protein. Our LGA code was used to identify multi-domain proteins with structure homology to both modeled structures, yielding a set of domain-fusion templates then used to model a hypothetical SpaK/SpaR complex. The models were used to identify putative functional residues and residues at the protein-protein interface, and bioinformatics was used to compare functionally and structurally relevant residues in corresponding positions among proteins with structural homology to the templates. Models of the complex were evaluated in light of known properties of the functional residues within two-component systems involving His-Asp phosphorelays. Based on this analysis, a phosphotransferase complexed with a beryllofluoride was selected as the optimal template for modeling a SpaK/SpaR complex conformation. In vitro phosphorylation studies performed using wild type and site-directed SpaK mutant proteins validated the predictions derived from application of the structure-driven domain-fusion method: SpaK was phosphorylated in the presence of (32P-ATP and the phosphate moiety was subsequently transferred to SpaR, supporting the hypothesis that SpaK and SpaR function as sensor and response regulator, respectively, in a two-component signal transduction system, and furthermore suggesting that the structure-driven domain-fusion approach correctly predicted a physical interaction between SpaK and SpaR. Our domain-fusion algorithm leverages quantitative structure information

  13. The sae locus of Staphylococcus aureus encodes a two-component regulatory system.

    Science.gov (United States)

    Giraudo, A T; Calzolari, A; Cataldi, A A; Bogni, C; Nagel, R

    1999-08-01

    Sae is a regulatory locus that activates the production of several exoproteins in Staphylococcus aureus. A 3.4-kb fragment of a S. aureus genomic library, screened with a probe adjacent to the transposon insertion of a sae::Tn551 mutant, was cloned into a bifunctional vector. This fragment was shown to carry the sae locus by restoration of exoprotein production in sae mutants. The sae locus was mapped to the SmaI-D fragment of the staphylococcal chromosome by pulse-field electrophoresis. Sequence analysis of the cloned fragment revealed the presence of two genes, designated saeR and saeS, encoding a response regulator and a histidine protein kinase, respectively, with high homology to other bacterial two-component regulatory systems.

  14. Phenotype MicroArray Analysis of Escherichia coli K-12 Mutants with Deletions of All Two-Component Systems

    Science.gov (United States)

    Zhou, Lu; Lei, Xiang-He; Bochner, Barry R.; Wanner, Barry L.

    2003-01-01

    Two-component systems are the most common mechanism of transmembrane signal transduction in bacteria. A typical system consists of a histidine kinase and a partner response regulator. The histidine kinase senses an environmental signal, which it transmits to its partner response regulator via a series of autophosphorylation, phosphotransfer, and dephosphorylation reactions. Much work has been done on particular systems, including several systems with regulatory roles in cellular physiology, communication, development, and, in the case of bacterial pathogens, the expression of genes important for virulence. We used two methods to investigate two-component regulatory systems in Escherichia coli K-12. First, we systematically constructed mutants with deletions of all two-component systems by using a now-standard technique of gene disruption (K. A. Datsenko and B. L. Wanner, Proc. Natl. Acad. Sci. USA 97:6640-6645, 2000). We then analyzed these deletion mutants with a new technology called Phenotype MicroArrays, which permits assays of nearly 2,000 growth phenotypes simultaneously. In this study we tested 100 mutants, including mutants with individual deletions of all two-component systems and several related genes, including creBC-regulated genes (cbrA and cbrBC), phoBR-regulated genes (phoA, phoH, phnCDEFGHIJKLMNOP, psiE, and ugpBAECQ), csgD, luxS, and rpoS. The results of this battery of nearly 200,000 tests provided a wealth of new information concerning many of these systems. Of 37 different two-component mutants, 22 showed altered phenotypes. Many phenotypes were expected, and several new phenotypes were also revealed. The results are discussed in terms of the biological roles and other information concerning these systems, including DNA microarray data for a large number of the same mutants. Other mutational effects are also discussed. PMID:12897016

  15. Mechanistic pathways and biological roles for receptor-independent activators of G-protein signaling.

    Science.gov (United States)

    Blumer, Joe B; Smrcka, Alan V; Lanier, Stephen M

    2007-03-01

    Signal processing via heterotrimeric G-proteins in response to cell surface receptors is a central and much investigated aspect of how cells integrate cellular stimuli to produce coordinated biological responses. The system is a target of numerous therapeutic agents and plays an important role in adaptive processes of organs; aberrant processing of signals through these transducing systems is a component of various disease states. In addition to G-protein coupled receptor (GPCR)-mediated activation of G-protein signaling, nature has evolved creative ways to manipulate and utilize the Galphabetagamma heterotrimer or Galpha and Gbetagamma subunits independent of the cell surface receptor stimuli. In such situations, the G-protein subunits (Galpha and Gbetagamma) may actually be complexed with alternative binding partners independent of the typical heterotrimeric Galphabetagamma. Such regulatory accessory proteins include the family of regulator of G-protein signaling (RGS) proteins that accelerate the GTPase activity of Galpha and various entities that influence nucleotide binding properties and/or subunit interaction. The latter group of proteins includes receptor-independent activators of G-protein signaling (AGS) proteins that play surprising roles in signal processing. This review provides an overview of our current knowledge regarding AGS proteins. AGS proteins are indicative of a growing number of accessory proteins that influence signal propagation, facilitate cross talk between various types of signaling pathways, and provide a platform for diverse functions of both the heterotrimeric Galphabetagamma and the individual Galpha and Gbetagamma subunits.

  16. Dynamics of coupled simplest chaotic two-component electronic circuits and its potential application to random bit generation

    Energy Technology Data Exchange (ETDEWEB)

    Modeste Nguimdo, Romain, E-mail: Romain.Nguimdo@vub.ac.be [Applied Physics Research Group, APHY, Vrije Universiteit Brussel, Pleinlaan 2, B-1050 Brussel (Belgium); Tchitnga, Robert [Laboratory of Electronics, Automation and Signal Processing, Department of Physics, University of Dschang, P.O. Box 67, Dschang (Cameroon); Woafo, Paul [Laboratory of Modelling and Simulation in Engineering and Biological Physics, Faculty of Science, University of Yaoundé I, P.O. Box 812, Yaoundé (Cameroon)

    2013-12-15

    We numerically investigate the possibility of using a coupling to increase the complexity in simplest chaotic two-component electronic circuits operating at high frequency. We subsequently show that complex behaviors generated in such coupled systems, together with the post-processing are suitable for generating bit-streams which pass all the NIST tests for randomness. The electronic circuit is built up by unidirectionally coupling three two-component (one active and one passive) oscillators in a ring configuration through resistances. It turns out that, with such a coupling, high chaotic signals can be obtained. By extracting points at fixed interval of 10 ns (corresponding to a bit rate of 100 Mb/s) on such chaotic signals, each point being simultaneously converted in 16-bits (or 8-bits), we find that the binary sequence constructed by including the 10(or 2) least significant bits pass statistical tests of randomness, meaning that bit-streams with random properties can be achieved with an overall bit rate up to 10×100 Mb/s =1Gbit/s (or 2×100 Mb/s =200 Megabit/s). Moreover, by varying the bias voltages, we also investigate the parameter range for which more complex signals can be obtained. Besides being simple to implement, the two-component electronic circuit setup is very cheap as compared to optical and electro-optical systems.

  17. Signal peptide hydrophobicity is critical for early stages in protein export by Bacillus subtilis

    NARCIS (Netherlands)

    Zanen, G; Meima, R; Tjalsma, H; Jongbloed, JDH; Westers, H; Oudega, B; Luirink, J; van Dijl, JM; Quax, WJ; Houben, E.N.G.

    2005-01-01

    Signal peptides that direct protein export in Bacillus subtilis are overall more hydrophobic than signal peptides in Escherichia coli. To study the importance of signal peptide hydrophobicity for protein export in both organisms, the alpha-amylase AmyQ was provided with leucine-rich (high hydrophobi

  18. Caenorhabditis elegans SMA-10/LRIG is a conserved transmembrane protein that enhances bone morphogenetic protein signaling.

    Directory of Open Access Journals (Sweden)

    Tina L Gumienny

    2010-05-01

    Full Text Available Bone morphogenetic protein (BMP pathways control an array of developmental and homeostatic events, and must themselves be exquisitely controlled. Here, we identify Caenorhabditis elegans SMA-10 as a positive extracellular regulator of BMP-like receptor signaling. SMA-10 acts genetically in a BMP-like (Sma/Mab pathway between the ligand DBL-1 and its receptors SMA-6 and DAF-4. We cloned sma-10 and show that it has fifteen leucine-rich repeats and three immunoglobulin-like domains, hallmarks of an LRIG subfamily of transmembrane proteins. SMA-10 is required in the hypodermis, where the core Sma/Mab signaling components function. We demonstrate functional conservation of LRIGs by rescuing sma-10(lf animals with the Drosophila ortholog lambik, showing that SMA-10 physically binds the DBL-1 receptors SMA-6 and DAF-4 and enhances signaling in vitro. This interaction is evolutionarily conserved, evidenced by LRIG1 binding to vertebrate receptors. We propose a new role for LRIG family members: the positive regulation of BMP signaling by binding both Type I and Type II receptors.

  19. Caenorhabditis elegans SMA-10/LRIG Is a Conserved Transmembrane Protein that Enhances Bone Morphogenetic Protein Signaling

    Science.gov (United States)

    Gumienny, Tina L.; MacNeil, Lesley; Zimmerman, Cole M.; Wang, Huang; Chin, Lena; Wrana, Jeffrey L.; Padgett, Richard W.

    2010-01-01

    Bone morphogenetic protein (BMP) pathways control an array of developmental and homeostatic events, and must themselves be exquisitely controlled. Here, we identify Caenorhabditis elegans SMA-10 as a positive extracellular regulator of BMP–like receptor signaling. SMA-10 acts genetically in a BMP–like (Sma/Mab) pathway between the ligand DBL-1 and its receptors SMA-6 and DAF-4. We cloned sma-10 and show that it has fifteen leucine-rich repeats and three immunoglobulin-like domains, hallmarks of an LRIG subfamily of transmembrane proteins. SMA-10 is required in the hypodermis, where the core Sma/Mab signaling components function. We demonstrate functional conservation of LRIGs by rescuing sma-10(lf) animals with the Drosophila ortholog lambik, showing that SMA-10 physically binds the DBL-1 receptors SMA-6 and DAF-4 and enhances signaling in vitro. This interaction is evolutionarily conserved, evidenced by LRIG1 binding to vertebrate receptors. We propose a new role for LRIG family members: the positive regulation of BMP signaling by binding both Type I and Type II receptors. PMID:20502686

  20. Competing G protein-coupled receptor kinases balance G protein and β-arrestin signaling.

    Science.gov (United States)

    Heitzler, Domitille; Durand, Guillaume; Gallay, Nathalie; Rizk, Aurélien; Ahn, Seungkirl; Kim, Jihee; Violin, Jonathan D; Dupuy, Laurence; Gauthier, Christophe; Piketty, Vincent; Crépieux, Pascale; Poupon, Anne; Clément, Frédérique; Fages, François; Lefkowitz, Robert J; Reiter, Eric

    2012-06-26

    Seven-transmembrane receptors (7TMRs) are involved in nearly all aspects of chemical communications and represent major drug targets. 7TMRs transmit their signals not only via heterotrimeric G proteins but also through β-arrestins, whose recruitment to the activated receptor is regulated by G protein-coupled receptor kinases (GRKs). In this paper, we combined experimental approaches with computational modeling to decipher the molecular mechanisms as well as the hidden dynamics governing extracellular signal-regulated kinase (ERK) activation by the angiotensin II type 1A receptor (AT(1A)R) in human embryonic kidney (HEK)293 cells. We built an abstracted ordinary differential equations (ODE)-based model that captured the available knowledge and experimental data. We inferred the unknown parameters by simultaneously fitting experimental data generated in both control and perturbed conditions. We demonstrate that, in addition to its well-established function in the desensitization of G-protein activation, GRK2 exerts a strong negative effect on β-arrestin-dependent signaling through its competition with GRK5 and 6 for receptor phosphorylation. Importantly, we experimentally confirmed the validity of this novel GRK2-dependent mechanism in both primary vascular smooth muscle cells naturally expressing the AT(1A)R, and HEK293 cells expressing other 7TMRs.

  1. Two-component signal transduction in Bacillus cereus and closely related bacteria

    NARCIS (Netherlands)

    Been, de M.W.H.J.

    2009-01-01

    Bacillus cereus is a Gram-positive pathogen that is recognised as an important cause of food-borne disease worldwide. Within the genus Bacillus, B. cereus and its closest relatives form a homogeneous subdivision that has been termed the B. cereus group. This group includes B. anthracis, a pathogen

  2. HBV X protein interacts with cytoskeletal signaling proteins through SH3 binding.

    Science.gov (United States)

    Feng, Huixing; Tan, Tuan Lin; Niu, Dandan; Chen, Wei Ning

    2010-01-01

    The aim of this study was to investigate interactions between cellular SH3-containing proteins and the proline-rich domain in Hepatitis B Virus (HBV) X protein (HBx) The proline-rich domain of HBx (amino acids 19-58) as well as the relevant site-directed mutagenesis (proline to alanine residues) were cloned into pGEX-5X-1 and expressed as GST-PXXP and GST-AXXA probes. Panomics SH3 domain arrays were probed using both GST-PXXP and GST-AXXA to identify potential interacting SH3 domain containing proteins. The specific interactions were confirmed by the immunoprecipitation of the full-length SH3 domain-containing protein. We report here the binding assay which demonstrated interaction between PXXP domain in HBx and the SH3-domain containing proteins, in particular various signaling proteins involved in cytoskeletal reorganization. Our findings were consistent with similar virus-host interactions via SH3 binding for other viruses such as hepatitis C virus (HCV) and human immunodeficiency virus (HIV) Further characterization of the proline-rich binding to SH3 domains could yield important information for the design of novel therapeutic measures against downstream disease causative effects of HBx in the liver cells.

  3. Initial data problems for the two-component Camassa-Holm system

    Directory of Open Access Journals (Sweden)

    Xiaohuan Wang

    2014-06-01

    Full Text Available This article concerns the study of some properties of the two-component Camassa-Holm system. By constructing two sequences of solutions of the two-component Camassa-Holm system, we prove that the solution map of the Cauchy problem of the two-component Camassa-Holm system is not uniformly continuous in $H^s(\\mathbb{R}$, $s>5/2$.

  4. The Cbln family of proteins interact with multiple signaling pathways.

    Science.gov (United States)

    Wei, Peng; Pattarini, Roberto; Rong, Yongqi; Guo, Hong; Bansal, Parmil K; Kusnoor, Sheila V; Deutch, Ariel Y; Parris, Jennifer; Morgan, James I

    2012-06-01

    Cerebellin precursor protein (Cbln1) is essential for synapse integrity in cerebellum through assembly into complexes that bridge pre-synaptic β-neurexins (Nrxn) to post-synaptic GluRδ2. However, GluRδ2 is largely cerebellum-specific, yet Cbln1 and its little studied family members, Cbln2 and Cbln4, are expressed throughout brain. Therefore, we investigated whether additional proteins mediate Cbln family actions. Whereas Cbln1 and Cbln2 bound to GluRδ2 and Nrxns1-3, Cbln4 bound weakly or not at all, suggesting it has distinct binding partners. In a candidate receptor-screening assay, Cbln4 (but not Cbln1 or Cbln2) bound selectively to the netrin receptor, (deleted in colorectal cancer (DCC) in a netrin-displaceable fashion. To determine whether Cbln4 had a netrin-like function, Cbln4-null mice were generated. Cbln4-null mice did not phenocopy netrin-null mice. Cbln1 and Cbln4 were likely co-localized in neurons thought to be responsible for synaptic changes in striatum of Cbln1-null mice. Furthermore, complexes containing Cbln1 and Cbln4 had greatly reduced affinity to DCC but increased affinity to Nrxns, suggesting a functional interaction. However, Cbln4-null mice lacked the striatal synaptic changes seen in Cbln null mice. Thus, Cbln family members interact with multiple receptors/signaling pathways in a subunit composition-dependent manner and have independent functions with Cbln4 potentially involved in the less well-characterized role of netrin/DCC in adult brain.

  5. Analytical method for yrast line states in the interacting two-component Bose-Einstein condensate

    Institute of Scientific and Technical Information of China (English)

    解炳昊; 景辉

    2002-01-01

    The yrast spectrum for the harmonically trapped two-component Bose-Einstein condensate (BEC), omitting thedifference between the two components, has been studied using an analytical method. The energy eigenstates andeigenvalues for L=0,1,2,3 are given. We illustrate that there are different eigenstate behaviours between the even Land odd L cases for the two-component BEC in two dimensions. Except for symmetric states, there are antisymmetricstates for the permutation of the two components, which cannot reduce to those in a single condensate case when thevalue of L is odd.

  6. Signal recognition particle and SecA cooperate during export of secretory proteins with highly hydrophobic signal sequences.

    Directory of Open Access Journals (Sweden)

    Yufan Zhou

    Full Text Available The Sec translocon of bacterial plasma membranes mediates the linear translocation of secretory proteins as well as the lateral integration of membrane proteins. Integration of many membrane proteins occurs co-translationally via the signal recognition particle (SRP-dependent targeting of ribosome-associated nascent chains to the Sec translocon. In contrast, translocation of classical secretory proteins across the Sec translocon is a post-translational event requiring no SRP but the motor protein SecA. Secretory proteins were, however, reported to utilize SRP in addition to SecA, if the hydrophobicity of their signal sequences exceeds a certain threshold value. Here we have analyzed transport of this subgroup of secretory proteins across the Sec translocon employing an entirely defined in vitro system. We thus found SecA to be both necessary and sufficient for translocation of secretory proteins with hydrophobic signal sequences, whereas SRP and its receptor improved translocation efficiency. This SRP-mediated boost of translocation is likely due to the early capture of the hydrophobic signal sequence by SRP as revealed by site-specific photo cross-linking of ribosome nascent chain complexes.

