scholarly journals Hybrid Two-Component Sensors for Identification of Bacterial Chemoreceptor Function

2019 ◽  
Vol 85 (22) ◽  
Author(s):  
Rita A. Luu ◽  
Rebecca A. Schomer ◽  
Ceanne N. Brunton ◽  
Richard Truong ◽  
Albert P. Ta ◽  
...  
Keyword(s):  
Gene Expression ◽  
Soil Bacteria ◽  
Tricarboxylic Acid Cycle ◽  
Chemical Stimulus ◽  
Content Type ◽  
E Coli ◽  
Wide Range ◽  
Component Systems ◽  
Two Component ◽  
Two Component Systems

ABSTRACT Soil bacteria adapt to diverse and rapidly changing environmental conditions by sensing and responding to environmental cues using a variety of sensory systems. Two-component systems are a widespread type of signal transduction system present in all three domains of life and typically are comprised of a sensor kinase and a response regulator. Many two-component systems function by regulating gene expression in response to environmental stimuli. The bacterial chemotaxis system is a modified two-component system with additional protein components and a response that, rather than regulating gene expression, involves behavioral adaptation and results in net movement toward or away from a chemical stimulus. Soil bacteria generally have 20 to 40 or more chemoreceptors encoded in their genomes. To simplify the identification of chemoeffectors (ligands) sensed by bacterial chemoreceptors, we constructed hybrid sensor proteins by fusing the sensor domains of Pseudomonas putida chemoreceptors to the signaling domains of the Escherichia coli NarX/NarQ nitrate sensors. Responses to potential attractants were monitored by β-galactosidase assays using an E. coli reporter strain in which the nitrate-responsive narG promoter was fused to lacZ. Hybrid receptors constructed from PcaY, McfR, and NahY, which are chemoreceptors for aromatic acids, tricarboxylic acid cycle intermediates, and naphthalene, respectively, were sensitive and specific for detecting known attractants, and the β-galactosidase activities measured in E. coli correlated well with results of chemotaxis assays in the native P. putida strain. In addition, a screen of the hybrid receptors successfully identified new ligands for chemoreceptor proteins and resulted in the identification of six receptors that detect propionate. IMPORTANCE Relatively few of the thousands of chemoreceptors encoded in bacterial genomes have been functionally characterized. More importantly, although methyl-accepting chemotaxis proteins, the major type of chemoreceptors present in bacteria, are easily identified bioinformatically, it is not currently possible to predict what chemicals will bind to a particular chemoreceptor. Chemotaxis is known to play roles in biodegradation as well as in host-pathogen and host-symbiont interactions, but many studies are currently limited by the inability to identify relevant chemoreceptor ligands. The use of hybrid receptors and this simple E. coli reporter system allowed rapid and sensitive screening for potential chemoeffectors. The fusion site chosen for this study resulted in a high percentage of functional hybrids, indicating that it could be used to broadly test chemoreceptor responses from phylogenetically diverse samples. Considering the wide range of chemical attractants detected by soil bacteria, hybrid receptors may also be useful as sensitive biosensors.

scholarly journals Elucidation of Regulatory Modes for Five Two-Component Systems in Escherichia coli Reveals Novel Relationships

mSystems ◽  
2020 ◽  
Vol 5 (6) ◽  
Author(s):  
Kumari Sonal Choudhary ◽  
Julia A. Kleinmanns ◽  
Katherine Decker ◽  
Anand V. Sastry ◽  
Ye Gao ◽  
...  
Keyword(s):  
Gene Expression ◽  
Escherichia Coli ◽  
Target Gene ◽  
Target Genes ◽  
Rna Seq ◽  
Content Type ◽  
E Coli ◽  
Component Systems ◽  
Two Component ◽  
Two Component Systems

