scholarly journals Systematic Reconstruction of the Complete Two-Component Sensorial Network in Staphylococcus aureus

mSystems ◽  
2020 ◽  
Vol 5 (4) ◽  
Author(s):  
B. Rapun-Araiz ◽  
A. F. Haag ◽  
V. De Cesare ◽  
C. Gil ◽  
P. Dorado-Morales ◽  
...  
Keyword(s):  
Gene Expression ◽  
Target Genes ◽  
Response Regulator ◽  
Active Form ◽  
Gain Of Function ◽  
Content Type ◽  
The Family ◽  
Two Component ◽  
Function Strategy ◽  
Two Component Signal Transduction

ABSTRACT In bacteria, adaptation to changes in the environment is mainly controlled through two-component signal transduction systems (TCSs). Most bacteria contain dozens of TCSs, each of them responsible for sensing a different range of signals and controlling the expression of a repertoire of target genes (regulon). Over the years, identification of the regulon controlled by each individual TCS in different bacteria has been a recurrent question. However, limitations associated with the classical approaches used have left our knowledge far from complete. In this report, using a pioneering approach in which a strain devoid of the complete nonessential TCS network was systematically complemented with the constitutively active form of each response regulator, we have reconstituted the regulon of each TCS of S. aureus in the absence of interference between members of the family. Transcriptome sequencing (RNA-Seq) and proteomics allowed us to determine the size, complexity, and insulation of each regulon and to identify the genes regulated exclusively by one or many TCSs. This gain-of-function strategy provides the first description of the complete TCS regulon in a living cell, which we expect will be useful to understand the pathobiology of this important pathogen. IMPORTANCE Bacteria are able to sense environmental conditions and respond accordingly. Their sensorial system relies on pairs of sensory and regulatory proteins, known as two-component systems (TCSs). The majority of bacteria contain dozens of TCSs, each of them responsible for sensing and responding to a different range of signals. Traditionally, the function of each TCS has been determined by analyzing the changes in gene expression caused by the absence of individual TCSs. Here, we used a bacterial strain deprived of the complete TC sensorial system to introduce, one by one, the active form of every TCS. This gain-of-function strategy allowed us to identify the changes in gene expression conferred by each TCS without interference of other members of the family.

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

2017 ◽  
Vol 199 (18) ◽  
Author(s):  
Jennifer K. Teschler ◽  
Andrew T. Cheng ◽  
Fitnat H. Yildiz
Keyword(s):  
Signal Transduction ◽  
Vibrio Cholerae ◽  
Histidine Kinase ◽  
Biofilm Formation ◽  
Pathogenic Bacteria ◽  
Response Regulator ◽  
Systematic Analysis ◽  
Content Type ◽  
Two Component ◽  
Two Component Signal Transduction

ABSTRACT 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. cholerae. vxrB 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 understanding of the role that TCSs play in the regulation of V. cholerae biofilm formation.

scholarly journals Development and Validation of a High-Throughput Cell-Based Screen To Identify Activators of a Bacterial Two-Component Signal Transduction System

2015 ◽  
Vol 59 (7) ◽  
pp. 3789-3799 ◽  
Author(s):  
Julia J. van Rensburg ◽  
Kate R. Fortney ◽  
Lan Chen ◽  
Andrew J. Krieger ◽  
Bruno P. Lima ◽  
...  
Keyword(s):  
Signal Transduction ◽  
High Throughput ◽  
Response Regulator ◽  
Phosphoryl Group ◽  
Signal Transduction System ◽  
Membrane Repair ◽  
Content Type ◽  
Two Component ◽  
Serovar Typhimurium ◽  
Two Component Signal Transduction

ABSTRACTCpxRA 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 inSalmonella entericaserovar Typhimurium andHaemophilus ducreyiare avirulent in mice and humans, respectively. Thus, the activation of CpxRA has high potential as a novel antimicrobial/antivirulence strategy. Using a series ofEscherichia colistrains containing a CpxR-P-responsivelacZreporter 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.

scholarly journals Identification of a Two-Component Signal Transduction System fromCorynebacterium diphtheriae That Activates Gene Expression in Response to the Presence of Heme and Hemoglobin

1999 ◽  
Vol 181 (17) ◽  
pp. 5330-5340 ◽  
Author(s):  
Michael P. Schmitt
Keyword(s):  
Gene Expression ◽  
Signal Transduction ◽  
Response Regulator ◽  
Open Reading Frames ◽  
Sensor Kinase ◽  
Dependent Manner ◽  
Signal Transduction System ◽  
Kinase Gene ◽  
Two Component ◽  
Two Component Signal Transduction