  7. Shc adaptor proteins are key transducers of mitogenic signaling mediated by the G protein-coupled thrombin receptor

    DEFF Research Database (Denmark)

    Chen, Y; Grall, D; Salcini, A E

    1996-01-01

    The serine protease thrombin activates G protein signaling systems that lead to Ras activation and, in certain cells, proliferation. Whereas the steps leading to Ras activation by G protein-coupled receptors are not well defined, the mechanisms of Ras activation by receptor tyrosine kinases have...... kinase activation, gene induction and cell growth. From these data, we conclude that Shc represents a crucial point of convergence between signaling pathways activated by receptor tyrosine kinases and G protein-coupled receptors....

  8. Protein tyrosine kinase and mitogen-activated protein kinase signalling pathways contribute to differences in heterophil-mediated innate immune responsiveness between two lines of broilers

    Science.gov (United States)

    Protein tyrosine phosphorylation mediates signal transduction of cellular processes, with protein tyrosine kinases (PTKs) regulating virtually all signaling events. The mitogen-activated protein kinase (MAPK) super-family consists of three conserved pathways that convert receptor activation into ce...

  9. The ankyrin repeat protein Diversin recruits Casein kinase Iepsilon to the beta-catenin degradation complex and acts in both canonical Wnt and Wnt/JNK signaling.

    Science.gov (United States)

    Schwarz-Romond, Thomas; Asbrand, Christian; Bakkers, Jeroen; Kühl, Michael; Schaeffer, Hans-Joerg; Huelsken, Jörg; Behrens, Jürgen; Hammerschmidt, Matthias; Birchmeier, Walter

    2002-08-15

    Wnt signals control decisive steps in development and can induce the formation of tumors. Canonical Wnt signals control the formation of the embryonic axis, and are mediated by stabilization and interaction of beta-catenin with Lef/Tcf transcription factors. An alternative branch of the Wnt pathway uses JNK to establish planar cell polarity in Drosophila and gastrulation movements in vertebrates. We describe here the vertebrate protein Diversin that interacts with two components of the canonical Wnt pathway, Casein kinase Iepsilon (CKIepsilon) and Axin/Conductin. Diversin recruits CKIepsilon to the beta-catenin degradation complex that consists of Axin/Conductin and GSK3beta and allows efficient phosphorylation of beta-catenin, thereby inhibiting beta-catenin/Tcf signals. Morpholino-based gene ablation in zebrafish shows that Diversin is crucial for axis formation, which depends on beta-catenin signaling. Diversin is also involved in JNK activation and gastrulation movements in zebrafish. Diversin is distantly related to Diego of Drosophila, which functions only in the pathway that controls planar cell polarity. Our data show that Diversin is an essential component of the Wnt-signaling pathway and acts as a molecular switch, which suppresses Wnt signals mediated by the canonical beta-catenin pathway and stimulates signaling via JNK.

  10. Quantitative Kinetic Analyses of Shutting Off a Two-Component System

    Directory of Open Access Journals (Sweden)

    Rong Gao

    2017-05-01

    Full Text Available Cells rely on accurate control of signaling systems to adapt to environmental perturbations. System deactivation upon stimulus removal is as important as activation of signaling pathways. The two-component system (TCS is one of the major bacterial signaling schemes. In many TCSs, phosphatase activity of the histidine kinase (HK is believed to play an essential role in shutting off the pathway and resetting the system to the prestimulus state. Two basic challenges are to understand the dynamic behavior of system deactivation and to quantitatively evaluate the role of phosphatase activity under natural cellular conditions. Here we report a kinetic analysis of the response to shutting off the archetype Escherichia coli PhoR-PhoB TCS pathway using both transcription reporter assays and in vivo phosphorylation analyses. Upon removal of the stimulus, the pathway is shut off by rapid dephosphorylation of the PhoB response regulator (RR while PhoB-regulated gene products gradually reset to prestimulus levels through growth dilution. We developed an approach combining experimentation and modeling to assess in vivo kinetic parameters of the phosphatase activity with kinetic data from multiple phosphatase-diminished mutants. This enabled an estimation of the PhoR phosphatase activity in vivo, which is much stronger than the phosphatase activity of PhoR cytoplasmic domains analyzed in vitro. We quantitatively modeled how strong the phosphatase activity needs to be to suppress nonspecific phosphorylation in TCSs and discovered that strong phosphatase activity of PhoR is required for cross-phosphorylation suppression.

  11. Role of the SaeRS two-component regulatory system in Staphylococcus epidermidis autolysis and biofilm formation

    Directory of Open Access Journals (Sweden)

    Francois Patrice

    2011-06-01

    Full Text Available Abstract Background Staphylococcus epidermidis (SE has emerged as one of the most important causes of nosocomial infections. The SaeRS two-component signal transduction system (TCS influences virulence and biofilm formation in Staphylococcus aureus. The deletion of saeR in S. epidermidis results in impaired anaerobic growth and decreased nitrate utilization. However, the regulatory function of SaeRS on biofilm formation and autolysis in S. epidermidis remains unclear. Results The saeRS genes of SE1457 were deleted by homologous recombination. The saeRS deletion mutant, SE1457ΔsaeRS, exhibited increased biofilm formation that was disturbed more severely (a 4-fold reduction by DNase I treatment compared to SE1457 and the complementation strain SE1457saec. Compared to SE1457 and SE1457saec, SE1457ΔsaeRS showed increased Triton X-100-induced autolysis (approximately 3-fold and decreased cell viability in planktonic/biofilm states; further, SE1457ΔsaeRS also released more extracellular DNA (eDNA in the biofilms. Correlated with the increased autolysis phenotype, the transcription of autolysis-related genes, such as atlE and aae, was increased in SE1457ΔsaeRS. Whereas the expression of accumulation-associated protein was up-regulated by 1.8-fold in 1457ΔsaeRS, the expression of an N-acetylglucosaminyl transferase enzyme (encoded by icaA critical for polysaccharide intercellular adhesin (PIA synthesis was not affected by the deletion of saeRS. Conclusions Deletion of saeRS in S. epidermidis resulted in an increase in biofilm-forming ability, which was associated with increased eDNA release and up-regulated Aap expression. The increased eDNA release from SE1457ΔsaeRS was associated with increased bacterial autolysis and decreased bacterial cell viability in the planktonic/biofilm states.

  12. Information theory in systems biology. Part II: protein-protein interaction and signaling networks.

    Science.gov (United States)

    Mousavian, Zaynab; Díaz, José; Masoudi-Nejad, Ali

    2016-03-01

    By the development of information theory in 1948 by Claude Shannon to address the problems in the field of data storage and data communication over (noisy) communication channel, it has been successfully applied in many other research areas such as bioinformatics and systems biology. In this manuscript, we attempt to review some of the existing literatures in systems biology, which are using the information theory measures in their calculations. As we have reviewed most of the existing information-theoretic methods in gene regulatory and metabolic networks in the first part of the review, so in the second part of our study, the application of information theory in other types of biological networks including protein-protein interaction and signaling networks will be surveyed. Copyright © 2015 Elsevier Ltd. All rights reserved.

  13. Two-Component Super AKNS Equations and Their Finite-Dimensional Integrable Super Hamiltonian System

    OpenAIRE

    Jing Yu; Jingwei Han

    2014-01-01

    Starting from a matrix Lie superalgebra, two-component super AKNS system is constructed. By making use of monononlinearization technique of Lax pairs, we find that the obtained two-component super AKNS system is a finite-dimensional integrable super Hamiltonian system. And its Lax representation and $r$ -matrix are also given in this paper.

  14. TASI 2011 lectures notes: two-component fermion notation and supersymmetry

    OpenAIRE

    Martin, Stephen P.

    2012-01-01

    These notes, based on work with Herbi Dreiner and Howie Haber, discuss how to do practical calculations of cross sections and decay rates using two-component fermion notation, as appropriate for supersymmetry and other beyond-the-Standard-Model theories. Included are a list of two-component fermion Feynman rules for the Minimal Supersymmetric Standard Model, and some example calculations.

  15. Relativistic two-component jet evolutions in 2D and 3D

    NARCIS (Netherlands)

    Meliani, Z.; Keppens, R.

    2009-01-01

    Observations of astrophysical jets and theoretical arguments suggest a transverse stratification with two components induced by intrinsic features of the central engine (accretion disk + black hole). We study two-component jet dynamics for an inner fast low density jet, surrounded by a slower, dense

  16. Two-Component Super AKNS Equations and Their Finite-Dimensional Integrable Super Hamiltonian System

    Directory of Open Access Journals (Sweden)

    Jing Yu

    2014-01-01

    Full Text Available Starting from a matrix Lie superalgebra, two-component super AKNS system is constructed. By making use of monononlinearization technique of Lax pairs, we find that the obtained two-component super AKNS system is a finite-dimensional integrable super Hamiltonian system. And its Lax representation and r-matrix are also given in this paper.

  17. Harmine promotes osteoblast differentiation through bone morphogenetic protein signaling

    Energy Technology Data Exchange (ETDEWEB)

    Yonezawa, Takayuki [Department of Nutriproteomics, Graduate School of Medicine, The University of Tokyo, 7-3-1 Hongo, Bunkyo-ku, Tokyo 113-0033 (Japan); Research Institute for Biological Functions, Chubu University, 1200 Matsumoto, Kasugai, Aichi 487-8501 (Japan); Lee, Ji-Won [Research Institute for Biological Functions, Chubu University, 1200 Matsumoto, Kasugai, Aichi 487-8501 (Japan); Hibino, Ayaka; Asai, Midori [Department of Biological Chemistry, College of Bioscience and Biotechnology, Chubu University, 1200 Matsumoto, Kasugai, Aichi 487-8501 (Japan); Hojo, Hironori [Center for Disease Biology and Integrative Medicine, Faculty of Medicine, The University of Tokyo, 7-3-1 Hongo, Bunkyo-ku, Tokyo 113-0033 (Japan); Cha, Byung-Yoon [Research Institute for Biological Functions, Chubu University, 1200 Matsumoto, Kasugai, Aichi 487-8501 (Japan); Teruya, Toshiaki [Research Institute for Biological Functions, Chubu University, 1200 Matsumoto, Kasugai, Aichi 487-8501 (Japan); Faculty of Education, University of the Ryukyus, 1 Senbaru, Nishihara, Okinawa 903-0213 (Japan); Nagai, Kazuo [Research Institute for Biological Functions, Chubu University, 1200 Matsumoto, Kasugai, Aichi 487-8501 (Japan); Department of Biological Chemistry, College of Bioscience and Biotechnology, Chubu University, 1200 Matsumoto, Kasugai, Aichi 487-8501 (Japan); Chung, Ung-Il [Center for Disease Biology and Integrative Medicine, Faculty of Medicine, The University of Tokyo, 7-3-1 Hongo, Bunkyo-ku, Tokyo 113-0033 (Japan); Yagasaki, Kazumi [Department of Nutriproteomics, Graduate School of Medicine, The University of Tokyo, 7-3-1 Hongo, Bunkyo-ku, Tokyo 113-0033 (Japan); Division of Applied Biological Chemistry, Institute of Agriculture, Tokyo Noko University, 3-5-8 Saiwai, Fuchu, Tokyo 183-8509 (Japan); and others

    2011-06-03

    Highlights: {yields} Harmine promotes the activity and mRNA expression of ALP. {yields} Harmine enhances the expressions of osteocalcin mRNA and protein. {yields} Harmine induces osteoblastic mineralization. {yields} Harmine upregulates the mRNA expressions of BMPs, Runx2 and Osterix. {yields} BMP signaling pathways are involved in the actions of harmine. -- Abstract: Bone mass is regulated by osteoblast-mediated bone formation and osteoclast-mediated bone resorption. We previously reported that harmine, a {beta}-carboline alkaloid, inhibits osteoclast differentiation and bone resorption in vitro and in vivo. In this study, we investigated the effects of harmine on osteoblast proliferation, differentiation and mineralization. Harmine promoted alkaline phosphatase (ALP) activity in MC3T3-E1 cells without affecting their proliferation. Harmine also increased the mRNA expressions of the osteoblast marker genes ALP and Osteocalcin. Furthermore, the mineralization of MC3T3-E1 cells was enhanced by treatment with harmine. Harmine also induced osteoblast differentiation in primary calvarial osteoblasts and mesenchymal stem cell line C3H10T1/2 cells. Structure-activity relationship studies using harmine-related {beta}-carboline alkaloids revealed that the C3-C4 double bond and 7-hydroxy or 7-methoxy group of harmine were important for its osteogenic activity. The bone morphogenetic protein (BMP) antagonist noggin and its receptor kinase inhibitors dorsomorphin and LDN-193189 attenuated harmine-promoted ALP activity. In addition, harmine increased the mRNA expressions of Bmp-2, Bmp-4, Bmp-6, Bmp-7 and its target gene Id1. Harmine also enhanced the mRNA expressions of Runx2 and Osterix, which are key transcription factors in osteoblast differentiation. Furthermore, BMP-responsive and Runx2-responsive reporters were activated by harmine treatment. Taken together, these results indicate that harmine enhances osteoblast differentiation probably by inducing the expressions of

  18. The role of the CGRP-receptor component protein (RCP) in adrenomedullin receptor signal transduction.

    Science.gov (United States)

    Prado, M A; Evans-Bain, B; Oliver, K R; Dickerson, I M

    2001-11-01

    G protein-coupled receptors are usually thought to act as monomer receptors that bind ligand and then interact with G proteins to initiate signal transduction. In this study we report an intracellular peripheral membrane protein named the calcitonin gene-related peptide (CGRP)-receptor component protein (RCP) required for signal transduction at the G protein-coupled receptor for adrenomedullin. Cell lines were made that expressed an antisense construct of the RCP cDNA, and in these cells diminished RCP expression correlated with loss of adrenomedullin signal transduction. In contrast, loss of RCP did not diminish receptor density or affinity, therefore RCP does not appear to act as a chaperone protein. Instead, RCP represents a novel class of protein required to couple the adrenomedullin receptor to the cellular signal transduction pathway. A candidate adrenomedullin receptor named the calcitonin receptor-like receptor (CRLR) has been described, which forms high affinity adrenomedullin receptors when co-expressed with the accessory protein receptor-activity modifying protein 2 (RAMP2). RCP co-immunoprecipitated with CRLR and RAMP2, indicating that a functional adrenomedullin receptor is composed of at least three proteins: the ligand binding protein (CRLR), an accessory protein (RAMP2), and a coupling protein for signal transduction (RCP).

  19. Predicting Pharmacodynamic Drug-Drug Interactions through Signaling Propagation Interference on Protein-Protein Interaction Networks.

    Directory of Open Access Journals (Sweden)

    Kyunghyun Park

    Full Text Available As pharmacodynamic drug-drug interactions (PD DDIs could lead to severe adverse effects in patients, it is important to identify potential PD DDIs in drug development. The signaling starting from drug targets is propagated through protein-protein interaction (PPI networks. PD DDIs could occur by close interference on the same targets or within the same pathways as well as distant interference through cross-talking pathways. However, most of the previous approaches have considered only close interference by measuring distances between drug targets or comparing target neighbors. We have applied a random walk with restart algorithm to simulate signaling propagation from drug targets in order to capture the possibility of their distant interference. Cross validation with DrugBank and Kyoto Encyclopedia of Genes and Genomes DRUG shows that the proposed method outperforms the previous methods significantly. We also provide a web service with which PD DDIs for drug pairs can be analyzed at http://biosoft.kaist.ac.kr/targetrw.

  20. A proteomic analysis of the regulon of the NarP two-component regulatory system response regulator in the bovine pathogen Mannheimia haemolytica A1

    Directory of Open Access Journals (Sweden)

    Inamoto Ichiro

    2011-11-01

    Full Text Available Abstract Background The response of the NarQP two-component signal transduction system regulon in response to the presence of nitrate for the bovine pathogen Mannheimia haemolytica A1 was investigated by proteomic analysis. Total proteins from a narP mutant and the parent SH1217 grown with or without NaNO3 supplement were examined by ISO-DALT 2D electrophoresis and liquid chromatography-mass spectrometry. Results Seventeen proteins were differentially expressed in the parent strain SH1217 in response to the addition of NaNO3 to the growth media. These responses were absent in the narP mutant, indicating that the altered production of these proteins is mediated by NarPMh. Interestingly, NarPMh mediated the increased production of some proteins which are not generally associated with nitrate respiration, such as the iron transporters FbpA and YfeA. The increased production of proteins such as superoxide dismutase, SodA, and GAPDH were also observed. The increased production of these iron-regulated proteins by NarPMh is thought to enhance the swift establishment of the nitrate respiration mechanism of M. haemolytica during pathogenesis. Conclusion The data suggested NarPMh acts as an important regulator which regulates the expression of a small set of proteins in response to nitrate availability. This may contribute to the prevalence of M. haemolytica A1 in its host during pathogenesis of BPP, through enhancing the effectiveness of nitrate respiration either directly or indirectly.

  1. Transcriptome analysis of the Brucella abortus BvrR/BvrS two-component regulatory system.

    Directory of Open Access Journals (Sweden)

    Cristina Viadas

    Full Text Available BACKGROUND: The two-component BvrR/BvrS system is essential for Brucella abortus virulence. It was shown previously that its dysfunction alters the expression of some major outer membrane proteins and the pattern of lipid A acylation. To determine the genes regulated by BvrR/BvrS, we performed a whole-genome microarray analysis using B. abortus RNA obtained from wild type and bvrR mutant cells grown in the same conditions. METHODOLOGY/PRINCIPAL FINDINGS: A total of 127 differentially expressed genes were found: 83 were over expressed and 44 were less expressed in the bvrR mutant. Two operons, the phosphotransferase system and the maltose transport system, were down-regulated. Several genes involved in cell envelope or outer membrane biogenesis were differentially expressed: genes for outer membrane proteins (omp25a, omp25d, lipoproteins, LPS and fatty acid biosynthesis, stress response proteins, chaperones, flagellar genes, and twelve genes encoding ABC transport systems. Ten genes related with carbon metabolism (pckA and fumB among others were up-regulated in the bvrR mutant, and denitrification genes (nirK, norC and nosZ were also regulated. Notably, seven transcriptional regulators were affected, including VjbR, ExoR and OmpR that were less expressed in the bvrR mutant. Finally, the expression of eleven genes which have been previously related with Brucella virulence was also altered. CONCLUSIONS/SIGNIFICANCE: All these data corroborate the impact of BvrR/BvrS on cell envelope modulation, confirm that this system controls the carbon and nitrogen metabolism, and suggest a cross-talk among some regulators to adjust the Brucella physiology to the shift expected to occur during the transit from the extracellular to the intracellular niche.