ABSTRACT Escherichia coli uses two-component systems (TCSs) to respond to environmental signals. TCSs affect gene expression and are parts of E. coli’s global transcriptional regulatory network (TRN). Here, we identified the regulons of five TCSs in E. coli MG1655: BaeSR and CpxAR, which were stimulated by ethanol stress; KdpDE and PhoRB, induced by limiting potassium and phosphate, respectively; and ZraSR, stimulated by zinc. We analyzed RNA-seq data using independent component analysis (ICA). ChIP-exo data were used to validate condition-specific target gene binding sites. Based on these data, we do the following: (i) identify the target genes for each TCS; (ii) show how the target genes are transcribed in response to stimulus; and (iii) reveal novel relationships between TCSs, which indicate noncognate inducers for various response regulators, such as BaeR to iron starvation, CpxR to phosphate limitation, and PhoB and ZraR to cell envelope stress. Our understanding of the TRN in E. coli is thus notably expanded. IMPORTANCE E. coli is a common commensal microbe found in the human gut microenvironment; however, some strains cause diseases like diarrhea, urinary tract infections, and meningitis. E. coli’s two-component systems (TCSs) modulate target gene expression, especially related to virulence, pathogenesis, and antimicrobial peptides, in response to environmental stimuli. Thus, it is of utmost importance to understand the transcriptional regulation of TCSs to infer bacterial environmental adaptation and disease pathogenicity. Utilizing a combinatorial approach integrating RNA sequencing (RNA-seq), independent component analysis, chromatin immunoprecipitation coupled with exonuclease treatment (ChIP-exo), and data mining, we suggest five different modes of TCS transcriptional regulation. Our data further highlight noncognate inducers of TCSs, which emphasizes the cross-regulatory nature of TCSs in E. coli and suggests that TCSs may have a role beyond their cognate functionalities. In summary, these results can lead to an understanding of the metabolic capabilities of bacteria and correctly predict complex phenotype under diverse conditions, especially when further incorporated with genome-scale metabolic models.

scholarly journals Elucidation of regulatory modes for five two-component systems in Escherichia coli reveals novel relationships

2020 ◽  
Author(s):  
Kumari Sonal Choudhary ◽  
Julia A. Kleinmanns ◽  
Katherine Decker ◽  
Anand V Sastry ◽  
Ye Gao ◽  
...  
Keyword(s):  
Gene Expression ◽  
Escherichia Coli ◽  
Transcriptional Regulation ◽  
Independent Component Analysis ◽  
Target Gene ◽  
Target Genes ◽  
E Coli ◽  
Component Systems ◽  
Two Component ◽  
Two Component Systems

AbstractEscherichia coli uses two-component systems (TCSs) to respond to environmental signals. TCSs affect gene expression and are parts of E. coli’s global transcriptional regulatory network (TRN). Here, we identified the regulons of five TCSs in E. coli MG1655: BaeSR and CpxAR, which were stimulated by ethanol stress; KdpDE and PhoRB, induced by limiting potassium and phosphate, respectively; and ZraSR, stimulated by zinc. We analyzed RNA-seq data using independent component analysis (ICA). ChIP-exo data was used to validate condition-specific target gene binding sites. Based on this data we (1) identify the target genes for each TCS; (2) show how the target genes are transcribed in response to stimulus; and (3) reveal novel relationships between TCSs, which indicate non-cognate inducers for various response regulators, such as BaeR to iron starvation, CpxR to phosphate limitation, and PhoB and ZraR to cell envelope stress. Our understanding of the TRN in E. coli is thus notably expanded.ImportanceE. coli is a common commensal microbe found in human gut microenvironment; however, some strains cause diseases like diarrhea, urinary tract infections and meningitis. E. coli’s two-component system (TCS) modulates target gene expression, specially related to virulence, pathogenesis and anti-microbial peptides, in response to environmental stimuli. Thus, it is of utmost importance to understand the transcriptional regulation of the TCSs to infer its environmental adaptation and disease pathogenicity. Utilizing a combinatorial approach integrating RNAseq, independent component analysis, ChIP-exo and data mining, we show that TCSs have five different modes of transcriptional regulation. Our data further highlights non-cognate inducers of TCSs emphasizing cross-regulatory nature of TCSs in E. coli and suggests that TCSs may have a role beyond their cognate functionalities. In summary, these results when further incorporated with genome scale metabolic models can lead to understanding of metabolic capabilities of bacteria and correctly predict complex phenotype under diverse conditions.

scholarly journals Cross-regulation between RpfG and HtsHs to regulate antibiotic biosynthesis in Lysobacter