ABSTRACT Corynebacterium diphtheriae, the causative agent of diphtheria, utilizes various host compounds to acquire iron. TheC. diphtheriae hmuO gene encodes a heme oxygenase that is involved in the utilization of heme and hemoglobin as iron sources. Transcription of the hmuO gene in C. diphtheriae is controlled under a dual regulatory mechanism in which the diphtheria toxin repressor protein (DtxR) and iron repress expression while either heme or hemoglobin is needed to activate transcription. In this study, two clones isolated from a C. diphtheriae chromosomal library were shown to activate transcription from the hmuO promoter in Escherichia coli. Sequence analysis revealed that these activator clones each carried distinct genes whose products had significant homology to response regulators of two-component signal transduction systems. Located upstream from each of these response regulator homologs are partial open reading frames that are predicted to encode the C-terminal portions of sensor kinases. The full-length sensor kinase gene for each of these systems was cloned from the C. diphtheriaechromosome, and constructs each carrying one complete sensor kinase gene and its cognate response regulator were constructed. One of these constructs, pTSB20, which carried the response regulator (chrA) and its cognate sensor kinase (chrS), was shown to strongly activate transcription from the hmuOpromoter in a heme-dependent manner in E. coli. A mutation in chrA (chrAD50N), which changed a conserved aspartic acid residue at position 50, the presumed site of phosphorylation by ChrS, to an asparagine, abolished heme-dependent activation. These findings suggest that the sensor kinase ChrS is involved in the detection of heme and the transduction of this signal, via a phosphotransfer mechanism, to the response regulator ChrA, which then activates transcription of the hmuO promoter. This is the first report of a bacterial two-component signal transduction system that controls gene expression through a heme-responsive mechanism.

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

2015 ◽  
Vol 198 (3) ◽  
pp. 377-385 ◽  
Author(s):  
Zaara Sarwar ◽  
Anthony G. Garza
Keyword(s):  
Gene Expression ◽  
Myxococcus Xanthus ◽  
Fruiting Bodies ◽  
Content Type ◽  
Body Development ◽  
Two Component ◽  
Fruiting Body Development ◽  
Two Component Signal Transduction ◽  
Temporal And Spatial ◽  
Sporulation Genes

When starved for nutrients,Myxococcus xanthusproduces 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 ofM. xanthussporulation 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.

scholarly journals Histidine Phosphotransfer Proteins in Fungal Two-Component Signal Transduction Pathways

2013 ◽  
Vol 12 (8) ◽  
pp. 1052-1060 ◽  
Author(s):  
Jan S. Fassler ◽  
Ann H. West
Keyword(s):  
Signal Transduction ◽  
Response Regulator ◽  
Single Gene ◽  
Signal Transduction Pathway ◽  
Pathogenic Fungi ◽  
Kinase Gene ◽  
Content Type ◽  
Two Component ◽  
Two Component Signal Transduction ◽  
Histidine Phosphotransfer

ABSTRACTThe histidine phosphotransfer (HPt) protein Ypd1 is an important participant in theSaccharomyces cerevisiaemultistep 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 bothS. cerevisiaeand inCryptococcus neoformans. These and other aspects of Ypd1 biology, combined with the availability of structural and mutational data inS. 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 onS. cerevisiaeYpd1 and to consider the potential benefits of conducting related studies in pathogenic fungi.

scholarly journals Transcription Analysis of a Lantibiotic Gene Cluster from Bifidobacterium longum DJO10A

2011 ◽  
Vol 77 (17) ◽  
pp. 5879-5887 ◽  
Author(s):  
Ju-Hoon Lee ◽  
Xiulan Li ◽  
Daniel J. O'Sullivan
Keyword(s):  
Gene Expression ◽  
Response Regulator ◽  
Bifidobacterium Longum ◽  
Regulatory System ◽  
Broth Culture ◽  
Northern Hybridization ◽  
Transcription Start ◽  
Transcription Analysis ◽  
Content Type ◽  
Two Component

ABSTRACTBifidobacterium longumDJO10A was previously demonstrated to produce a lantibiotic, but only during growth on agar media. To evaluate the feasibility of production of this lantibiotic in broth media, a transcription analysis of thelanAgene was undertaken. Comparative microarray analysis of broth and agar cultures ofB. longumDJO10A revealed that the lantibiotic production, modification, transport/peptidase, and immunity genes were significantly upregulated in agar cultures, while the two-component regulatory genes were expressed equally under both conditions. This suggested that the signal transduction regulatory system should function in broth cultures. Real-time PCR and Northern hybridization confirmed thatlanAgene expression was significantly repressed in broth cultures. A crude lantibiotic preparation from an agar-grown culture was obtained, and its antimicrobial spectrum analysis revealed a broad inhibition range. Addition of this extract to broth cultures ofB. longumDJO10A inducedlanAgene expression in a dose-dependent fashion. Subinoculation using >10% of an induced broth culture maintainedlanAexpression. The expression oflanAwas log-phase specific, being significantly downregulated in stationary phase. Transcription start analysis oflanArevealed a 284-bp 5′ untranslated region, which was proposed to be involved in repression of transcription, while an inverted repeat structure located at bp −75 relative to the transcription start was strategically located to likely function as a binding site for the two-component response regulator. Understanding the transcription regulation of thislanAgene is the first step toward enabling production of this novel and potentially interesting lantibiotic in broth cultures.