  2. Signal peptide peptidase (SPP) assembles with substrates and misfolded membrane proteins into distinct oligomeric complexes

    Science.gov (United States)

    Schrul, Bianca; Kapp, Katja; Sinning, Irmgard; Dobberstein, Bernhard

    2010-01-01

    SPP (signal peptide peptidase) is an aspartyl intramembrane cleaving protease, which processes a subset of signal peptides, and is linked to the quality control of ER (endoplasmic reticulum) membrane proteins. We analysed SPP interactions with signal peptides and other membrane proteins by co-immunoprecipitation assays. We found that SPP interacts specifically and tightly with a large range of newly synthesized membrane proteins, including signal peptides, preproteins and misfolded membrane proteins, but not with all co-expressed type II membrane proteins. Signal peptides are trapped by the catalytically inactive SPP mutant SPPD/A. Preproteins and misfolded membrane proteins interact with both SPP and the SPPD/A mutant, and are not substrates for SPP-mediated intramembrane proteolysis. Proteins interacting with SPP are found in distinct complexes of different sizes. A signal peptide is mainly trapped in a 200 kDa SPP complex, whereas a preprotein is predominantly found in a 600 kDa SPP complex. A misfolded membrane protein is detected in 200, 400 and 600 kDa SPP complexes. We conclude that SPP not only processes signal peptides, but also collects preproteins and misfolded membrane proteins that are destined for disposal. PMID:20196774

  3. The Role of Cgrp-Receptor Component Protein (Rcp in Cgrp-Mediated Signal Transduction

    Directory of Open Access Journals (Sweden)

    M. A. Prado

    2001-01-01

    Full Text Available The calcitonin gene-related peptide (CGRP-receptor component protein (RCP is a 17-kDa intracellular peripheral membrane protein required for signal transduction at CGRP receptors. To determine the role of RCP in CGRP-mediated signal transduction, RCP was depleted from NIH3T3 cells using antisense strategy. Loss of RCP protein correlated with loss of cAMP production by CGRP in the antisense cells. In contrast, loss of RCP had no effect on CGRP-mediated binding; therefore RCP is not acting as a chaperone for the CGRP receptor. Instead, RCP is a novel signal transduction molecule that couples the CGRP receptor to the cellular signal transduction machinery. RCP thus represents a prototype for a new class of signal transduction proteins that are required for regulation of G protein-coupled receptors.

  4. Prostaglandin E2 negatively regulates AMP-activated protein kinase via protein kinase A signaling pathway.

    Science.gov (United States)

    Funahashi, Koji; Cao, Xia; Yamauchi, Masako; Kozaki, Yasuko; Ishiguro, Naoki; Kambe, Fukushi

    2009-01-01

    We investigated possible involvement of prostaglandin (PG) E2 in regulation of AMP-activated protein kinase (AMPK). When osteoblastic MG63 cells were cultured in serum-deprived media, Thr-172 phosphorylation of AMPK alpha-subunit was markedly increased. Treatment of the cells with PGE2 significantly reduced the phosphorylation. Ser-79 phosphorylation of acetyl-CoA carboxylase, a direct target for AMPK, was also reduced by PGE2. On the other hand, PGE2 reciprocally increased Ser-485 phosphorylation of the alpha-subunit that could be associated with inhibition of AMPK activity. These effects of PGE2 were mimicked by PGE2 receptor EP2 and EP4 agonists and forskolin, but not by EP1 and EP3 agonists, and the effects were suppressed by an adenylate cyclase inhibitor SQ22536 and a protein kinase A inhibitor H89. Additionally, the PGE2 effects were duplicated in primary calvarial osteoblasts. Together, the present study demonstrates that PGE2 negatively regulates AMPK activity via activation of protein kinase A signaling pathway.

  5. Preparation and frictional investigation of the two-components silanes deposited on alumina surface

    Energy Technology Data Exchange (ETDEWEB)

    Kośla, K.; Grobelny, J.; Cichomski, M., E-mail: mcichom@uni.lodz.pl

    2014-09-30

    Highlights: • The two-component silane films on the alumina surface were obtained by a combination of soft lithography and vapor phase deposition method. • The effectiveness of modification procedure was monitored by AFM topography images. • By using gas phase deposition method succeeded in obtaining a good reproduction of pattern. • Silane films with low surface free energy and coefficient of friction values were obtained. • The frictional performance in milli-Newton load range of one- and two-component films was investigated by microtribometry. - Abstract: Functionalization and pattering technique that permits two-component pattern-specific modification of alumina surface with silanes molecules are reported. The method relies on a two-component molecular system that simultaneously decreases coefficient of friction of the alumina surface and provides uniform chemical functionality suitable for further elaboration. Pattern/two-component modification is achieved via gas-phase deposition of the silanes using polydimethylsiloxane stamp. The frictional behaviors of the two-component films of the silane molecules with different chain length covalently absorbed on alumina surfaces, were characterized by the ball-disk (microtribometer) tester. The surfaces of the substrate modified by two-component molecular films were examined by atomic force microscopy (AFM). The measured tribological results showed that the mixing of the fluoroalkylsilane and alkylsilane enhance the lubrication and decrease the friction compared to the one-component thin films.

  6. Modulation of the MAP kinase signaling cascade by Raf kinase inhibitory protein

    Institute of Scientific and Technical Information of China (English)

    Nicholas TRAKUL; Marsha R. ROSNER

    2005-01-01

    Proteins like Raf kinase inhibitory protein (RKIP) that serve as modulators of signaling pathways, either by promoting or inhibiting the formation of productive signaling complexes through protein-protein interactions, have been demonstrated to play an increasingly important role in a number of cell types and organisms. These proteins have been implicated in development as well as the progression of cancer. RKIP is a particularly interesting regulator, as it is a highly conserved, ubiquitously expressed protein that has been shown to play a role in growth and differentiation in a number of organisms and can regulate multiple signaling pathways. RKIP is also the first MAP kinase signaling modulator to be identified as playing a role in cancer metastasis, and identification of the mechanism by which it regulates Raf-1 activation provides new targets for therapeutic intervention.

  7. In silico characterization of three two-component systems of Ehrlichia canis and evaluation of a natural plant-derived inhibitor.

    Science.gov (United States)

    Santos, E V; Silva, G; Cardozo, G P; Bitencourt, T A; França, S C; Fachin, A L; Marins, M

    2012-10-04

    Two-component signal transduction systems (TCS) are important elements in the interaction of endobacteria with host cells. They are basically composed of two proteins, an environmental signal sensor and a response regulator, which activate genes involved in a wide range of bacterial responses to their environment. We analyzed three sets of genes corresponding to TCS of Ehrlichia canis, a common tick-borne canine pathogen and the etiologic agent of canine monocytic ehrlichiosis, in order to identify the characteristic domains of the sensor and response regulator components. Analysis of sequence alignments of the corresponding proteins indicated a high degree of similarity to other members of the Anaplasmataceae TCS proteins, demonstrating that they could be useful as universal targets for development of new drugs against these bacteria. We also evaluated by quantitative PCR inhibition of E. canis by (2H)-1,4-benzoxazin-3(4H)-one (BOA), the core compound of the plant phenolic compound DIMBOA, which shows inhibitory action against TCS of the phytopathogen Agrobacterium tumefasciens. This bacterium exerts its pathogenicity by transferring oncogenic DNA (T-DNA) into plant cells; this transfer is mediated through a type-IV secretion system, which is regulated by the VirA/VirG TCS. The process of infection and pathogenesis of E. canis is associated with the secretion of effector proteins into the host cell cytoplasm through a T4SS system, which blocks the cell defense response. We suggest that BOA, and possibly other plant phenolic compounds that are TCS inhibitors, can be exploited in the search for new antiehrlichial drugs to be used alone or as complements in the treatment of canine monocytic ehrlichiosis.

  8. Methods of producing epoxides from alkenes using a two-component catalyst system

    Science.gov (United States)

    Kung, Mayfair C.; Kung, Harold H.; Jiang, Jian

    2013-07-09

    Methods for the epoxidation of alkenes are provided. The methods include the steps of exposing the alkene to a two-component catalyst system in an aqueous solution in the presence of carbon monoxide and molecular oxygen under conditions in which the alkene is epoxidized. The two-component catalyst system comprises a first catalyst that generates peroxides or peroxy intermediates during oxidation of CO with molecular oxygen and a second catalyst that catalyzes the epoxidation of the alkene using the peroxides or peroxy intermediates. A catalyst system composed of particles of suspended gold and titanium silicalite is one example of a suitable two-component catalyst system.

  9. Two-component generalizations of the periodic Camassa-Holm and Degasperis-Procesi equations

    CERN Document Server

    Escher, Joachim; Lenells, Jonatan

    2010-01-01

    We use geometric methods to study two natural two-component generalizations of the periodic Camassa-Holm and Degasperis-Procesi equations. We show that these generalizations can be regarded as geodesic equations on the semidirect product of the diffeomorphism group of the circle $\\Diff(S^1)$ with some space of sufficiently smooth functions on the circle. Our goals are to understand the geometric properties of these two-component systems and to prove local well-posedness in various function spaces. Furthermore, we perform some explicit curvature calculations for the two-component Camassa-Holm equation, giving explicit examples of large subspaces of positive curvature.

  10. Travelling wave solutions for some two-component shallow water models

    Science.gov (United States)

    Dutykh, Denys; Ionescu-Kruse, Delia

    2016-07-01

    In the present study we perform a unified analysis of travelling wave solutions to three different two-component systems which appear in shallow water theory. Namely, we analyze the celebrated Green-Naghdi equations, the integrable two-component Camassa-Holm equations and a new two-component system of Green-Naghdi type. In particular, we are interested in solitary and cnoidal-type solutions, as two most important classes of travelling waves that we encounter in applications. We provide a complete phase-plane analysis of all possible travelling wave solutions which may arise in these models. In particular, we show the existence of new type of solutions.

  11. Two-Component Wadati-Konno-Ichikawa Equation and Its Symmetry Reductions

    Institute of Scientific and Technical Information of China (English)

    QU Chang-Zheng; YAO Ruo-Xia; LI Zhi-Bin

    2004-01-01

    @@ It is shown that two-component Wadati-Konno-Ichikawa (WKI) equation, i.e. a generalization of the well-known WKI equation, is obtained from the motion of space curves in Euclidean geometry, and it is exactly a system for the graph of the curves when the curve motion is governed by the two-component modified Korteweg-de Vries flow. Group-invariant solutions of the two-component WKI equation which corresponds to an optimal system of its Lie point symmetry groups are obtained, and its similarity reductions to systems of ordinary differential equations are also given.

  12. Protein conservation and variation suggest mechanisms of cell type-specific modulation of signaling pathways.

    Directory of Open Access Journals (Sweden)

    Martin H Schaefer

    2014-06-01

    Full Text Available Many proteins and signaling pathways are present in most cell types and tissues and yet perform specialized functions. To elucidate mechanisms by which these ubiquitous pathways are modulated, we overlaid information about cross-cell line protein abundance and variability, and evolutionary conservation onto functional pathway components and topological layers in the pathway hierarchy. We found that the input (receptors and the output (transcription factors layers evolve more rapidly than proteins in the intermediary transmission layer. In contrast, protein expression variability decreases from the input to the output layer. We observed that the differences in protein variability between the input and transmission layer can be attributed to both the network position and the tendency of variable proteins to physically interact with constitutively expressed proteins. Differences in protein expression variability and conservation are also accompanied by the tendency of conserved and constitutively expressed proteins to acquire somatic mutations, while germline mutations tend to occur in cell type-specific proteins. Thus, conserved core proteins in the transmission layer could perform a fundamental role in most cell types and are therefore less tolerant to germline mutations. In summary, we propose that the core signal transmission machinery is largely modulated by a variable input layer through physical protein interactions. We hypothesize that the bow-tie organization of cellular signaling on the level of protein abundance variability contributes to the specificity of the signal response in different cell types.

  13. Identification, Functional Characterization and Regulon Prediction of a Novel Two Component System Comprising BAS0540-BAS0541 of Bacillus anthracis

    Science.gov (United States)

    Gopalani, Monisha; Kandari, Divya; Bhatnagar, Rakesh

    2016-01-01

    Two component systems (TCSs) can be envisaged as complex molecular devices that help the bacteria to sense its environment and respond aptly. 41 TCSs are predicted in Bacillus anthracis, a potential bioterrorism agent, of which only four have been studied so far. Thus, the intricate signaling network contributed by TCSs remains largely unmapped in B. anthracis and needs comprehensive exploration. In this study, we functionally characterized one such system composed of BAS0540 (Response regulator) and BAS0541 (Histidine kinase). BAS0540-BAS0541, the closest homolog of CiaRH of Streptococcus in B. anthracis, forms a functional TCS with BAS0541 displaying autophosphorylation and subsequent phosphotransfer to BAS0540. BAS0540 was also found to accept phosphate from physiologically relevant small molecule phosphodonors like acetyl phosphate and carbamoyl phosphate. Results of qRT-PCR and immunoblotting demonstrated that BAS0540 exhibits a constitutive expression throughout the growth of B. anthracis. Regulon prediction for BAS0540 in B. anthracis was done in silico using the consensus DNA binding sequence of CiaR of Streptococcus. The predicted regulon of BAS0540 comprised of 23 genes, which could be classified into 8 functionally diverse categories. None of the proven virulence factors were a part of the predicted regulon, an observation contrasting with the regulon of CiaRH in Streptococci. Electrophoretic mobility shift assay was used to show direct binding of purified BAS0540 to the upstream regions of 5 putative regulon candidates- BAS0540 gene itself; a gene predicted to encode cell division protein FtsA; a self–immunity gene; a RND family transporter gene and a gene encoding stress (heat) responsive protein. A significant enhancement in the DNA binding ability of BAS0540 was observed upon phosphorylation. Overexpression of response regulator BAS0540 in B. anthracis led to a prodigious increase of ~6 folds in the cell length, thereby conferring it a filamentous

  14. A mechanism regulating G protein-coupled receptor signaling that requires cycles of protein palmitoylation and depalmitoylation.

    Science.gov (United States)

    Jia, Lixia; Chisari, Mariangela; Maktabi, Mohammad H; Sobieski, Courtney; Zhou, Hao; Konopko, Aaron M; Martin, Brent R; Mennerick, Steven J; Blumer, Kendall J

    2014-02-28

    Reversible attachment and removal of palmitate or other long-chain fatty acids on proteins has been hypothesized, like phosphorylation, to control diverse biological processes. Indeed, palmitate turnover regulates Ras trafficking and signaling. Beyond this example, however, the functions of palmitate turnover on specific proteins remain poorly understood. Here, we show that a mechanism regulating G protein-coupled receptor signaling in neuronal cells requires palmitate turnover. We used hexadecyl fluorophosphonate or palmostatin B to inhibit enzymes in the serine hydrolase family that depalmitoylate proteins, and we studied R7 regulator of G protein signaling (RGS)-binding protein (R7BP), a palmitoylated allosteric modulator of R7 RGS proteins that accelerate deactivation of Gi/o class G proteins. Depalmitoylation inhibition caused R7BP to redistribute from the plasma membrane to endomembrane compartments, dissociated R7BP-bound R7 RGS complexes from Gi/o-gated G protein-regulated inwardly rectifying K(+) (GIRK) channels and delayed GIRK channel closure. In contrast, targeting R7BP to the plasma membrane with a polybasic domain and an irreversibly attached lipid instead of palmitate rendered GIRK channel closure insensitive to depalmitoylation inhibitors. Palmitate turnover therefore is required for localizing R7BP to the plasma membrane and facilitating Gi/o deactivation by R7 RGS proteins on GIRK channels. Our findings broaden the scope of biological processes regulated by palmitate turnover on specific target proteins. Inhibiting R7BP depalmitoylation may provide a means of enhancing GIRK activity in neurological disorders.

  15. Signals fly when kinases meet Rho-of-plants (ROP) small G-proteins.

    Science.gov (United States)

    Fehér, Attila; Lajkó, Dézi Bianka

    2015-08-01

    Rho-type small GTP-binding plant proteins function as two-state molecular switches in cellular signalling. There is accumulating evidence that Rho-of-plants (ROP) signalling is positively controlled by plant receptor kinases, through the ROP guanine nucleotide exchange factor proteins. These signalling modules regulate cell polarity, cell shape, hormone responses, and pathogen defence, among other things. Other ROP-regulatory proteins might also be subjected to protein phosphorylation by cellular kinases (e.g., mitogen-activated protein kinases or calcium-dependent protein kinases), in order to integrate various cellular signalling pathways with ROP GTPase-dependent processes. In contrast to the role of kinases in upstream ROP regulation, much less is known about the potential link between ROP GTPases and downstream kinase signalling. In other eukaryotes, Rho-type G-protein-activated kinases are widespread and have a key role in many cellular processes. Recent data indicate the existence of structurally different ROP-activated kinases in plants, but their ROP-dependent biological functions still need to be validated. In addition to these direct interactions, ROPs may also indirectly control the activity of mitogen-activated protein kinases or calcium-dependent protein kinases. These kinases may therefore function as upstream as well as downstream kinases in ROP-mediated signalling pathways, such as the phosphatidylinositol monophosphate kinases involved in cell polarity establishment.

  16. Histaminylation of glutamine residues is a novel posttranslational modification implicated in G-protein signaling.

    Science.gov (United States)

    Vowinckel, Jakob; Stahlberg, Silke; Paulmann, Nils; Bluemlein, Katharina; Grohmann, Maik; Ralser, Markus; Walther, Diego J

    2012-11-02

    Posttranslational modifications (PTM) have been shown to be essential for protein function and signaling. Here we report the identification of a novel modification, protein transfer of histamine, and provide evidence for its function in G protein signaling. Histamine, known as neurotransmitter and mediator of the inflammatory response, was found incorporated into mastocytoma proteins. Histaminylation was dependent on transglutaminase II. Mass spectrometry confirmed histamine modification of the small and heterotrimeric G proteins Cdc42, Gαo1 and Gαq. The modification was specific for glutamine residues in the catalytic core, and triggered their constitutive activation. TGM2-mediated histaminylation is thus a novel PTM that functions in G protein signaling. Protein αmonoaminylations, thus including histaminylation, serotonylation, dopaminylation and norepinephrinylation, hence emerge as a novel class of regulatory PTMs.