2020 ◽  
Author(s):  
Kaihuai Li ◽  
Gaoge Xu ◽  
Bo Wang ◽  
Guichun Wu ◽  
Fengquan Liu
Keyword(s):  
Gene Expression ◽  
Environmental Changes ◽  
Antibiotic Biosynthesis ◽  
Two Component System ◽  
Independent Manner ◽  
Heat Stable ◽  
Component Systems ◽  
Two Component ◽  
Two Component Systems ◽  
Sense And Respond

AbstractBacterial two-component systems (TCSs) sense and respond to environmental changes and modulate downstream gene expression. However, the mechanism of cross-talk between multiple TCSs is unclear. In this study, we report a previously uncharacterized mechanism by which the TCS protein RpfG interacts with hybrid two-component system (HyTCS) proteins HtsH1, HtsH2 and HtsH3 to regulate antibiotic biosynthesis in Lysobacter. RpfG, a phosphodiesterase (PDE), can degrade c-di-GMP to 5’-pGpG and can regulate antibiotic heat-stable antifungal factor (HSAF) biosynthesis in a PDE- independent manner. Thus, we wondered whether RpfG regulate HSAF biosynthesis through interactions with other factors. Subsequently, we demonstrated that RpfG interacts with three HyTCS proteins (HtsH1, HtsH2 and HtsH3), that can inhibit the PDE enzymatic activity of RpfG. Importantly, deletion of htsH1, htsH2 and htsH3 resulted in significantly decreased HSAF production, and we showed that HtsH1, HtsH2 and HtsH3 depend on their phosphorylation activity to directly regulate HSAF biosynthesis gene expression. Our results reveal that RpfG does not depend on PDE activity to regulate HSAF biosynthesis, rather it interacts with HtsH1, HtsH2 and HtsH3 to do so, a regulatory mechanism that may be a conserved paradigm in Lysobacter and Xanthomonas.

scholarly journals Effects of Regulatory Network Organization and Environment on PmrD Connector Activity and Polymyxin Resistance in Klebsiella pneumoniae and Escherichia coli

2020 ◽  
Vol 65 (3) ◽  
Author(s):  
Annie I. Chen ◽  
Francisco Javier Albicoro ◽  
Jun Zhu ◽  
Mark Goulian
Keyword(s):  
Escherichia Coli ◽  
Klebsiella Pneumoniae ◽  
Critical Role ◽  
Negative Regulator ◽  
Network Organization ◽  
Content Type ◽  
Resistance Pathway ◽  
Component Systems ◽  
Two Component ◽  
Two Component Systems

ABSTRACT Polymyxins are a class of cyclic peptides with antimicrobial activity against Gram-negative bacteria. In Enterobacteriaceae, the PhoQ/PhoP and PmrB/PmrA two-component systems regulate many genes that confer resistance to both polymyxins and host antimicrobial peptides. The activities of these two-component systems are modulated by additional proteins that are conserved across Enterobacteriaceae, such as MgrB, a negative regulator of PhoQ, and PmrD, a “connector” protein that activates PmrB/PmrA in response to PhoQ/PhoP stimulation. Despite the conservation of many protein components of the PhoQ/PhoP-PmrD-PmrB/PmrA network, the specific molecular interactions and regulatory mechanisms vary across different genera. Here, we explore the role of PmrD in modulating this signaling network in Klebsiella pneumoniae and Escherichia coli. We show that in K. pneumoniae, PmrD is not required for polymyxin resistance arising from mutation of mgrB—the most common cause of spontaneous polymyxin resistance in this bacterium—suggesting that direct activation of polymyxin resistance genes by PhoQ/PhoP plays a critical role in this resistance pathway. However, for conditions of low pH or intermediate iron concentrations, both of which stimulate PmrB/PmrA, we find that PmrD does contribute to resistance. We further show that in E. coli, PmrD functions as a connector between PhoQ/PhoP and PmrB/PmrA, in contrast with previous reports. In this case, activity also depends on PmrB/PmrA stimulation, or on very high activation of PhoQ/PhoP. Our results indicate that the importance of the PmrD connector in modulating the polymyxin resistance network depends on both the network organization and on the environmental conditions associated with PmrB stimulation.

scholarly journals Multiple Two-Component Systems of Streptococcus mutans Regulate Agmatine Deiminase Gene Expression and Stress Tolerance