scholarly journals Orphan Hybrid Histidine Protein Kinase SinK Acts as a Signal Integrator To Fine-Tune Multicellular Behavior inMyxococcus xanthus

2019 ◽  
Vol 201 (6) ◽  
Author(s):  
Maike M. Glaser ◽  
Penelope I. Higgs
Keyword(s):  
Environmental Conditions ◽  
Histidine Kinase ◽  
Response Regulator ◽  
Myxococcus Xanthus ◽  
Histidine Kinases ◽  
Content Type ◽  
Environmental Bacterium ◽  
Two Component ◽  
Two Component Signal Transduction ◽  
Fine Tune

ABSTRACTHis-Asp phosphorelay (also known as two-component signal transduction) proteins are the predominant mechanism used in most bacteria to control behavior in response to changing environmental conditions. In addition to systems consisting of a simple two-component system utilizing an isolated histidine kinase/response regulator pair, some bacteria are enriched in histidine kinases that serve as signal integration proteins; these kinases are usually characterized by noncanonical domain architecture, and the responses that they regulate may be difficult to identify. The environmental bacteriumMyxococcus xanthusis highly enriched in these noncanonical histidine kinases.M. xanthusis renowned for a starvation-induced multicellular developmental program in which some cells are induced to aggregate into fruiting bodies and then differentiate into environmentally resistant spores. Here, we characterize theM. xanthusorphan hybrid histidine kinase SinK (Mxan_4465), which consists of a histidine kinase transmitter followed by two receiver domains (REC1and REC2). NonphosphorylatablesinKmutants were analyzed under two distinct developmental conditions and using a new high-resolution developmental assay. These assays revealed that SinK autophosphorylation and REC1impact the onset of aggregation and/or the mobility of aggregates, while REC2impacts sporulation efficiency. SinK activity is controlled by a genus-specific hypothetical protein (SinM; Mxan_4466). We propose that SinK serves to fine-tune fruiting body morphology in response to environmental conditions.IMPORTANCEBiofilms are multicellular communities of microorganisms that play important roles in host disease or environmental biofouling. Design of preventative strategies to block biofilms depends on understanding the molecular mechanisms used by microorganisms to build them. The production of biofilms in bacteria often involves two-component signal transduction systems in which one protein component (a kinase) detects an environmental signal and, through phosphotransfer, activates a second protein component (a response regulator) to change the transcription of genes necessary to produce a biofilm. We show that an atypical kinase, SinK, modulates several distinct stages of specialized biofilm produced by the environmental bacteriumMyxococcus xanthus. SinK likely integrates multiple signals to fine-tune biofilm formation in response to distinct environmental conditions.

scholarly journals Regulation of Virulence by Two-Component Systems in Pathogenic Burkholderia

2020 ◽  
Vol 88 (7) ◽  
Author(s):  
Matthew M. Schaefers
Keyword(s):  
Histidine Kinase ◽  
Burkholderia Cepacia ◽  
Target Genes ◽  
Response Regulator ◽  
Pharmaceutical Products ◽  
Phosphoryl Group ◽  
Content Type ◽  
Component Systems ◽  
Two Component ◽  
Two Component Systems

ABSTRACT The regulation and timely expression of bacterial genes during infection is critical for a pathogen to cause an infection. Bacteria have multiple mechanisms to regulate gene expression in response to their environment, one of which is two-component systems (TCS). TCS have two components. One component is a sensory histidine kinase (HK) that autophosphorylates when activated by a signal. The activated sensory histidine kinase then transfers the phosphoryl group to the second component, the response regulator, which activates transcription of target genes. The genus Burkholderia contains members that cause human disease and are often extensively resistant to many antibiotics. The Burkholderia cepacia complex (BCC) can cause severe lung infections in patients with cystic fibrosis (CF) or chronic granulomatous disease (CGD). BCC members have also recently been associated with several outbreaks of bacteremia from contaminated pharmaceutical products. Separate from the BCC is Burkholderia pseudomallei, which is the causative agent of melioidosis, a serious disease that occurs in the tropics, and a potential bioterrorism weapon. Bioinformatic analysis of sequenced Burkholderia isolates predicts that most strains have at least 40 TCS. The vast majority of these TCS are uncharacterized both in terms of the signals that activate them and the genes that are regulated by them. This review will highlight TCS that have been described to play a role in virulence in either the BCC or B. pseudomallei. Since many of these TCS are involved in virulence, TCS are potential novel therapeutic targets, and elucidating their function is critical for understanding Burkholderia pathogenesis.

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