  17. Signaling via G proteins mediates tumorigenic effects of GPR87

    DEFF Research Database (Denmark)

    Arfelt, Kristine Niss; Fares, Suzan; Sparre-Ulrich, Alexander H.;

    2017-01-01

    G protein-coupled receptors (GPCRs) constitute a large protein family of seven transmembrane (7TM) spanning proteins that regulate multiple physiological functions. GPR87 is overexpressed in several cancers and plays a role in tumor cell survival. Here, the basal activity of GPR87 was investigated...... in transiently transfected HEK293 cells, revealing ligand-independent coupling to Gαi, Gαq and Gα12/13. Furthermore, GPR87 showed a ligand-independent G protein-dependent activation of the downstream transcription factors CREB, NFκB, NFAT and SRE. In tetracycline-induced Flp-In T-Rex-293 cells, GPR87 induced...

  18. Insulin Signalling and Regulation of Protein Kinase B in Adipocytes

    OpenAIRE

    Göransson, Olga

    2003-01-01

    Insulin resistance is a hallmark of type 2 diabetes, an increasingly common disorder. The cause of insulin resistance is supposedly failures in the processes used by insulin to signal to the interior of its target cells. These failing steps are still unknown, most probably because of incomplete knowledge of how the insulin signals are transmitted. Since insulin resistance is strongly linked to obesity, defects in lipid metabolism or other adipocyte functions, may be an important factor in the...

  19. Scale and Contour: Two Components of a Theory of Memory for Melodies.

    Science.gov (United States)

    Dowling, W. Jay

    1978-01-01

    The author concentrates on two components of memory which contribute to the reproduction and recognition of melodies, namely, melodic contour and musical scale. A new experiment is reported that shows the interdependence of both components. (Author/RK)

  20. Laser controlling chaotic region of a two-component Bose-Einstein condensate

    Institute of Scientific and Technical Information of China (English)

    Boli Xia; Wenhua Hai

    2005-01-01

    @@ For a weakly and periodically driven two-component Bose-Einstein condensate (BEC) the Melnikov chaotic solution and boundedness conditions are derived from a direct perturbation theory that leads to the chaotic regions in the parameter space.

  1. Two component injection moulding: an interface quality and bond strength dilemma

    DEFF Research Database (Denmark)

    Islam, Mohammad Aminul; Hansen, Hans Nørgaard; Tang, Peter Torben

    2008-01-01

    Two component injection moulding is a special branch of injection moulding where two different polymers are combined in to a single part to exploit the different material properties in the final product. Considering the technical and economical importance of the process, this paper investigates...... on quality parameters of the two component parts. Most engineering applications of two component injection moulding calls for high bond strength between the two polymers, on the other hand a sharp and well-defined interface between the two polymers are required for applications like selective metallization...... conditions for a sharp and well-defined interface are exactly the opposite of what is congenial for higher bond strength. So in the production of two component injection moulded parts, there is a compromise to make between the interface quality and the bond strength of the two polymers. Also the injection...

  2. Photoisomerization-induced morphology and transparency transition in an azobenzene based two-component organogel system.

    Science.gov (United States)

    Cao, Xinhua; Liu, Xue; Chen, Liming; Mao, Yueyuan; Lan, Haichuang; Yi, Tao

    2015-11-15

    A two-component gel containing long chain alkylated gallic acid (GA) and photochromic phenazopyridine (PAP) was prepared. The gel was thoroughly characterized by UV-visible and IR spectra, SEM and POM images, XRD diffraction and dynamic oscillatory measurements. The structure and transparency of the two-component gel can be reversibly changed by alternative UV light irradiation and warming in the palm of the hand. This kind of soft material has potential application in upscale surface functional materials.

  3. Competitive Adsorption of a Two-Component Gas on a Deformable Adsorbent

    OpenAIRE

    Usenko, A. S.

    2013-01-01

    We investigate the competitive adsorption of a two-component gas on the surface of an adsorbent whose adsorption properties vary in adsorption due to the adsorbent deformation. The essential difference of adsorption isotherms for a deformable adsorbent both from the classical Langmuir adsorption isotherms of a two-component gas and from the adsorption isotherms of a one-component gas taking into account variations in adsorption properties of the adsorbent in adsorption is obtained. We establi...

  4. Variational derivation of two-component Camassa-Holm shallow water system

    CERN Document Server

    Ionescu-Kruse, Delia

    2012-01-01

    By a variational approach in the Lagrangian formalism, we derive the nonlinear integrable two-component Camassa-Holm system (1). We show that the two-component Camassa-Holm system (1) with the plus sign arises as an approximation to the Euler equations of hydrodynamics for propagation of irrotational shallow water waves over a flat bed. The Lagrangian used in the variational derivation is not a metric.

  5. Conformational analysis of g protein-coupled receptor signaling by hydrogen/deuterium exchange mass spectrometry.

    Science.gov (United States)

    Li, Sheng; Lee, Su Youn; Chung, Ka Young

    2015-01-01

    Conformational change and protein-protein interactions are two major mechanisms of membrane protein signal transduction, including G protein-coupled receptors (GPCRs). Upon agonist binding, GPCRs change conformation, resulting in interaction with downstream signaling molecules such as G proteins. To understand the precise signaling mechanism, studies have investigated the structural mechanism of GPCR signaling using X-ray crystallography, nuclear magnetic resonance (NMR), or electron paramagnetic resonance. In addition to these techniques, hydrogen/deuterium exchange mass spectrometry (HDX-MS) has recently been used in GPCR studies. HDX-MS measures the rate at which peptide amide hydrogens exchange with deuterium in the solvent. Exposed or flexible regions have higher exchange rates and excluded or ordered regions have lower exchange rates. Therefore, HDX-MS is a useful tool for studying protein-protein interfaces and conformational changes after protein activation or protein-protein interactions. Although HDX-MS does not give high-resolution structures, it analyzes protein conformations that are difficult to study with X-ray crystallography or NMR. Furthermore, conformational information from HDX-MS can help in the crystallization of X-ray crystallography by suggesting highly flexible regions. Interactions between GPCRs and downstream signaling molecules are not easily analyzed by X-ray crystallography or NMR because of the large size of the GPCR-signaling molecule complexes, hydrophobicity, and flexibility of GPCRs. HDX-MS could be useful for analyzing the conformational mechanism of GPCR signaling. In this chapter, we discuss details of HDX-MS for analyzing GPCRs using the β2AR-G protein complex as a model system.

  6. Making Sense of G Proteins: Genetic analysis of sensory G protein signaling in the nematode C. elegans

    NARCIS (Netherlands)

    H. Lans (Hannes)

    2005-01-01

    textabstractAmong the key molecules involved in sensory perception are G proteins, which act in every cell to activate a cascade of signaling molecules in response to certain environmental cues. In this thesis, several studies on the role of G proteins in the sensory system of C. elegans are describ

  7. Compartmentalization Role of A-Kinase Anchoring Proteins (AKAPs in Mediating Protein Kinase A (PKA Signaling and Cardiomyocyte Hypertrophy

    Directory of Open Access Journals (Sweden)

    Abeer Rababa'h

    2014-12-01

    Full Text Available The Beta-adrenergic receptors (β-ARs stimulation enhances contractility through protein kinase-A (PKA substrate phosphorylation. This PKA signaling is conferred in part by PKA binding to A-kinase anchoring proteins (AKAPs. AKAPs coordinate multi-protein signaling networks that are targeted to specific intracellular locations, resulting in the localization of enzyme activity and transmitting intracellular actions of neurotransmitters and hormones to its target substrates. In particular, mAKAP (muscle-selective AKAP has been shown to be present on the nuclear envelope of cardiomyocytes with various proteins including: PKA-regulatory subunit (RIIα, phosphodiesterase-4D3, protein phosphatase-2A, and ryanodine receptor (RyR2. Therefore, through the coordination of spatial-temporal signaling of proteins and enzymes, mAKAP controls cyclic-adenosine monophosphate (cAMP levels very tightly and functions as a regulator of PKA-mediated substrate phosphorylation leading to changes in calcium availability and myofilament calcium sensitivity. The goal of this review is to elucidate the critical compartmentalization role of mAKAP in mediating PKA signaling and regulating cardiomyocyte hypertrophy by acting as a scaffolding protein. Based on our literature search and studying the structure–function relationship between AKAP scaffolding protein and its binding partners, we propose possible explanations for the mechanism by which mAKAP promotes cardiac hypertrophy.

  8. Making Sense of G Proteins: Genetic analysis of sensory G protein signaling in the nematode C. elegans

    NARCIS (Netherlands)

    H. Lans (Hannes)

    2005-01-01

    textabstractAmong the key molecules involved in sensory perception are G proteins, which act in every cell to activate a cascade of signaling molecules in response to certain environmental cues. In this thesis, several studies on the role of G proteins in the sensory system of C. elegans are describ

  9. Methods for the Analysis of Protein Phosphorylation-Mediated Cellular Signaling Networks

    Science.gov (United States)

    White, Forest M.; Wolf-Yadlin, Alejandro

    2016-06-01

    Protein phosphorylation-mediated cellular signaling networks regulate almost all aspects of cell biology, including the responses to cellular stimulation and environmental alterations. These networks are highly complex and comprise hundreds of proteins and potentially thousands of phosphorylation sites. Multiple analytical methods have been developed over the past several decades to identify proteins and protein phosphorylation sites regulating cellular signaling, and to quantify the dynamic response of these sites to different cellular stimulation. Here we provide an overview of these methods, including the fundamental principles governing each method, their relative strengths and weaknesses, and some examples of how each method has been applied to the analysis of complex signaling networks. When applied correctly, each of these techniques can provide insight into the topology, dynamics, and regulation of protein phosphorylation signaling networks.

  10. Role of Rab GTPases and their interacting proteins in mediating metabolic signalling and regulation.

    Science.gov (United States)

    Chua, Christelle En Lin; Tang, Bor Luen

    2015-06-01

    The vesicular transport pathways, which shuttle materials to and from the cell surface and within the cell, and the metabolic (growth factor and nutrient) signalling pathways, which integrate a variety of extracellular and intracellular signals to mediate growth, proliferation or survival, are both important for cellular physiology. There is evidence to suggest that the transport and metabolic signalling pathways intersect-vesicular transport can affect the regulation of metabolic signals and vice versa. The Rab family GTPases regulate the specificity of vesicular transport steps in the cell. Together with their interacting proteins, Rabs would likely constitute the points of intersection between vesicular transport and metabolic signalling pathways. Examples of these points would include growth factor signalling, glucose and lipid metabolism, as well as autophagy. Many of these processes involve mechanistic/mammalian target of rapamycin (mTOR) complex 1 (mTORC1) in downstream cascades, or are regulated by TORC signalling. A general functionality of the vesicular transport processes controlled by the Rabs is also important for spatial and temporal regulation of the transmission of metabolic signals between the cell surface and the nucleus. In other cases, specific Rabs and their interacting proteins are known to function in recruiting metabolism-related proteins to target membranes, or may compete with other factors in the TORC signalling pathway as a means of metabolic regulation. We review and discuss herein examples of how Rabs and their interacting proteins can mediate metabolic signalling and regulation in cells.

  11. A connecter-like factor, CacA, links RssB/RpoS and the CpxR/CpxA two-component system in Salmonella

    Directory of Open Access Journals (Sweden)

    Kato Akinori

    2012-10-01

    Full Text Available Abstract Background Bacteria integrate numerous environmental stimuli when generating cellular responses. Increasing numbers of examples describe how one two-component system (TCS responds to signals detected by the sensor of another TCS. However, the molecular mechanisms underlying this phenomenon remain poorly defined. Results Here, we report a connector-like factor that affects the activity of the CpxR/CpxA two-component system in Salmonella enterica serovar Typhimurium. We isolated a clone that induced the expression of a cpxP-lac gene fusion from a high-copy-number plasmid pool of random Salmonella genomic fragments. A 63-amino acid protein, CacA, was responsible for the CpxA/CpxR-dependent activation of the cpxP gene. The CpxR-activated genes cpxP and spy exhibited approximately 30% and 50% reductions in transcription, respectively, in a clean cacA deletion mutant strain in comparison to wild-type. From 33 response regulator (RR deletion mutants, we identified that the RssB regulator represses cacA transcription. Substitution mutations in a conserved -10 region harboring the RNA polymerase recognition sequence, which is well conserved with a known RpoS -10 region consensus sequence, rendered the cacA promoter RpoS-independent. The CacA-mediated induction of cpxP transcription was affected in a trxA deletion mutant, which encodes thioredoxin 1, suggesting a role for cysteine thiol-disulfide exchange(s in CacA-dependent Cpx activation. Conclusions We identified CacA as an activator of the CpxR/CpxA system in the plasmid clone. We propose that CacA may integrate the regulatory status of RssB/RpoS into the CpxR/CpxA system. Future investigations are necessary to thoroughly elucidate how CacA activates the CpxR/CpxA system.

  12. Gene Regulation by the LiaSR Two-Component System in Streptococcus mutans.

    Directory of Open Access Journals (Sweden)

    Manoharan Shankar

    Full Text Available The LiaSR two-component signal transduction system regulates cellular responses to several environmental stresses, including those that induce cell envelope damages. Downstream regulons of the LiaSR system have been implicated in tolerance to acid, antibiotics and detergents. In the dental pathogen Streptococcus mutans, the LiaSR system is necessary for tolerance against acid, antibiotics, and cell wall damaging stresses during growth in the oral cavity. To understand the molecular mechanisms by which LiaSR regulates gene expression, we created a mutant LiaR in which the conserved aspartic acid residue (the phosphorylation site, was changed to alanine residue (D58A. As expected, the LiaR-D58A variant was unable to acquire the phosphate group and bind to target promoters. We also noted that the predicted LiaR-binding motif upstream of the lia operon does not appear to be well conserved. Consistent with this observation, we found that LiaR was unable to bind to the promoter region of lia; however, we showed that LiaR was able to bind to the promoters of SMU.753, SMU.2084 and SMU.1727. Based on sequence analysis and DNA binding studies we proposed a new 25-bp conserved motif essential for LiaR binding. Introducing alterations at fully conserved positions in the 25-bp motif affected LiaR binding, and the binding was dependent on the combination of positions that were altered. By scanning the S. mutans genome for the occurrence of the newly defined LiaR binding motif, we identified the promoter of hrcA (encoding a key regulator of the heat shock response that contains a LiaR binding motif, and we showed that hrcA is negatively regulated by the LiaSR system. Taken together our results suggest a putative role of the LiaSR system in heat shock responses of S. mutans.

  13. Landscape mapping of functional proteins in insulin signal transduction and insulin resistance: a network-based protein-protein interaction analysis.

    Directory of Open Access Journals (Sweden)

    Chiranjib Chakraborty

    Full Text Available The type 2 diabetes has increased rapidly in recent years throughout the world. The insulin signal transduction mechanism gets disrupted sometimes and it's known as insulin-resistance. It is one of the primary causes associated with type-2 diabetes. The signaling mechanisms involved several proteins that include 7 major functional proteins such as INS, INSR, IRS1, IRS2, PIK3CA, Akt2, and GLUT4. Using these 7 principal proteins, multiple sequences alignment has been created. The scores between sequences also have been developed. We have constructed a phylogenetic tree and modified it with node and distance. Besides, we have generated sequence logos and ultimately developed the protein-protein interaction network. The small insulin signal transduction protein arrangement shows complex network between the functional proteins.

  14. Inhibition of Hsp70 by Methylene Blue Affects Signaling Protein Function and Ubiquitination and Modulates Polyglutamine Protein Degradation*

    OpenAIRE

    Wang, Adrienne M; Morishima, Yoshihiro; Clapp, Kelly M.; Peng, Hwei-Ming; Pratt, William B.; Gestwicki, Jason E.; Osawa, Yoichi; Lieberman, Andrew P.

    2010-01-01

    The Hsp90/Hsp70-based chaperone machinery regulates the activity and degradation of many signaling proteins. Cycling with Hsp90 stabilizes client proteins, whereas Hsp70 interacts with chaperone-dependent E3 ubiquitin ligases to promote protein degradation. To probe these actions, small molecule inhibitors of Hsp70 would be extremely useful; however, few have been identified. Here we test the effects of methylene blue, a recently described inhibitor of Hsp70 ATPase activity, in three well est...

  15. Sizn1 is a novel protein that functions as a transcriptional coactivator of bone morphogenic protein signaling.

    Science.gov (United States)

    Cho, Ginam; Lim, Youngshin; Zand, Dina; Golden, Jeffrey A

    2008-03-01

    Bone morphogenic proteins (BMPs) play pleotrophic roles in nervous system development, and their signaling is highly regulated at virtually every step in the pathway. We have cloned a novel gene, Sizn1 (Smad-interacting zinc finger protein), which functions as a transcriptional coactivator of BMP signaling. It positively modulates BMP signaling by interacting with Smad family members and associating with CBP in the transcription complex. Sizn1 is expressed in the ventral embryonic forebrain, where, as we will show, it contributes to BMP-dependent, cholinergic-neuron-specific gene expression. These data indicate that Sizn1 is a positive modulator of BMP signaling and provide further insight into how BMP signaling can be modulated in neuronal progenitor subsets to influence cell-type-specific gene expression and development.

  16. Intra-leukocyte expression of two-component systems in Ehrlichia chaffeensis and Anaplasma phagocytophilum and effects of the histidine kinase inhibitor closantel.