2009 ◽  
Vol 191 (23) ◽  
pp. 7363-7366 ◽  
Author(s):  
Yaling Liu ◽  
Robert A. Burne
Keyword(s):  
Gene Expression ◽  
Stress Tolerance ◽  
Streptococcus Mutans ◽  
Thermal Stresses ◽  
Low Ph ◽  
Component Systems ◽  
Two Component ◽  
Two Component Systems ◽  
Agmatine Deiminase

ABSTRACT Induction of the agmatine deiminase system (AgDS) of Streptococcus mutans requires agmatine and is optimal at low pH. We show here that the VicRK, ComDE, and CiaRH two-component systems influence AgDS gene expression in response to acidic and thermal stresses.

scholarly journals The ChrSA and HrrSA Two-Component Systems Are Required for Transcriptional Regulation of thehemAPromoter in Corynebacterium diphtheriae

2016 ◽  
Vol 198 (18) ◽  
pp. 2419-2430 ◽  
Author(s):  
Jonathan M. Burgos ◽  
Michael P. Schmitt
Keyword(s):  
Signal Transduction ◽  
Kinase Activity ◽  
Content Type ◽  
Component Systems ◽  
Two Component ◽  
Two Component Signal Transduction ◽  
Two Component Systems ◽  
Signal Transduction Systems

ABSTRACTCorynebacterium diphtheriaeutilizes heme and hemoglobin (Hb) as iron sources for growth in low-iron environments. InC. diphtheriae, the two-component signal transduction systems (TCSs) ChrSA and HrrSA are responsive to Hb levels and regulate the transcription of promoters forhmuO,hrtAB, andhemA. ChrSA and HrrSA activate transcription at thehmuOpromoter and repress transcription athemAin an Hb-dependent manner. In this study, we show that HrrSA is the predominant repressor athemAand that its activity results in transcriptional repression in the presence and absence of Hb, whereas repression ofhemAby ChrSA is primarily responsive to Hb. DNA binding studies showed that both ChrA and HrrA bind to thehemApromoter region at virtually identical sequences. ChrA binding was enhanced by phosphorylation, while binding to DNA by HrrA was independent of its phosphorylation state. ChrA and HrrA are phosphorylatedin vitroby the sensor kinase ChrS, whereas no kinase activity was observed with HrrSin vitro. Phosphorylated ChrA was not observedin vivo, even in the presence of Hb, which is likely due to the instability of the phosphate moiety on ChrA. However, phosphorylation of HrrA was observedin vivoregardless of the presence of the Hb inducer, and genetic analysis indicates that ChrS is responsible for most of the phosphorylation of HrrAin vivo. Phosphorylation studies strongly suggest that HrrS functions primarily as a phosphatase and has only minimal kinase activity. These findings collectively show a complex mechanism of regulation at thehemApromoter, where both two-component systems act in concert to optimize expression of heme biosynthetic enzymes.IMPORTANCEUnderstanding the mechanism by which two-component signal transduction systems function to respond to environmental stimuli is critical to the study of bacterial pathogenesis. The current study expands on the previous analyses of the ChrSA and HrrSA TCSs in the human pathogenC. diphtheriae. The findings here underscore the complex interactions between the ChrSA and HrrSA systems in the regulation of thehemApromoter and demonstrate how the two systems complement one another to refine and control transcription in the presence and absence of Hb.

scholarly journals Chloroplast two-component systems: evolution of the link between photosynthesis and gene expression

2009 ◽  
Vol 276 (1665) ◽  
pp. 2133-2145 ◽  
Author(s):  
Sujith Puthiyaveetil ◽  
John F. Allen
Keyword(s):  
Gene Expression ◽  
Genomic Analysis ◽  
Mendelian Inheritance ◽  
Comparative Genomic ◽  
Photosynthetic Eukaryotes ◽  
Specific Distribution ◽  
Component Systems ◽  
Two Component ◽  
Sensor Kinases ◽  
Two Component Systems