    Science.gov (United States)

    Cheng, Zhihui; Kumagai, Yumi; Lin, Mingqun; Zhang, Chunbin; Rikihisa, Yasuko

    2006-08-01

    The two-component system (TCS) composed of a pair of a sensor histidine kinase and a response regulator, allows bacteria to sense signals and respond to changes in their environment through specific gene activation or repression. The present study examined TCS in the obligatory intracellular bacteria Ehrlichia chaffeensis and Anaplasma phagocytophilum, that cause human monocytic ehrlichiosis (HME) and human granulocytic anaplasmosis (HGA) respectively. The genomes of E. chaffeensis and A. phagocytophilum were each predicted to encode three pairs of TCSs. All six genes encoding three histidine kinases and three response regulators were expressed in both E. chaffeensis and A. phagocytophilum cultured in human leukocytes. Pretreatment of host cell-free E. chaffeensis or A. phagocytophilum with closantel, an inhibitor of histidine kinases, completely blocked the infection of host cells. Treatment of infected cells 1 day post infection with closantel cleared infection in dose-dependent manner. All six genes in E. chaffeensis were cloned, recombinant proteins were expressed, and polyclonal antibodies were produced. Double immunofluorescence labelling and Western blot analysis revealed that all six proteins were expressed in cell culture. Autokinase activities of the three recombinant histidine kinases from E. chaffeensis were inhibited by closantel in vitro. A number of E. chaffeensis genes, including the six TCS genes, were downregulated within 5-60 min post closantel treatment. These results suggest that these TCSs play an essential role in infection and survival of E. chaffeensis and A. phagocytophilum in human leukocytes.

  17. Identification and characterization of the two-component NtrY/NtrX regulatory system in Azospirillum brasilense

    Directory of Open Access Journals (Sweden)

    M.L. Ishida

    2002-06-01

    Full Text Available Two Azospirillum brasilense open reading frames (ORFs exhibited homology with the two-component NtrY/NtrX regulatory system from Azorhizobium caulinodans. These A. brasilense ORFs, located downstream to the nifR3ntrBC operon, were isolated, sequenced and characterized. The present study suggests that ORF1 and ORF2 correspond to the A. brasilense ntrY and ntrX genes, respectively. The amino acid sequences of A. brasilense NtrY and NtrX proteins showed high similarity to sensor/kinase and regulatory proteins, respectively. Analysis of lacZ transcriptional fusions by the ß-galactosidase assay in Escherichia coli ntrC mutants showed that the NtrY/NtrX proteins failed to activate transcription of the nifA promoter of A. brasilense. The ntrYX operon complemented a nifR3ntrBC deletion mutant of A. brasilense for nitrate-dependent growth, suggesting a possible cross-talk between the NtrY/X and NtrB/C sensor/regulator pairs. Our data support the existence of another two-component regulatory system in A. brasilense, the NtrY/NtrX system, probably involved in the regulation of nitrate assimilation.

  18. Novel protein kinase signaling systems regulating lifespan identified by small molecule library screening using Drosophila.

    Directory of Open Access Journals (Sweden)

    Stephen R Spindler

    Full Text Available Protein kinase signaling cascades control most aspects of cellular function. The ATP binding domains of signaling protein kinases are the targets of most available inhibitors. These domains are highly conserved from mammals to flies. Herein we describe screening of a library of small molecule inhibitors of protein kinases for their ability to increase Drosophila lifespan. We developed an assay system which allowed screening using the small amounts of materials normally present in commercial chemical libraries. The studies identified 17 inhibitors, the majority of which targeted tyrosine kinases associated with the epidermal growth factor receptor (EGFR, platelet-derived growth factor (PDGF/vascular endothelial growth factor (VEGF receptors, G-protein coupled receptor (GPCR, Janus kinase (JAK/signal transducer and activator of transcription (STAT, the insulin and insulin-like growth factor (IGFI receptors. Comparison of the protein kinase signaling effects of the inhibitors in vitro defined a consensus intracellular signaling profile which included decreased signaling by p38MAPK (p38, c-Jun N-terminal kinase (JNK and protein kinase C (PKC. If confirmed, many of these kinases will be novel additions to the signaling cascades known to regulate metazoan longevity.

  19. Novel protein kinase signaling systems regulating lifespan identified by small molecule library screening using Drosophila.

    Science.gov (United States)

    Spindler, Stephen R; Li, Rui; Dhahbi, Joseph M; Yamakawa, Amy; Sauer, Frank

    2012-01-01

    Protein kinase signaling cascades control most aspects of cellular function. The ATP binding domains of signaling protein kinases are the targets of most available inhibitors. These domains are highly conserved from mammals to flies. Herein we describe screening of a library of small molecule inhibitors of protein kinases for their ability to increase Drosophila lifespan. We developed an assay system which allowed screening using the small amounts of materials normally present in commercial chemical libraries. The studies identified 17 inhibitors, the majority of which targeted tyrosine kinases associated with the epidermal growth factor receptor (EGFR), platelet-derived growth factor (PDGF)/vascular endothelial growth factor (VEGF) receptors, G-protein coupled receptor (GPCR), Janus kinase (JAK)/signal transducer and activator of transcription (STAT), the insulin and insulin-like growth factor (IGFI) receptors. Comparison of the protein kinase signaling effects of the inhibitors in vitro defined a consensus intracellular signaling profile which included decreased signaling by p38MAPK (p38), c-Jun N-terminal kinase (JNK) and protein kinase C (PKC). If confirmed, many of these kinases will be novel additions to the signaling cascades known to regulate metazoan longevity.

  20. Ric-8A, a Gα protein guanine nucleotide exchange factor potentiates taste receptor signaling

    Directory of Open Access Journals (Sweden)

    Claire J Fenech

    2009-10-01

    Full Text Available Taste receptors for sweet, bitter and umami tastants are G-protein coupled receptors (GPCRs. While much effort has been devoted to understanding G-protein-receptor interactions and identifying the components of the signalling cascade downstream of these receptors, at the level of the G-protein the modulation of receptor signal transduction remains relatively unexplored. In this regard a taste-specific regulator of G-protein signaling (RGS, RGS21, has recently been identified. To study whether guanine nucleotide exchange factors (GEFs are involved in the transduction of the signal downstream of the taste GPCRs we investigated the expression of Ric-8A and Ric-8B in mouse taste cells and their interaction with G-protein subunits found in taste buds. Mammalian Ric-8 proteins were initially identified as potent GEFs for a range of Gα subunits and Ric-8B has recently been shown to amplify olfactory signal transduction. We find that both Ric-8A and Ric-8B are expressed in a large portion of taste bud cells and that most of these cells contain IP3R-3 a marker for sweet, umami and bitter taste receptor cells. Ric-8A interacts with Gα-gustducin and Gαi2 through which it amplifies the signal transduction of hTas2R16, a receptor for bitter compounds. Overall, these findings are consistent with a role for Ric-8 in mammalian taste signal transduction.

  1. High expression of G-protein signaling modulator 2 in hepatocellular carcinoma facilitates tumor growth and metastasis by activating the PI3K/AKT signaling pathway.

    Science.gov (United States)

    He, Xiao-Qin; Zhang, Yue-Feng; Yu, Jia-Jun; Gan, Yuan-Yuan; Han, Na-Na; Zhang, Mei-Xia; Ge, Wei; Deng, Jun-Jian; Zheng, Yong-Fa; Xu, Xi-Ming

    2017-03-01

    The aim of this study was to investigate the role of G-protein signaling modulator 2 in the carcinogenesis and progression of hepatocellular carcinoma. We previously showed that G-protein signaling modulator 2 was upregulated in hepatitis B virus-related hepatocellular carcinoma tissues through a hierarchical clustering analysis. With this study, we first assessed the expression pattern of G-protein signaling modulator 2 in hepatocellular carcinoma specimens and adjacent noncancerous tissues; clinical data were analyzed, along survival times, utilizing the Kaplan-Meier method. Moreover, the functions of G-protein signaling modulator 2 were examined using small-interfering RNAs in vitro. The results showed that G-protein signaling modulator 2 was clearly overexpressed in hepatocellular carcinoma tissues and cell lines and that the G-protein signaling modulator 2 expression level was related to tumor size and hepatitis B virus infection. Furthermore, G-protein signaling modulator 2 knockdown studies suggested that G-protein signaling modulator 2 accelerates cell growth, cell cycle, migration, and invasion and inhibits apoptosis, acting as an oncogene in hepatocellular carcinoma. Western blotting indicated that silencing of G-protein signaling modulator 2 in HepG2 and SMMC-7721 cells increased the expression levels of Bax, caspase-3, and E-cadherin, while notably suppressing the cyclin-dependent kinase 4, cyclin-dependent kinase 6, CyclinD1, Snail1, Vimentin, and matrix metallopeptidase 9 expression levels, compared with that in the control groups. In addition, we found that G-protein signaling modulator 2 can affect the expression of key proteins involved in protein kinase B activation. In conclusion, high expression of G-protein signaling modulator 2 was involved in the pathological processes of hepatocellular carcinoma through activation of the phosphatidylinositol 3-kinase/protein kinase B signaling pathway, which may provide an attractive potential diagnostic

  2. Negative regulation of RIG-I-mediated antiviral signaling by TRK-fused gene (TFG) protein

    Energy Technology Data Exchange (ETDEWEB)

    Lee, Na-Rae; Shin, Han-Bo; Kim, Hye-In; Choi, Myung-Soo; Inn, Kyung-Soo, E-mail: innks@khu.ac.kr

    2013-07-19

    Highlights: •TRK-fused gene product (TFG) interacts with TRIM25 upon viral infection. •TFG negatively regulates RIG-I mediated antiviral signaling. •TFG depletion leads to enhanced viral replication. •TFG act downstream of MAVS. -- Abstract: RIG-I (retinoic acid inducible gene I)-mediated antiviral signaling serves as the first line of defense against viral infection. Upon detection of viral RNA, RIG-I undergoes TRIM25 (tripartite motif protein 25)-mediated K63-linked ubiquitination, leading to type I interferon (IFN) production. In this study, we demonstrate that TRK-fused gene (TFG) protein, previously identified as a TRIM25-interacting protein, binds TRIM25 upon virus infection and negatively regulates RIG-I-mediated type-I IFN signaling. RIG-I-mediated IFN production and nuclear factor (NF)-κB signaling pathways were upregulated by the suppression of TFG expression. Furthermore, vesicular stomatitis virus (VSV) replication was significantly inhibited by small inhibitory hairpin RNA (shRNA)-mediated knockdown of TFG, supporting the suppressive role of TFG in RIG-I-mediated antiviral signaling. Interestingly, suppression of TFG expression increased not only RIG-I-mediated signaling but also MAVS (mitochondrial antiviral signaling protein)-induced signaling, suggesting that TFG plays a pivotal role in negative regulation of RNA-sensing, RIG-I-like receptor (RLR) family signaling pathways.

  3. Extracting protein dynamics information from overlapped NMR signals using relaxation dispersion difference NMR spectroscopy

    Energy Technology Data Exchange (ETDEWEB)

    Konuma, Tsuyoshi [Icahn School of Medicine at Mount Sinai, Department of Structural and Chemical Biology (United States); Harada, Erisa [Suntory Foundation for Life Sciences, Bioorganic Research Institute (Japan); Sugase, Kenji, E-mail: sugase@sunbor.or.jp, E-mail: sugase@moleng.kyoto-u.ac.jp [Kyoto University, Department of Molecular Engineering, Graduate School of Engineering (Japan)

    2015-12-15

    Protein dynamics plays important roles in many biological events, such as ligand binding and enzyme reactions. NMR is mostly used for investigating such protein dynamics in a site-specific manner. Recently, NMR has been actively applied to large proteins and intrinsically disordered proteins, which are attractive research targets. However, signal overlap, which is often observed for such proteins, hampers accurate analysis of NMR data. In this study, we have developed a new methodology called relaxation dispersion difference that can extract conformational exchange parameters from overlapped NMR signals measured using relaxation dispersion spectroscopy. In relaxation dispersion measurements, the signal intensities of fluctuating residues vary according to the Carr-Purcell-Meiboon-Gill pulsing interval, whereas those of non-fluctuating residues are constant. Therefore, subtraction of each relaxation dispersion spectrum from that with the highest signal intensities, measured at the shortest pulsing interval, leaves only the signals of the fluctuating residues. This is the principle of the relaxation dispersion difference method. This new method enabled us to extract exchange parameters from overlapped signals of heme oxygenase-1, which is a relatively large protein. The results indicate that the structural flexibility of a kink in the heme-binding site is important for efficient heme binding. Relaxation dispersion difference requires neither selectively labeled samples nor modification of pulse programs; thus it will have wide applications in protein dynamics analysis.

  4. The RGS protein Crg2 regulates both pheromone and cAMP signalling in Cryptococcus neoformans.

    Science.gov (United States)

    Xue, Chaoyang; Hsueh, Yen-Ping; Chen, Lydia; Heitman, Joseph

    2008-10-01

    G proteins orchestrate critical cellular functions by transducing extracellular signals into internal signals and controlling cellular responses to environmental cues. G proteins typically function as switches that are activated by G protein-coupled receptors (GPCRs) and negatively controlled by regulator of G protein signalling (RGS) proteins. In the human fungal pathogen Cryptococcus neoformans, three G protein alpha subunits (Gpa1, Gpa2 and Gpa3) have been identified. In a previous study, we identified the RGS protein Crg2 involved in regulating the pheromone response pathway through Gpa2 and Gpa3. In this study, a role for Crg2 was established in the Gpa1-cAMP signalling pathway that governs mating and virulence. We show that Crg2 physically interacts with Gpa1 and crg2 mutations increase cAMP production. crg2 mutations also enhance mating filament hyphae production, but reduce cell-cell fusion and sporulation efficiency during mating. Although crg2 mutations and the Gpa1 dominant active allele GPA1(Q284L) enhanced melanin production under normally repressive conditions, virulence was attenuated in a murine model. We conclude that Crg2 participates in controlling both Gpa1-cAMP-virulence and pheromone-mating signalling cascades and hypothesize it may serve as a molecular interface between these two central signalling conduits.

  5. Protein Phosphatase 2A Signaling in Human Prostate Cancer

    Science.gov (United States)

    2014-08-01

    Altogether, these findings may aid in the development of novel therapeutic strategies targeting the PP2A signaling network and/or better treatment planning ...restoration resulted in decreased nuclear accumulation and transcriptional activity of b-catenin/NF-kB, and restitution of their activity abrogated

  6. Mitogen-activated protein kinase and abscisic acid signal transduction

    NARCIS (Netherlands)

    Heimovaara-Dijkstra, S.; Testerink, C.; Wang, M.

    1998-01-01

    The phytohormone abscisic acid (ABA) is a classical plant hormone, responsible for regulation of abscission, diverse aspects of plant and seed development, stress responses and germination. It was found that ABA signal transduction in plants can involve the activity of type 2C-phosphatases (PP2C), c

  7. Large-scale Models Reveal the Two-component Mechanics of Striated Muscle

    Directory of Open Access Journals (Sweden)

    Robert Jarosch

    2008-12-01

    Full Text Available This paper provides a comprehensive explanation of striated muscle mechanics and contraction on the basis of filament rotations. Helical proteins, particularly the coiled-coils of tropomyosin, myosin and α-actinin, shorten their H-bonds cooperatively and produce torque and filament rotations when the Coulombic net-charge repulsion of their highly charged side-chains is diminished by interaction with ions. The classical “two-component model” of active muscle differentiated a “contractile component” which stretches the “series elastic component” during force production. The contractile components are the helically shaped thin filaments of muscle that shorten the sarcomeres by clockwise drilling into the myosin cross-bridges with torque decrease (= force-deficit. Muscle stretch means drawing out the thin filament helices off the cross-bridges under passive counterclockwise rotation with torque increase (= stretch activation. Since each thin filament is anchored by four elastic α-actinin Z-filaments (provided with forceregulating sites for Ca2+ binding, the thin filament rotations change the torsional twist of the four Z-filaments as the “series elastic components”. Large scale models simulate the changes of structure and force in the Z-band by the different Z-filament twisting stages A, B, C, D, E, F and G. Stage D corresponds to the isometric state. The basic phenomena of muscle physiology, i. e. latency relaxation, Fenn-effect, the force-velocity relation, the length-tension relation, unexplained energy, shortening heat, the Huxley-Simmons phases, etc. are explained and interpreted with the help of the model experiments.

  8. Cell-type dependent modulation of Notch signaling by the amyloid precursor protein.

    Science.gov (United States)

    Oh, Sun Young; Chen, Ci-Di; Abraham, Carmela R

    2010-04-01

    The amyloid precursor protein is a ubiquitously expressed transmembrane protein that has been long implicated in the pathogenesis of Alzheimer's disease but its normal biological function has remained elusive despite extensive effort. We have previously reported the identification of Notch2 as an amyloid precursor protein interacting protein in E18 rat neurons. Here, we sought to reveal the physiologic consequences of this interaction. We report a functional relationship between amyloid precursor protein and Notch1, which does not affect Delta ligand binding. First, we observed interactions between the amyloid precursor protein and Notch in mouse embryonic stem cells lacking both presenilin 1 and presenilin 2, the active proteolytic components of the gamma-secretase complex, suggesting that these two transmembrane proteins can interact in the absence of presenilin. Next, we demonstrated that the amyloid precursor protein affects Notch signaling by using Notch-dependent luciferase assays in two cell lines, the human embryonic kidney 293 and the monkey kidney, COS7. We found that the amyloid precursor protein exerts opposing effects on Notch signaling in human embryonic kidney 293 vs. COS7 cells. Finally, we show that more Notch Intracellular Domain is found in the nucleus in the presence of exogenous amyloid precursor protein or its intracellular domain, suggesting the mechanism by which the amyloid precursor protein affects Notch signaling in certain cells. Our results provide evidence of potentially important communications between the amyloid precursor protein and Notch.

  9. Phosphorelay of non-orthodox two component systems functions through a bi-molecular mechanism in vivo

    DEFF Research Database (Denmark)

    Jovanovic, Goran; Sheng, Xia; Ale, Angelique;

    2015-01-01

    the functional relevance of the dimerization of a non-orthodox or hybrid histidine kinase along which the phosphorelay takes place has been a subject of debate. We use a combination of molecular and genetic approaches, coupled to mathematical and statistical modelling, to demonstrate that the different possible...... intra- and inter-molecular mechanisms of phosphotransfer are formally non-identifiable in Escherichia coli expressing the ArcB non-orthodox histidine kinase used in anoxic redox control. In order to resolve this issue we further analyse the mathematical model in order to identify discriminatory...... in the histidine kinase. This is the first detailed mechanistic analysis of the molecular processes involved in non-orthodox two-component signalling and our results suggest strongly that dimerization facilitates more discriminatory proof-reading of external signals, via these allosteric reactions, prior to them...

  10. Phospholipase D and phosphatidic acid in plant defence response: from protein-protein and lipid-protein interactions to hormone signalling.

    Science.gov (United States)

    Zhao, Jian

    2015-04-01

    Phospholipase Ds (PLDs) and PLD-derived phosphatidic acids (PAs) play vital roles in plant hormonal and environmental responses and various cellular dynamics. Recent studies have further expanded the functions of PLDs and PAs into plant-microbe interaction. The molecular diversities and redundant functions make PLD-PA an important signalling complex regulating lipid metabolism, cytoskeleton dynamics, vesicle trafficking, and hormonal signalling in plant defence through protein-protein and protein-lipid interactions or hormone signalling. Different PLD-PA signalling complexes and their targets have emerged as fast-growing research topics for understanding their numerous but not yet established roles in modifying pathogen perception, signal transduction, and downstream defence responses. Meanwhile, advanced lipidomics tools have allowed researchers to reveal further the mechanisms of PLD-PA signalling complexes in regulating lipid metabolism and signalling, and their impacts on jasmonic acid/oxylipins, salicylic acid, and other hormone signalling pathways that essentially mediate plant defence responses. This review attempts to summarize the progress made in spatial and temporal PLD/PA signalling as well as PLD/PA-mediated modification of plant defence. It presents an in-depth discussion on the functions and potential mechanisms of PLD-PA complexes in regulating actin filament/microtubule cytoskeleton, vesicle trafficking, and hormonal signalling, and in influencing lipid metabolism-derived metabolites as critical signalling components in plant defence responses. The discussion puts PLD-PA in a broader context in order to guide future research.

  11. Products of lipid, protein and RNA oxidation as signals and regulators of gene expression in plants

    Directory of Open Access Journals (Sweden)

    Jagna eChmielowska-Bąk

    2015-06-01

    Full Text Available Reactive oxygen species (ROS are engaged in several processes essential for normal cell functioning, such as differentiation, anti-microbial defense, stimulus sensing and signaling. Interestingly, recent studies imply that cellular signal transduction and gene regulation are mediated not only directly by ROS but also by the molecules derived from ROS-mediated oxidation. Lipid peroxidation leads to non-enzymatic formation of oxylipins. These molecules were shown to modulate expression of signaling associated genes including genes encoding phosphatases, kinases and transcription factors. Oxidized peptides derived from protein oxidation might be engaged in organelle-specific ROS signaling. In turn, oxidation of particular mRNAs leads to decrease in the level of encoded proteins and thus, contributes to the post-transcriptional regulation of gene expression. Present mini review summarizes latest findings concerning involvement of products of lipid, protein and RNA oxidation in signal transduction and gene regulation.

  12. Role of SRC-like adaptor protein (SLAP) in immune and malignant cell signaling.

    Science.gov (United States)

    Kazi, Julhash U; Kabir, Nuzhat N; Rönnstrand, Lars

    2015-07-01

    SRC-like adaptor protein (SLAP) is an adaptor protein structurally similar to the SRC family protein kinases. Like SRC, SLAP contains an SH3 domain followed by an SH2 domain but the kinase domain has been replaced by a unique C-terminal region. SLAP is expressed in a variety of cell types. Current studies suggest that it regulates signaling of various cell surface receptors including the B cell receptor, the T cell receptor, cytokine receptors and receptor tyrosine kinases which are important regulator of immune and cancer cell signaling. SLAP targets receptors, or its associated components, by recruiting the ubiquitin machinery and thereby destabilizing signaling. SLAP directs receptors to ubiquitination-mediated degradation and controls receptors turnover as well as signaling. Thus, SLAP appears to be an important component in regulating signal transduction required for immune and malignant cells.

  13. Application of native signal sequences for recombinant proteins secretion in Pichia pastoris

    DEFF Research Database (Denmark)

    Borodina, Irina; Do, Duy Duc; Eriksen, Jens C.;

    alpha‐mating factor (MF) prepropeptide from Saccharomyces cerevisiae is most commonly used. Our aim was to test whether signal peptides from P. pastoris native secreted proteins could be used to direct secretion of recombinant proteins. Results Eleven native signal peptides from P. pastoris were tested...... with the promoter and signal peptide of choice (or a mix thereof) and the basic expression vector. The mix is treated with USER enzyme and transformed into E. coli. The plasmids (or plasmid mixes) are further purified, linearized and transformed into P. pastoris. We illustrate the commodity of the system...... by optimization of expression of three different proteins in P. pastoris. Conclusions Native signal peptides from P. pastoris can be used to direct secretion of recombinant proteins. A novel USER‐based P. pastoris system allows easy cloning of protein‐coding gene with the promoter and leader sequence of choice....

  14. How a Mycoparasite Employs G-Protein Signaling: Using the Example of Trichoderma

    Directory of Open Access Journals (Sweden)

    Markus Omann

    2010-01-01

    Full Text Available Mycoparasitic Trichoderma spp. act as potent biocontrol agents against a number of plant pathogenic fungi, whereupon the mycoparasitic attack includes host recognition followed by infection structure formation and secretion of lytic enzymes and antifungal metabolites leading to the host's death. Host-derived signals are suggested to be recognized by receptors located on the mycoparasite's cell surface eliciting an internal signal transduction cascade which results in the transcription of mycoparasitism-relevant genes. Heterotrimeric G proteins of fungi transmit signals originating from G-protein-coupled receptors mainly to the cAMP and the MAP kinase pathways resulting in regulation of downstream effectors. Components of the G-protein signaling machinery such as G subunits and G-protein-coupled receptors were recently shown to play crucial roles in Trichoderma mycoparasitism as they govern processes such as the production of extracellular cell wall lytic enzymes, the secretion of antifungal metabolites, and the formation of infection structures.

  15. Cell signaling, post-translational protein modifications and NMR spectroscopy

    Energy Technology Data Exchange (ETDEWEB)

    Theillet, Francois-Xavier [In-cell NMR Group, Department of NMR-Supported Structural Biology, Leibniz Institute of Molecular Pharmacology (FMP Berlin) (Germany); Smet-Nocca, Caroline [Universite Lille Nord de France, CNRS UMR 8576 (France); Liokatis, Stamatios; Thongwichian, Rossukon; Kosten, Jonas [In-cell NMR Group, Department of NMR-Supported Structural Biology, Leibniz Institute of Molecular Pharmacology (FMP Berlin) (Germany); Yoon, Mi-Kyung; Kriwacki, Richard W. [St. Jude Children' s Research Hospital, Department of Structural Biology (United States); Landrieu, Isabelle; Lippens, Guy [Universite Lille Nord de France, CNRS UMR 8576 (France); Selenko, Philipp, E-mail: selenko@fmp-berlin.de [In-cell NMR Group, Department of NMR-Supported Structural Biology, Leibniz Institute of Molecular Pharmacology (FMP Berlin) (Germany)

    2012-11-15

    Post-translationally modified proteins make up the majority of the proteome and establish, to a large part, the impressive level of functional diversity in higher, multi-cellular organisms. Most eukaryotic post-translational protein modifications (PTMs) denote reversible, covalent additions of small chemical entities such as phosphate-, acyl-, alkyl- and glycosyl-groups onto selected subsets of modifiable amino acids. In turn, these modifications induce highly specific changes in the chemical environments of individual protein residues, which are readily detected by high-resolution NMR spectroscopy. In the following, we provide a concise compendium of NMR characteristics of the main types of eukaryotic PTMs: serine, threonine, tyrosine and histidine phosphorylation, lysine acetylation, lysine and arginine methylation, and serine, threonine O-glycosylation. We further delineate the previously uncharacterized NMR properties of lysine propionylation, butyrylation, succinylation, malonylation and crotonylation, which, altogether, define an initial reference frame for comprehensive PTM studies by high-resolution NMR spectroscopy.

  16. Protein signaling networks from single cell fluctuations and information theory profiling.

    Science.gov (United States)

    Shin, Young Shik; Remacle, F; Fan, Rong; Hwang, Kiwook; Wei, Wei; Ahmad, Habib; Levine, R D; Heath, James R

    2011-05-18

    Protein signaling networks among cells play critical roles in a host of pathophysiological processes, from inflammation to tumorigenesis. We report on an approach that integrates microfluidic cell handling, in situ protein secretion profiling, and information theory to determine an extracellular protein-signaling network and the role of perturbations. We assayed 12 proteins secreted from human macrophages that were subjected to lipopolysaccharide challenge, which emulates the macrophage-based innate immune responses against Gram-negative bacteria. We characterize the fluctuations in protein secretion of single cells, and of small cell colonies (n = 2, 3,···), as a function of colony size. Measuring the fluctuations permits a validation of the conditions required for the application of a quantitative version of the Le Chatelier's principle, as derived using information theory. This principle provides a quantitative prediction of the role of perturbations and allows a characterization of a protein-protein interaction network.

  17. Direct Modulation of Heterotrimeric G Protein-coupled Signaling by a Receptor Kinase Complex.

    Science.gov (United States)

    Tunc-Ozdemir, Meral; Urano, Daisuke; Jaiswal, Dinesh Kumar; Clouse, Steven D; Jones, Alan M

    2016-07-01

    Plants and some protists have heterotrimeric G protein complexes that activate spontaneously without canonical G protein-coupled receptors (GPCRs). In Arabidopsis, the sole 7-transmembrane regulator of G protein signaling 1 (AtRGS1) modulates the G protein complex by keeping it in the resting state (GDP-bound). However, it remains unknown how a myriad of biological responses is achieved with a single G protein modulator. We propose that in complete contrast to G protein activation in animals, plant leucine-rich repeat receptor-like kinases (LRR RLKs), not GPCRs, provide this discrimination through phosphorylation of AtRGS1 in a ligand-dependent manner. G protein signaling is directly activated by the pathogen-associated molecular pattern flagellin peptide 22 through its LRR RLK, FLS2, and co-receptor BAK1.

  18. G Protein-Coupled Receptor Signaling in Stem Cells and Cancer

    Directory of Open Access Journals (Sweden)

    Jennifer R. Lynch

    2016-05-01

    Full Text Available G protein-coupled receptors (GPCRs are a large superfamily of cell-surface signaling proteins that bind extracellular ligands and transduce signals into cells via heterotrimeric G proteins. GPCRs are highly tractable drug targets. Aberrant expression of GPCRs and G proteins has been observed in various cancers and their importance in cancer stem cells has begun to be appreciated. We have recently reported essential roles for G protein-coupled receptor 84 (GPR84 and G protein subunit Gαq in the maintenance of cancer stem cells in acute myeloid leukemia. This review will discuss how GPCRs and G proteins regulate stem cells with a focus on cancer stem cells, as well as their implications for the development of novel targeted cancer therapies.

  19. Protein Signaling Networks from Single Cell Fluctuations and Information Theory Profiling

    Science.gov (United States)

    Shin, Young Shik; Remacle, F.; Fan, Rong; Hwang, Kiwook; Wei, Wei; Ahmad, Habib; Levine, R.D.; Heath, James R.

    2011-01-01

    Protein signaling networks among cells play critical roles in a host of pathophysiological processes, from inflammation to tumorigenesis. We report on an approach that integrates microfluidic cell handling, in situ protein secretion profiling, and information theory to determine an extracellular protein-signaling network and the role of perturbations. We assayed 12 proteins secreted from human macrophages that were subjected to lipopolysaccharide challenge, which emulates the macrophage-based innate immune responses against Gram-negative bacteria. We characterize the fluctuations in protein secretion of single cells, and of small cell colonies (n = 2, 3,···), as a function of colony size. Measuring the fluctuations permits a validation of the conditions required for the application of a quantitative version of the Le Chatelier's principle, as derived using information theory. This principle provides a quantitative prediction of the role of perturbations and allows a characterization of a protein-protein interaction network. PMID:21575571

  20. Signaling by bone morphogenetic proteins directs formation of an ectodermal signaling center that regulates craniofacial development.

    Science.gov (United States)

    Foppiano, Silvia; Hu, Diane; Marcucio, Ralph S

    2007-12-01

    We previously described a signaling center, the Frontonasal Ectodermal Zone (FEZ) that regulates growth and patterning of the frontonasal process (FNP). The FEZ is comprised of FNP ectoderm flanking a boundary between Sonic hedgehog (Shh) and Fibroblast growth factor 8 (Fgf8) expression domains. Our objective was to examine BMP signaling during formation of the FEZ. We blocked BMP signaling throughout the FNP prior to FEZ formation by infecting chick embryos at stage 10 (HH10) with a replication-competent avian retrovirus encoding the BMP antagonist Noggin. We assessed gene expression patterns in the FNP 72 h after infection (approximately HH22) and observed that Shh expression was reduced or absent. In the mesenchyme, we observed that Bmp2 transcripts were absent while the Bmp4 expression domain was expanded proximally. In addition to the molecular changes, infected embryos also exhibited facial malformations at 72 and 96 h after infection suggesting that the FEZ did not form. Our data indicate that reduced cell proliferation, but not apoptosis, in the mesenchyme contributed to the phenotype that we observed. Additionally, adding exogenous SHH into the mesenchyme of RCAS-Noggin-infected embryos did not restore Bmp2 and Bmp4 to a normal pattern of expression. These data indicate that BMP signaling mediates interactions between tissues in the FNP that regulate FEZ formation; and that the correct pattern of Bmp2 and Bmp4, but not Bmp7, expression in the FNP mesenchyme requires signaling by the BMP pathway.

  1. A key temporal delay in the circadian cycle of Drosophila is mediated by a nuclear localization signal in the timeless protein.

    Science.gov (United States)

    Saez, Lino; Derasmo, Mary; Meyer, Pablo; Stieglitz, J; Young, Michael W

    2011-07-01

    Regulated nuclear entry of the Period (PER) and Timeless (TIM) proteins, two components of the Drosophila circadian clock, is essential for the generation and maintenance of circadian behavior. PER and TIM shift from the cytoplasm to the nucleus daily, and the length of time that PER and TIM reside in the cytoplasm is an important determinant of the period length of the circadian rhythm. Here we identify a TIM nuclear localization signal (NLS) that is required for appropriately timed nuclear accumulation of both TIM and PER. Transgenic flies with a mutated TIM NLS produced circadian rhythms with a period of ∼30 hr. In pacemaker cells of the brain, PER and TIM proteins rise to abnormally high levels in the cytoplasm of tim(ΔNLS) mutants, but show substantially reduced nuclear accumulation. In cultured S2 cells, the mutant TIM(ΔNLS) protein significantly delays nuclear accumulation of both TIM and wild-type PER proteins. These studies confirm that TIM is required for the nuclear localization of PER and point to a key role for the TIM NLS in the regulated nuclear accumulation of both proteins.

  2. Intrinsic Disorder in Transmembrane Proteins: Roles in Signaling and Topology Prediction.

    Directory of Open Access Journals (Sweden)

    Jérôme Bürgi

    Full Text Available Intrinsically disordered regions (IDRs are peculiar stretches of amino acids that lack stable conformations in solution. Intrinsic Disorder containing Proteins (IDP are defined by the presence of at least one large IDR and have been linked to multiple cellular processes including cell signaling, DNA binding and cancer. Here we used computational analyses and publicly available databases to deepen insight into the prevalence and function of IDRs specifically in transmembrane proteins, which are somewhat neglected in most studies. We found that 50% of transmembrane proteins have at least one IDR of 30 amino acids or more. Interestingly, these domains preferentially localize to the cytoplasmic side especially of multi-pass transmembrane proteins, suggesting that disorder prediction could increase the confidence of topology prediction algorithms. This was supported by the successful prediction of the topology of the uncharacterized multi-pass transmembrane protein TMEM117, as confirmed experimentally. Pathway analysis indicated that IDPs are enriched in cell projection and axons and appear to play an important role in cell adhesion, signaling and ion binding. In addition, we found that IDP are enriched in phosphorylation sites, a crucial post translational modification in signal transduction, when compared to fully ordered proteins and to be implicated in more protein-protein interaction events. Accordingly, IDPs were highly enriched in short protein binding regions called Molecular Recognition Features (MoRFs. Altogether our analyses strongly support the notion that the transmembrane IDPs act as hubs in cellular signal events.

  3. Identification of two functional nuclear localization signals in the capsid protein of duck circovirus

    Energy Technology Data Exchange (ETDEWEB)

    Xiang, Qi-Wang; Zou, Jin-Feng; Wang, Xin [Department of Preventive Veterinary Medicine, College of Veterinary Medicine, Shandong Agricultural University, Shandong, Taian 271018 (China); Shandong Provincial Key Laboratory of Animal Biotechnology and Disease Control and Prevention, Shandong, Taian 271018 (China); Sun, Ya-Ni [College of Veterinary Medicine, Northwest A and F University, Shanxi, Yangling 712100 (China); Gao, Ji-Ming; Xie, Zhi-Jing [Department of Preventive Veterinary Medicine, College of Veterinary Medicine, Shandong Agricultural University, Shandong, Taian 271018 (China); Shandong Provincial Key Laboratory of Animal Biotechnology and Disease Control and Prevention, Shandong, Taian 271018 (China); Wang, Yu [Department of Basic Medical Sciences, Taishan Medical College, Shandong, Taian 271000 (China); Zhu, Yan-Li [Department of Preventive Veterinary Medicine, College of Veterinary Medicine, Shandong Agricultural University, Shandong, Taian 271018 (China); Shandong Provincial Key Laboratory of Animal Biotechnology and Disease Control and Prevention, Shandong, Taian 271018 (China); Jiang, Shi-Jin, E-mail: sjjiang@sdau.edu.cn [Department of Preventive Veterinary Medicine, College of Veterinary Medicine, Shandong Agricultural University, Shandong, Taian 271018 (China); Shandong Provincial Key Laboratory of Animal Biotechnology and Disease Control and Prevention, Shandong, Taian 271018 (China)

    2013-02-05

    The capsid protein (CP) of duck circovirus (DuCV) is the major immunogenic protein and has a high proportion of arginine residues concentrated at the N terminus of the protein, which inhibits efficient mRNA translation in prokaryotic expression systems. In this study, we investigated the subcellular distribution of DuCV CP expressed via recombinant baculoviruses in Sf9 cells and the DNA binding activities of the truncated recombinant DuCV CPs. The results showed that two independent bipartite nuclear localization signals (NLSs) situated at N-terminal 1-17 and 18-36 amino acid residue of the CP. Moreover, two expression level regulatory signals (ELRSs) and two DNA binding signals (DBSs) were also mapped to the N terminus of the protein and overlapped with the two NLSs. The ability of CP to bind DNA, coupled with the karyophilic nature of this protein, strongly suggests that it may be responsible for nuclear targeting of the viral genome.