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 inherited from an ancestral cyanobacterial symbiont are no longer present in chloroplasts. Recent research now shows that two-component systems have survived in chloroplasts as products of both chloroplast and nuclear genes. Comparative genomic analysis of photosynthetic eukaryotes shows a lineage-specific distribution of chloroplast two-component systems. The components and the systems they comprise have homologues in extant cyanobacterial lineages, indicating their ancient cyanobacterial origin. Sequence and functional characteristics of chloroplast two-component systems point to their fundamental role in linking photosynthesis with gene expression. We propose that two-component systems provide a coupling between photosynthesis and gene expression that serves to retain genes in chloroplasts, thus providing the basis of cytoplasmic, non-Mendelian inheritance of plastid-associated characters. We discuss the role of this coupling in the chronobiology of cells and in the dialogue between nuclear and cytoplasmic genetic systems.

Age replacement policies for two-component systems with stochastic dependence

2015 ◽  
Vol 21 (3) ◽  
pp. 346-357 ◽  
Author(s):  
Zouheir Malki ◽  
Daoud Ait-Kadi ◽  
Mohamed-Salah Ouali
Keyword(s):  
Random Variable ◽  
Replacement Policy ◽  
Replacement Cost ◽  
Stochastic Dependence ◽  
Content Type ◽  
Preventive Replacement ◽  
Component Systems ◽  
Two Component ◽  
Two Component Systems ◽  
Age Replacement

Purpose – The purpose of this paper is to investigate age replacement policies for two-component parallel system with stochastic dependence. The stochastic dependence considered, is modeled by a one-sided domino effect. The failure of component 1 at instant t may induce the failure of component 2 at instant t+τ with probability p 1→2. The time delay τ is a random variable with known probability density function h p 1→2 (.). The system is considered in a failed state when both components are failed. The proposed replacement policies suggest to replace the system upon failure or at age T whichever occurs first. Design/methodology/approach – In the first policy, costs and durations associated with maintenance activities are supposed to be constant. In the second replacement policy, the preventive replacement cost depends on the system’s state and age. The expected cost per unit of time over an infinite span is derived and numerical examples are presented. Findings – In this paper and especially in the second policy, the authors find that the authors can get a more economical policy if the authors consider that the preventive replacement cost is not constant but depends on T. Originality/value – In this paper, the authors take into account of the stochastic dependence between system components. This dependence affects the global reliability of the system and replacement’s periodicity. It can be used to measure the performance of the system et introduced into design phase of the system.

scholarly journals Menaquinone Analogs Inhibit Growth of Bacterial Pathogens

2013 ◽  
Vol 57 (11) ◽  
pp. 5432-5437 ◽  
Author(s):  
Patrick M. Schlievert ◽  
Joseph A. Merriman ◽  
Wilmara Salgado-Pabón ◽  
Elizabeth A. Mueller ◽  
Adam R. Spaulding ◽  
...  
Keyword(s):  
Plasma Membranes ◽  
Rabbit Model ◽  
Two Component System ◽  
Content Type ◽  
Gram Positive Bacteria ◽  
Component Systems ◽  
Two Component ◽  
Topical Antibacterial ◽  
Two Component Systems ◽  
Antibacterial Target

ABSTRACTGram-positive bacteria cause serious human illnesses through combinations of cell surface and secreted virulence factors. We initiated studies with four of these organisms to develop novel topical antibacterial agents that interfere with growth and exotoxin production, focusing on menaquinone analogs. Menadione, 1,4-naphthoquinone, and coenzymes Q1 to Q3 but not menaquinone, phylloquinone, or coenzyme Q10 inhibited the growth and to a greater extent exotoxin production ofStaphylococcus aureus,Bacillus anthracis,Streptococcus pyogenes, andStreptococcus agalactiaeat concentrations of 10 to 200 μg/ml. Coenzyme Q1 reduced the ability ofS. aureusto cause toxic shock syndrome in a rabbit model, inhibited the growth of four Gram-negative bacteria, and synergized with another antimicrobial agent, glycerol monolaurate, to inhibitS. aureusgrowth. The staphylococcal two-component system SrrA/B was shown to be an antibacterial target of coenzyme Q1. We hypothesize that menaquinone analogs both induce toxic reactive oxygen species and affect bacterial plasma membranes and biosynthetic machinery to interfere with two-component systems, respiration, and macromolecular synthesis. These compounds represent a novel class of potential topical therapeutic agents.

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