  4. Adult neurogenesis requires Smad4-mediated bone morphogenic protein signaling in stem cells.

    NARCIS (Netherlands)

    Colak, D.; Mori, T.; Brill, M.S; Pfeifer, A.; Falk, S.; Deng, C.; Monteiro, R.; Mummery, C.L.; Sommer, L.; Gotz, M.

    2008-01-01

    In the mammalian brain, neurogenesis continues only in few regions of the forebrain. The molecular signals governing neurogenesis in these unique neurogenic niches, however, are still ill defined. Here, we show that bone morphogenic protein (BMP)-mediated signaling is active in adult neural stem cel

  5. Cross talk between insulin and bone morphogenetic protein signaling systems in brown adipogenesis

    DEFF Research Database (Denmark)

    Zhang, Hongbin; Schulz, Tim J; Espinoza, Daniel O;

    2010-01-01

    Both insulin and bone morphogenetic protein (BMP) signaling systems are important for adipocyte differentiation. Analysis of gene expression in BMP7-treated fibroblasts revealed a coordinated change in insulin signaling components by BMP7. To further investigate the cross talk between insulin and...

  6. Bone morphogenetic protein signaling suppresses tumorigenesis at gastric epithelial transition zones in mice

    NARCIS (Netherlands)

    Bleuming, Sylvia A.; He, Xi C.; Kodach, Liudmila L.; Hardwick, James C.; Koopman, Frieda A.; ten Kate, Fiebo J.; van Deventer, Sander J. H.; Hommes, Daniel W.; Peppelenbosch, Maikel P.; Offerhaus, G. Johan; Li, Linheng; van den Brink, Gijs R.

    2007-01-01

    Bone morphogenetic protein (BMP) signaling is known to suppress oncogenesis in the small and large intestine of mice and humans. We examined the role of Bmpr1a signaling in the stomach. On conditional inactivation of Bmpr1a, mice developed neoplastic lesions specifically in the squamocolumnar and ga

  7. Protein tyrosine phosphatase PTPRR isoforms in cellular signaling and trafficking

    NARCIS (Netherlands)

    Dilaver, Gönül

    2005-01-01

    Previous work has revealed the existence of two Protein Tyrosine Phosphatases in mouse, PTPBR7 and PTP-SL, that were in part identical, suggesting that they originated from the same gene, termed Ptprr (1,5,6). In this thesis, I report on the characterization of the various PTPRR isoforms in neuronal

  8. Protein Kinase Signalling in the Moss Physcomitrella patens

    DEFF Research Database (Denmark)

    Azevedo de Silva, Raquel

    , such as salt or osmotic stress. Moreover, the PpMEKK1 homologs seem to be involved in ABA-dependent responses, such as, dehydration. Rescue of mekk1 knockouts phenotype when exogenous ABA is applied and downregulation of ABA biosynthesis genes suggests that these proteins function upstream or parallel...

  9. Signaling pathways of the ING proteins in apoptosis.

    Science.gov (United States)

    Shah, Sitar; Riabowol, Karl

    2009-05-01

    Members of the ING family of type II tumor suppressors reside in different chromatin regulatory complexes and are stoichiometeric members of histone acetyltransferase (HAT) and histone deacetylase (HDAC) complexes. It has been frequently observed that expressing ING proteins promotes apoptosis in both normal and transformed cells of different species. They have also been reported to either rely upon p53, or to add to its ability to promote programmed cell death (apoptosis) although whether ING proteins require p53 to induce apoptosis is now questionable based upon observations using knockout cell lines and animal models. Genetic studies in model organisms, and particularly in Caenorhabditis elegans, have identified different pathways involved in apoptosis during development, in the germ line and in response to various forms of stress including DNA damage. In this review we summarize structural features of the INGs and recent observations made in knockout models of Mus musculus and Caenorhabditis elegans that have helped to further clarify the functions of the ING proteins in biochemical pathways leading to apoptosis. Based upon these observations we propose a model for how ING proteins may act both independently and in concert with p53 to promote apoptosis.

  10. Mapping membrane protein interactions in cell signaling systems.

    Energy Technology Data Exchange (ETDEWEB)

    Light, Yooli Kim; Hadi, Masood Z.; Lane, Pamela; Jacobsen, Richard B.; Hong, Joohee; Ayson, Marites J.; Wood, Nichole L.; Schoeniger, Joseph S.; Young, Malin M.

    2003-12-01

    We proposed to apply a chemical cross-linking, mass spectrometry and modeling method called MS3D to the structure determination of the rhodopsin-transducin membrane protein complex (RTC). Herein we describe experimental progress made to adapt the MS3D approach for characterizing membrane protein systems, and computational progress in experimental design, data analysis and protein structure modeling. Over the past three years, we have developed tailored experimental methods for all steps in the MS3D method for rhodopsin, including protein purification, a functional assay, cross-linking, proteolysis and mass spectrometry. In support of the experimental effort. we have out a data analysis pipeline in place that automatically selects the monoisotopic peaks in a mass spectrometric spectrum, assigns them and stores the results in a database. Theoretical calculations using 24 experimentally-derived distance constraints have resulted in a backbone-level model of the activated form of rhodopsin, which is a critical first step towards building a model of the RTC. Cross-linked rhodopsin-transducin complexes have been isolated via gel electrophoresis and further mass spectrometric characterization of the cross-links is underway.

  11. Connecting G protein signaling to chemoattractant-mediated cell polarity and cytoskeletal reorganization.

    Science.gov (United States)

    Liu, Youtao; Lacal, Jesus; Firtel, Richard A; Kortholt, Arjan

    2016-10-07

    The directional movement towards extracellular chemical gradients, a process called chemotaxis, is an important property of cells. Central to eukaryotic chemotaxis is the molecular mechanism by which chemoattractant-mediated activation of G-protein coupled receptors (GPCRs) induces symmetry breaking in the activated downstream signaling pathways. Studies with mainly Dictyostelium and mammalian neutrophils as experimental systems have shown that chemotaxis is mediated by a complex network of signaling pathways. Recently, several labs have used extensive and efficient proteomic approaches to further unravel this dynamic signaling network. Together these studies showed the critical role of the interplay between heterotrimeric G-protein subunits and monomeric G proteins in regulating cytoskeletal rearrangements during chemotaxis. Here we highlight how these proteomic studies have provided greater insight into the mechanisms by which the heterotrimeric G protein cycle is regulated, how heterotrimeric G proteins-induced symmetry breaking is mediated through small G protein signaling, and how symmetry breaking in G protein signaling subsequently induces cytoskeleton rearrangements and cell migration.

  12. Modeling Thermal Dust Emission with Two Components: Application to the Planck High Frequency Instrument Maps

    Science.gov (United States)

    Meisner, Aaron M.; Finkbeiner, Douglas P.

    2015-01-01

    We apply the Finkbeiner et al. two-component thermal dust emission model to the Planck High Frequency Instrument maps. This parameterization of the far-infrared dust spectrum as the sum of two modified blackbodies (MBBs) serves as an important alternative to the commonly adopted single-MBB dust emission model. Analyzing the joint Planck/DIRBE dust spectrum, we show that two-component models provide a better fit to the 100-3000 GHz emission than do single-MBB models, though by a lesser margin than found by Finkbeiner et al. based on FIRAS and DIRBE. We also derive full-sky 6.'1 resolution maps of dust optical depth and temperature by fitting the two-component model to Planck 217-857 GHz along with DIRBE/IRAS 100 μm data. Because our two-component model matches the dust spectrum near its peak, accounts for the spectrum's flattening at millimeter wavelengths, and specifies dust temperature at 6.'1 FWHM, our model provides reliable, high-resolution thermal dust emission foreground predictions from 100 to 3000 GHz. We find that, in diffuse sky regions, our two-component 100-217 GHz predictions are on average accurate to within 2.2%, while extrapolating the Planck Collaboration et al. single-MBB model systematically underpredicts emission by 18.8% at 100 GHz, 12.6% at 143 GHz, and 7.9% at 217 GHz. We calibrate our two-component optical depth to reddening, and compare with reddening estimates based on stellar spectra. We find the dominant systematic problems in our temperature/reddening maps to be zodiacal light on large angular scales and the cosmic infrared background anisotropy on small angular scales.

  13. Bacillus subtilis Two-Component System Sensory Kinase DegS Is Regulated by Serine Phosphorylation in Its Input Domain

    Science.gov (United States)

    Jers, Carsten; Kobir, Ahasanul; Søndergaard, Elsebeth Oline; Jensen, Peter Ruhdal; Mijakovic, Ivan

    2011-01-01

    Bacillus subtilis two-component system DegS/U is well known for the complexity of its regulation. The cytosolic sensory kinase DegS does not receive a single predominant input signal like most two-component kinases, instead it integrates a wide array of metabolic inputs that modulate its activity. The phosphorylation state of the response regulator DegU also does not confer a straightforward “on/off” response; it is fine-tuned and at different levels triggers different sub-regulons. Here we describe serine phosphorylation of the DegS sensing domain, which stimulates its kinase activity. We demonstrate that DegS phosphorylation can be carried out by at least two B. subtilis Hanks-type kinases in vitro, and this stimulates the phosphate transfer towards DegU. The consequences of this process were studied in vivo, using phosphomimetic (Ser76Asp) and non-phosphorylatable (Ser76Ala) mutants of DegS. In a number of physiological assays focused on different processes regulated by DegU, DegS S76D phosphomimetic mutant behaved like a strain with intermediate levels of DegU phosphorylation, whereas DegS S76A behaved like a strain with lower levels of DegU phophorylation. These findings suggest a link between DegS phosphorylation at serine 76 and the level of DegU phosphorylation, establishing this post-translational modification as an additional trigger for this two-component system. PMID:21304896

  14. Bacillus subtilis two-component system sensory kinase DegS is regulated by serine phosphorylation in its input domain.

    Directory of Open Access Journals (Sweden)

    Carsten Jers

    Full Text Available Bacillus subtilis two-component system DegS/U is well known for the complexity of its regulation. The cytosolic sensory kinase DegS does not receive a single predominant input signal like most two-component kinases, instead it integrates a wide array of metabolic inputs that modulate its activity. The phosphorylation state of the response regulator DegU also does not confer a straightforward "on/off" response; it is fine-tuned and at different levels triggers different sub-regulons. Here we describe serine phosphorylation of the DegS sensing domain, which stimulates its kinase activity. We demonstrate that DegS phosphorylation can be carried out by at least two B. subtilis Hanks-type kinases in vitro, and this stimulates the phosphate transfer towards DegU. The consequences of this process were studied in vivo, using phosphomimetic (Ser76Asp and non-phosphorylatable (Ser76Ala mutants of DegS. In a number of physiological assays focused on different processes regulated by DegU, DegS S76D phosphomimetic mutant behaved like a strain with intermediate levels of DegU phosphorylation, whereas DegS S76A behaved like a strain with lower levels of DegU phophorylation. These findings suggest a link between DegS phosphorylation at serine 76 and the level of DegU phosphorylation, establishing this post-translational modification as an additional trigger for this two-component system.

  15. The protein interaction network of a taxis signal transduction system in a Halophilic Archaeon

    Directory of Open Access Journals (Sweden)

    Schlesner Matthias

    2012-11-01

    Full Text Available Abstract Background The taxis signaling system of the extreme halophilic archaeon Halobacterium (Hbt. salinarum differs in several aspects from its model bacterial counterparts Escherichia coli and Bacillus subtilis. We studied the protein interactions in the Hbt. salinarum taxis signaling system to gain an understanding of its structure, to gain knowledge about its known components and to search for new members. Results The interaction analysis revealed that the core signaling proteins are involved in different protein complexes and our data provide evidence for dynamic interchanges between them. Fifteen of the eighteen taxis receptors (halobacterial transducers, Htrs can be assigned to four different groups depending on their interactions with the core signaling proteins. Only one of these groups, which contains six of the eight Htrs with known signals, shows the composition expected for signaling complexes (receptor, kinase CheA, adaptor CheW, response regulator CheY. From the two Hbt. salinarum CheW proteins, only CheW1 is engaged in signaling complexes with Htrs and CheA, whereas CheW2 interacts with Htrs but not with CheA. CheY connects the core signaling structure to a subnetwork consisting of the two CheF proteins (which build a link to the flagellar apparatus, CheD (the hub of the subnetwork, two CheC complexes and the receptor methylesterase CheB. Conclusions Based on our findings, we propose two hypotheses. First, Hbt. salinarum might have the capability to dynamically adjust the impact of certain Htrs or Htr clusters depending on its current needs or environmental conditions. Secondly, we propose a hypothetical feedback loop from the response regulator to Htr methylation made from the CheC proteins, CheD and CheB, which might contribute to adaptation analogous to the CheC/CheD system of B. subtilis.

  16. Domain Walls and Textured Vortices in a Two-Component Ginzburg-Landau Model

    DEFF Research Database (Denmark)

    Madsen, Søren Peder; Gaididei, Yu. B.; Christiansen, Peter Leth

    2005-01-01

    We look for domain wall and textured vortex solutions in a two-component Ginzburg-Landau model inspired by two-band superconductivity. The two-dimensional two-component model, with equal coherence lengths and no magnetic field, shows some interesting properties. In the absence of a Josephson type...... coupling between the two order parameters a ''textured vortex'' is found by analytical and numerical solution of the Ginzburg-Landau equations. With a Josephson type coupling between the two order parameters we find the system to split up in two domains separated by a domain wall, where the order parameter...

  17. Block algebra in two-component BKP and D type Drinfeld-Sokolov hierarchies

    Science.gov (United States)

    Li, Chuanzhong; He, Jingsong

    2013-11-01

    We construct generalized additional symmetries of a two-component BKP hierarchy defined by two pseudo-differential Lax operators. These additional symmetry flows form a Block type algebra with some modified (or additional) terms because of a B type reduction condition of this integrable hierarchy. Further we show that the D type Drinfeld-Sokolov hierarchy, which is a reduction of the two-component BKP hierarchy, possess a complete Block type additional symmetry algebra. That D type Drinfeld-Sokolov hierarchy has a similar algebraic structure as the bigraded Toda hierarchy which is a differential-discrete integrable system.

  18. Rabi Oscillations in Two-Component Bose-Einstein Condensates with a Coupling Drive

    Institute of Scientific and Technical Information of China (English)

    LI Wei-Dong; FAN Wen-Bing; ZHOU Xiao-Ji; WANG Yi-Qiu; LIANG Jiu-Qing

    2002-01-01

    The Rabi oscillations in two-component Bose-Einstein condensates with a coupling drive are studiedby means of a pair of bosonic operators. The coupling drive and initial phase difference will affect the amplitudeand the period of the Rabi oscillations. The Rabi oscillations will vanish in the evolution of the condensate densityfor some special initial phase differences (ψ = 0 or π). Our theory provides not only an analytical framework forquantitative predictions for two-component condensates, but also gives an intuitive understanding of some mysteriousfeatures observed in experiments and numerical. simulations.

  19. Modulational instability of two-component Bose-Einstein condensates in an optical lattice

    CERN Document Server

    Jin, G R; Nahm, K; Jin, Guang-Ri; Kim, Chul Koo; Nahm, Kyun

    2004-01-01

    We study modulational instability of two-component Bose-Einstein condensates in a deep optical lattice, which is modelled as a coupled discrete nonlinear Schr\\"{o}dinger equation. The excitation spectrum and the modulational instability condition of the total system are presented analytically. In the long-wavelength limit, our results agree with the homogeneous two-component Bose-Einstein condensates case. The discreteness effects result in the appearance of the modulational instability for the condensates in miscible region. The numerical calculations confirm our analytical results and show that the interspecies coupling can transfer the instability from one component to another.

  20. Domain Walls and Textured Vortices in a Two-Component Ginzburg-Landau Model

    DEFF Research Database (Denmark)

    Madsen, Søren Peder; Gaididei, Yu. B.; Christiansen, Peter Leth

    2005-01-01

    We look for domain wall and textured vortex solutions in a two-component Ginzburg-Landau model inspired by two-band superconductivity. The two-dimensional two-component model, with equal coherence lengths and no magnetic field, shows some interesting properties. In the absence of a Josephson type...... coupling between the two order parameters a ''textured vortex'' is found by analytical and numerical solution of the Ginzburg-Landau equations. With a Josephson type coupling between the two order parameters we find the system to split up in two domains separated by a domain wall, where the order parameter...

  1. Block algebra in two-component BKP and D type Drinfeld-Sokolov hierarchies

    Energy Technology Data Exchange (ETDEWEB)

    Li, Chuanzhong, E-mail: lichuanzhong@nbu.edu.cn; He, Jingsong, E-mail: hejingsong@nbu.edu.cn [Department of Mathematics, Ningbo University, Ningbo 315211 (China)

    2013-11-15

    We construct generalized additional symmetries of a two-component BKP hierarchy defined by two pseudo-differential Lax operators. These additional symmetry flows form a Block type algebra with some modified (or additional) terms because of a B type reduction condition of this integrable hierarchy. Further we show that the D type Drinfeld-Sokolov hierarchy, which is a reduction of the two-component BKP hierarchy, possess a complete Block type additional symmetry algebra. That D type Drinfeld-Sokolov hierarchy has a similar algebraic structure as the bigraded Toda hierarchy which is a differential-discrete integrable system.

  2. Presenilin dependence of phospholipase C and protein kinase C signaling

    DEFF Research Database (Denmark)

    Dehvari, Nodi; Cedazo-Minguez, Angel; Isacsson, Ola

    2007-01-01

    -stimulated phospholipase C (PLC) activity which was gamma-secretase dependent. To further evaluate the dependence of PLC on PSs we measured PLC activity and the activation of variant protein kinase C (PKC) isoforms in mouse embryonic fibroblasts (MEFs) lacking either PS1, PS2, or both. PLC activity and PKCalpha...... and PKCgamma activations were significantly lower in PS1 and PS2 double knockout MEFs after PLC stimulation. Protein levels of PKCalpha and PKCgamma were lower in PS1 and PS2 double knockout MEFs. In contrast, PKCdelta levels were significantly elevated in PS1 and PS2 double knockout as well as in PS1 knockout......). These results show that PLC and PKC activations are modulated by PS and also that PSs differentially regulate the expression of PKC isoforms by both APP/AICD-dependent and independent mechanisms....

  3. Structural diversity in twin-arginine signal peptide-binding proteins

    NARCIS (Netherlands)

    Maillard, J.; Spronk, C.A.E.M.; Buchanan, G.; Lyall, V.; Richardson, D.J.; Palmer, T.; Vuister, G.W.; Sargent, F.

    2007-01-01

    The twin-arginine transport (Tat) system is dedicated to the translocation of folded proteins across the bacterial cytoplasmic membrane. Proteins are targeted to the Tat system by signal peptides containing a twin-arginine motif. In Escherichia coli, many Tat substrates bind redox-active cofactors i

  4. Membrane Recruitment of the Non-receptor Protein GIV/Girdin (Gα-interacting, Vesicle-associated Protein/Girdin) Is Sufficient for Activating Heterotrimeric G Protein Signaling.

    Science.gov (United States)

    Parag-Sharma, Kshitij; Leyme, Anthony; DiGiacomo, Vincent; Marivin, Arthur; Broselid, Stefan; Garcia-Marcos, Mikel

    2016-12-30

    GIV (aka Girdin) is a guanine nucleotide exchange factor that activates heterotrimeric G protein signaling downstream of RTKs and integrins, thereby serving as a platform for signaling cascade cross-talk. GIV is recruited to the cytoplasmic tail of receptors upon stimulation, but the mechanism of activation of its G protein regulatory function is not well understood. Here we used assays in humanized yeast models and G protein activity biosensors in mammalian cells to investigate the role of GIV subcellular compartmentalization in regulating its ability to promote G protein signaling. We found that in unstimulated cells GIV does not co-fractionate with its substrate G protein Gαi3 on cell membranes and that constitutive membrane anchoring of GIV in yeast cells or rapid membrane translocation in mammalian cells via chemically induced dimerization leads to robust G protein activation. We show that membrane recruitment of the GIV "Gα binding and activating" motif alone is sufficient for G protein activation and that it does not require phosphomodification. Furthermore, we engineered a synthetic protein to show that recruitment of the GIV "Gα binding and activating" motif to membranes via association with active RTKs, instead of via chemically induced dimerization, is also sufficient for G protein activation. These results reveal that recruitment of GIV to membranes in close proximity to its substrate G protein is a major mechanism responsible for the activation of its G protein regulatory function.

  5. Characterization of an archaeal two-component system that regulates methanogenesis in Methanosaeta harundinacea.

    Directory of Open Access Journals (Sweden)

    Jie Li

    Full Text Available Two-component signal transduction systems (TCSs are a major mechanism used by bacteria in response to environmental changes. Although many sequenced archaeal genomes encode TCSs, they remain poorly understood. Previously, we reported that a methanogenic archaeon, Methanosaeta harundinacea, encodes FilI, which synthesizes carboxyl-acyl homoserine lactones, to regulate transitions of cellular morphology and carbon metabolic fluxes. Here, we report that filI, the cotranscribed filR2, and the adjacent filR1 constitute an archaeal TCS. FilI possesses a cytoplasmic kinase domain (histidine kinase A and histidine kinase-like ATPase and its cognate response regulator. FilR1 carries a receiver (REC domain coupled with an ArsR-related domain with potential DNA-binding ability, while FilR2 carries only a REC domain. In a phosphorelay assay, FilI was autophosphorylated and specifically transferred the phosphoryl group to FilR1 and FilR2, confirming that the three formed a cognate TCS. Through chromatin immunoprecipitation-quantitative polymerase chain reaction (ChIP-qPCR using an anti-FilR1 antibody, FilR1 was shown to form in vivo associations with its own promoter and the promoter of the filI-filR2 operon, demonstrating a regulatory pattern common among TCSs. ChIP-qPCR also detected FilR1 associations with key genes involved in acetoclastic methanogenesis, acs4 and acs1. Electrophoretic mobility shift assays confirmed the in vitro tight binding of FilR1 to its own promoter and those of filI-filR2, acs4, and mtrABC. This also proves the DNA-binding ability of the ArsR-related domain, which is found primarily in Archaea. The archaeal promoters of acs4, filI, acs1, and mtrABC also initiated FilR1-modulated expression in an Escherichia coli lux reporter system, suggesting that FilR1 can up-regulate both archaeal and bacterial transcription. In conclusion, this work identifies an archaeal FilI/FilRs TCS that regulates the methanogenesis of M. harundinacea.

  6. Activator of G-protein signaling 8 is involved in VEGF-mediated signal processing during angiogenesis.

    Science.gov (United States)

    Hayashi, Hisaki; Al Mamun, Abdullah; Sakima, Miho; Sato, Motohiko

    2016-03-15

    Activator of G-protein signaling 8 (AGS8, also known as FNDC1) is a receptor-independent accessory protein for the Gβγ subunit, which was isolated from rat heart subjected to repetitive transient ischemia with the substantial development of collaterals. Here, we report the role of AGS8 in vessel formation by endothelial cells. Knockdown of AGS8 by small interfering RNA (siRNA) inhibited vascular endothelial growth factor (VEGF)-induced tube formation, as well as VEGF-stimulated cell growth and migration. VEGF stimulated the phosphorylation of the VEGF receptor-2 (VEGFR-2, also known as KDR), ERK1/2 and p38 MAPK; however, knockdown of AGS8 inhibited these signaling events. Signal alterations by AGS8 siRNA were associated with a decrease of cell surface VEGFR-2 and an increase of VEGFR-2 in the cytosol. Endocytosis blockers did not influence the decrease of VEGFR-2 by AGS8 siRNA, suggesting the involvement of AGS8 in VEGFR-2 trafficking to the plasma membrane. VEGFR-2 formed a complex with AGS8 in cells, and a peptide designed to disrupt AGS8-Gβγ interaction inhibited VEGF-induced tube formation. These data suggest a potential role for AGS8-Gβγ in VEGF signal processing. AGS8 might play a key role in tissue adaptation by regulating angiogenic events.

  7. A core viral protein binds host nucleosomes to sequester immune danger signals.

    Science.gov (United States)

    Avgousti, Daphne C; Herrmann, Christin; Kulej, Katarzyna; Pancholi, Neha J; Sekulic, Nikolina; Petrescu, Joana; Molden, Rosalynn C; Blumenthal, Daniel; Paris, Andrew J; Reyes, Emigdio D; Ostapchuk, Philomena; Hearing, Patrick; Seeholzer, Steven H; Worthen, G Scott; Black, Ben E; Garcia, Benjamin A; Weitzman, Matthew D

    2016-07-01

    Viral proteins mimic host protein structure and function to redirect cellular processes and subvert innate defenses. Small basic proteins compact and regulate both viral and cellular DNA genomes. Nucleosomes are the repeating units of cellular chromatin and play an important part in innate immune responses. Viral-encoded core basic proteins compact viral genomes, but their impact on host chromatin structure and function remains unexplored. Adenoviruses encode a highly basic protein called protein VII that resembles cellular histones. Although protein VII binds viral DNA and is incorporated with viral genomes into virus particles, it is unknown whether protein VII affects cellular chromatin. Here we show that protein VII alters cellular chromatin, leading us to hypothesize that this has an impact on antiviral responses during adenovirus infection in human cells. We find that protein VII forms complexes with nucleosomes and limits DNA accessibility. We identified post-translational modifications on protein VII that are responsible for chromatin localization. Furthermore, proteomic analysis demonstrated that protein VII is sufficient to alter the protein composition of host chromatin. We found that protein VII is necessary and sufficient for retention in the chromatin of members of the high-mobility-group protein B family (HMGB1, HMGB2 and HMGB3). HMGB1 is actively released in response to inflammatory stimuli and functions as a danger signal to activate immune responses. We showed that protein VII can directly bind HMGB1 in vitro and further demonstrated that protein VII expression in mouse lungs is sufficient to decrease inflammation-induced HMGB1 content and neutrophil recruitment in the bronchoalveolar lavage fluid. Together, our in vitro and in vivo results show that protein VII sequesters HMGB1 and can prevent its release. This study uncovers a viral strategy in which nucleosome binding is exploited to control extracellular immune signaling.

  8. Characterization of the yehUT two-component regulatory system of Salmonella enterica Serovar Typhi and Typhimurium.

    Science.gov (United States)

    Wong, Vanessa K; Pickard, Derek J; Barquist, Lars; Sivaraman, Karthikeyan; Page, Andrew J; Hart, Peter J; Arends, Mark J; Holt, Kathryn E; Kane, Leanne; Mottram, Lynda F; Ellison, Louise; Bautista, Ruben; McGee, Chris J; Kay, Sally J; Wileman, Thomas M; Kenney, Linda J; MacLennan, Calman A; Kingsley, Robert A; Dougan, Gordon

    2013-01-01

    Proteins exhibiting hyper-variable sequences within a bacterial pathogen may be associated with host adaptation. Several lineages of the monophyletic pathogen Salmonella enterica serovar Typhi (S. Typhi) have accumulated non-synonymous mutations in the putative two-component regulatory system yehUT. Consequently we evaluated the function of yehUT in S. Typhi BRD948 and S. Typhimurium ST4/74. Transcriptome analysis identified the cstA gene, encoding a carbon starvation protein as the predominantly yehUT regulated gene in both these serovars. Deletion of yehUT had no detectable effect on the ability of these mutant Salmonella to invade cultured epithelial cells (S. Typhi and S. Typhimurium) or induce colitis in a murine model (S. Typhimurium only). Growth, metabolic and antimicrobial susceptibility tests identified no obvious influences of yehUT on these phenotypes.

  9. The importance of Ca2+/Zn2+ signaling S100 proteins and RAGE in translational medicine.

    Science.gov (United States)

    Leclerc, Estelle; Heizmann, Claus W

    2011-06-01

    The Receptor for Advanced Glycation Endproducts (RAGE) is a multiligand receptor involved in a large number of human disorders. Identified first as the receptor for the Advanced Glycation Endproducts (AGEs), RAGE has emerged in recent years as a major receptor for many members of the S100 calcium and zinc binding protein family. The interaction with and the signaling triggered by several S100 proteins such as S100B and S100A12 have been studied in details and have shown concentration and cell type dependent signaling cascades. The S100 protein family consists of more than 20 members which present high amino-acid sequence and structural similarities. These small EF-hand calcium binding proteins interact with a large number of protein targets and are almost all been shown to be involved in cancer. In this review we discuss the recent knowledge about the role of S100 proteins and RAGE in human disorders.

  10. Exercise and Glycemic Control: Focus on Redox Homeostasis and Redox-Sensitive Protein Signaling

    Science.gov (United States)

    Parker, Lewan; Shaw, Christopher S.; Stepto, Nigel K.; Levinger, Itamar

    2017-01-01

    Physical inactivity, excess energy consumption, and obesity are associated with elevated systemic oxidative stress and the sustained activation of redox-sensitive stress-activated protein kinase (SAPK) and mitogen-activated protein kinase signaling pathways. Sustained SAPK activation leads to aberrant insulin signaling, impaired glycemic control, and the development and progression of cardiometabolic disease. Paradoxically, acute exercise transiently increases oxidative stress and SAPK signaling, yet postexercise glycemic control and skeletal muscle function are enhanced. Furthermore, regular exercise leads to the upregulation of antioxidant defense, which likely assists in the mitigation of chronic oxidative stress-associated disease. In this review, we explore the complex spatiotemporal interplay between exercise, oxidative stress, and glycemic control, and highlight exercise-induced reactive oxygen species and redox-sensitive protein signaling as important regulators of glucose homeostasis. PMID:28529499

  11. The mitogaligin protein is addressed to the nucleus via a non-classical localization signal

    Energy Technology Data Exchange (ETDEWEB)

    Robinet, Pauline; Mollet, Lucile; Gonzalez, Patrick; Normand, Thierry; Charpentier, Stephane; Brule, Fabienne; Dubois, Martine [Centre de Biophysique Moleculaire (Affiliated with the University of Orleans), CNRS UPR4301, Rue Charles Sadron, 45071 Orleans Cedex 2 (France); Legrand, Alain, E-mail: alain.legrand@cnrs-orleans.fr [Centre de Biophysique Moleculaire (Affiliated with the University of Orleans), CNRS UPR4301, Rue Charles Sadron, 45071 Orleans Cedex 2 (France)

    2010-01-29

    Mitogaligin, a protein encoded by galig, an internal cytotoxic gene of the galectin-3 locus, is mostly a mitochondrial protein. Mitochondrial targeting is due to an already identified mitochondrial localization signal. Interaction of mitogaligin with mitochondria leads to cytochrome c cytosolic leakage and ultimately to cell death. We have previously pointed out that mitogaligin can also be directed to the nucleus when the mitochondrial addressing signal is inactivated, indicating a possible dual intracellular localization of the protein. When expressed in the nucleus, mitogaligin exhibits also apoptotic properties leading to cell death. In this report, we show that nuclear addressing of mitogaligin depends on a sequence differing from classical signals containing basic, lysine or proline-tyrosine rich residues. The signal consists of a long sequence of amino acids residues based on a series of a short repetitive degenerated sequence.

  12. The Role of Cgrp-Receptor Component Protein (Rcp) in Cgrp-Mediated Signal Transduction

    OpenAIRE

    Prado, M.A.; B. Evans-Bain; Santi, S. L.; Dickerson, I M

    2001-01-01

    The calcitonin gene-related peptide (CGRP)-receptor component protein (RCP) is a 17-kDa intracellular peripheral membrane protein required for signal transduction at CGRP receptors. To determine the role of RCP in CGRP-mediated signal transduction, RCP was depleted from NIH3T3 cells using antisense strategy. Loss of RCP protein correlated with loss of cAMP production by CGRP in the antisense cells. In contrast, loss of RCP had no effect on CGRP-mediated binding; therefore RCP is not acting as...

  13. G protein signaling governing cell fate decisions involves opposing Gα subunits in Cryptococcus neoformans

    OpenAIRE

    Hsueh, Yen-Ping; Xue, Chaoyang; Heitman, Joseph

    2007-01-01

    Communication between cells and their environments is often mediated by G protein-coupled receptors and cognate G proteins. In fungi, one such signaling cascade is the mating pathway triggered by pheromone/pheromone receptor recognition. Unlike Saccharomyces cerevisiae, which expresses two Gα subunits, most filamentous ascomycetes and basidiomycetes have three Gα subunits. Previous studies have defined the Gα subunit acting upstream of the cAMP-protein kinase A pathway, but it has been unclea...

  14. Novel two-component regulatory systems play a role in biofilm formation of Lactobacillus reuteri rodent isolate 100-23.

    Science.gov (United States)

    Su, Marcia Shu-Wei; Gänzle, Michael G

    2014-04-01

    This study characterized the two-component regulatory systems encoded by bfrKRT and cemAKR, and assessed their influence on biofilm formation by Lactobacillus reuteri 100-23. A method for deletion of multiple genes was employed to disrupt the genetic loci of two-component systems. The operons bfrKRT and cemAKR showed complementary organization. Genes bfrKRT encode a histidine kinase, a response regulator and an ATP-binding cassette-type transporter with a bacteriocin-processing peptidase domain, respectively. Genes cemAKR code for a signal peptide, a histidine kinase and a response regulator, respectively. Deletion of single or multiple genes in the operons bfrKRT and cemAKR did not affect cell morphology, growth or the sensitivity to various stressors. However, gene disruption affected biofilm formation; this effect was dependent on the carbon source. Deletion of bfrK or cemA increased sucrose-dependent biofilm formation in vitro. Glucose-dependent biofilm formation was particularly increased by deletion of cemK. The expression of cemK and cemR was altered by deletion of bfrK, indicating cross-talk between these two regulatory systems. These results may contribute to our understanding of the genetic factors related to the biofilm formation and competitiveness of L. reuteri in intestinal ecosystems.

  15. Analysis of the BarA/UvrY two-component system in Shewanella oneidensis MR-1.

    Directory of Open Access Journals (Sweden)

    Lucas Binnenkade

    Full Text Available The BarA/UvrY two-component system is well conserved in species of the γ-proteobacteria and regulates numerous processes predominantly by controlling the expression of a subset of noncoding small RNAs. In this study, we identified and characterized the BarA/UvrY two-component system in the gammaproteobacterium Shewanella oneidensis MR-1. Functional interaction of sensor kinase BarA and the cognate response regulator UvrY was indicated by in vitro phosphotransfer studies. The expression of two predicted small regulatory RNAs (sRNAs, CsrB1 and CsrB2, was dependent on UvrY. Transcriptomic analysis by microarrays revealed that UvrY is a global regulator and directly or indirectly affects transcript levels of more than 200 genes in S. oneidensis. Among these are genes encoding key enzymes of central carbon metabolism such as ackA, aceAB, and pflAB. As predicted of a signal transduction pathway that controls aspects of central metabolism, mutants lacking UvrY reach a significantly higher OD than the wild type during aerobic growth on N-acetylglucosamine (NAG while under anaerobic conditions the mutant grew more slowly. A shorter lag phase occurred with lactate as carbon source. In contrast, significant growth phenotypes were absent in complex medium. Based on these studies we hypothesize that, in S. oneidensis MR-1, the global BarA/UvrY/Csr regulatory pathway is involved in central carbon metabolism processes.

  16. The Cbln Family of Proteins Interact with Multiple Signaling Pathways

    OpenAIRE

    Wei, Peng; Pattarini, Roberto; Rong, Yongqi; Guo, Hong; Bansal, Parmil K; Kusnoor, Sheila V; Deutch, Ariel Y.; Parris, Jennifer; Morgan, James I.

    2012-01-01

    Cbln1 is essential for synapse integrity