Exploring the mode of action of inhibitors targeting the PhoP response regulator of Salmonella enterica through comprehensive pharmacophore approaches.
Ontology highlight
ABSTRACT:
SUBMITTER: Tsai KC
PROVIDER: S-EPMC9062048 | biostudies-literature | 2019 Mar
REPOSITORIES: biostudies-literature
Publications
Exploring the mode of action of inhibitors targeting the PhoP response regulator of <i>Salmonella enterica</i> through comprehensive pharmacophore approaches.
Tsai Keng-Chang KC Hung Po-Pin PP Cheng Ching-Feng CF Chen Chinpan C Tseng Tien-Sheng TS Tseng Tien-Sheng TS
RSC advances 20190321 16
The PhoQ/PhoP two-component system regulates the physiological and virulence functions of <i>Salmonella enterica</i>. However, the mode of action of known PhoP inhibitors is unclear. We systematically constructed a pharmacophore model of inhibitors to probe the interface pharmacophore model of the PhoP dimer, coupling it with Ligplot analysis. We found that these inhibitors bind on the α5-helix, altering the conformation and interfering with PhoP binding on DNA. ...[more]
Similar Datasets
Project description:The PhoP-PhoQ two-component system is necessary for the virulence of Salmonella spp. and is responsible for regulating several modifications of the lipopolysaccharide (LPS). Mutagenesis of the transcriptional regulator phoP resulted in the identification of a mutant able to activate transcription of regulated genes approximately 100-fold in the absence of PhoQ. Sequence analysis showed two single-base alterations resulting in amino acid changes at positions 93 (S93N) and 203 (Q203R). These mutations were individually created, and although each resulted in a constitutive phenotype, the double mutant displayed a synergistic effect both in the induction of PhoP-activated gene expression and in resistance to antimicrobial peptides. The constitutive phoP gene was placed under the control of an arabinose-inducible promoter to examine the kinetics of PhoP-activated gene induction and the resultant modifications of LPS. Gene induction and 2-hydroxymyristate modification of the lipid A were shown to occur within minutes of the addition of arabinose and to peak at 4 h. As the first constitutive mutant of phoP identified, this allele will be invaluable to future genetic and biochemical studies of this and likely other regulatory systems.
| S-EPMC97674 | biostudies-literature
Project description:Two-component signal transduction (TCST) is the predominant signaling scheme used in bacteria to sense and respond to environmental changes in order to survive and thrive. A typical TCST system consists of a sensor histidine kinase to detect external signals and an effector response regulator to respond to external changes. In the signaling scheme, the histidine kinase phosphorylates and activates the response regulator, which functions as a transcription factor to modulate gene expression. One promising strategy toward antibacterial development is to target TCST regulatory systems, specifically the response regulators to disrupt the expression of genes important for virulence. In Salmonella enterica, the PhoQ/PhoP signal transduction system is used to sense and respond to low magnesium levels and regulates the expression for over 40 genes necessary for growth under these conditions, and more interestingly, genes that are important for virulence. In this study, a hybrid approach coupling computational and experimental methods was applied to identify drug-like compounds to target the PhoP response regulator. A computational approach of structure-based virtual screening combined with a series of biochemical and biophysical assays was used to test the predictability of the computational strategy and to characterize the mode of action of the compounds. Eight compounds from virtual screening inhibit the formation of the PhoP-DNA complex necessary for virulence gene regulation. This investigation served as an initial case study for targeting TCST response regulators to modulate the gene expression of a signal transduction pathway important for bacterial virulence. With the increasing resistance of pathogenic bacteria to current antibiotics, targeting TCST response regulators that control virulence is a viable strategy for the development of antimicrobial therapeutics with novel modes of action.
| S-EPMC3445336 | biostudies-literature
Project description:Genetic and genomic approaches have been successfully used to assign genes to distinct regulatory networks. However, the present challenge of distinguishing differentially regulated genes within a network is particularly hard because members of a given network tend to have similar regulatory features. We have addressed this challenge by developing a method, termed Gene Promoter Scan, that discriminates coregulated promoters by simultaneously considering both multiple cis promoter features and gene expression. Here, we apply this method to probe the regulatory networks governed by the PhoP/PhoQ two-component system in the enteric bacteria Escherichia coli and Salmonella enterica. Our analysis uncovered members of the PhoP regulon and interactions with other regulatory systems that were not discovered in previous approaches. The predictions made by Gene Promoter Scan were experimentally validated to establish that the PhoP protein uses multiple mechanisms to control gene transcription, regulates acid resistance determinants, and is a central element in a highly connected network.
| S-EPMC548500 | biostudies-literature
Project description:Infection with Salmonella enterica serovar Typhi in humans causes the life-threatening disease typhoid fever. In the laboratory, typhoid fever can be modeled through the inoculation of susceptible mice with Salmonella enterica serovar Typhimurium. Using this murine model, we previously characterized the interactions between Salmonella Typhimurium and host cells in the gallbladder and showed that this pathogen can successfully invade gallbladder epithelial cells and proliferate. Additionally, we showed that Salmonella Typhimurium can use bile phospholipids to grow at high rates. These abilities are likely important for quick colonization of the gallbladder during typhoid fever and further pathogen dissemination through fecal shedding. To further characterize the interactions between Salmonella and the gallbladder environment, we compared the transcriptomes of Salmonella cultures grown in LB broth or physiological murine bile. Our data showed that many genes involved in bacterial central metabolism are affected by bile, with the citric acid cycle being repressed and alternative respiratory systems being activated. Additionally, our study revealed a new aspect of Salmonella interactions with bile through the identification of the global regulator phoP as a bile-responsive gene. Repression of phoP expression could also be achieved using physiological, but not commercial, bovine bile. The biological activity does not involve PhoPQ sensing of a bile component and is not caused by bile acids, the most abundant organic components of bile. Bioactivity-guided purification allowed the identification of a subset of small molecules from bile that can elicit full activity; however, a single compound with phoP inhibitory activity could not be isolated, suggesting that multiple molecules may act in synergy to achieve this effect. Due to the critical role of phoP in Salmonella virulence, further studies in this area will likely reveal aspects of the interaction between Salmonella and bile that are relevant to disease.
| S-EPMC3347055 | biostudies-literature
Project description:Salmonella typhimurium is a facultative intracellular pathogen capable of surviving within host phagocytic cells. Salmonella strains carrying phoP mutations are avirulent, unable to survive in macrophages, and extremely sensitive to peptides having antimicrobial activity such as the host-derived defensins. We present here the DNA sequence of the phoP gene and show that the deduced amino acid sequence of phoP has extensive homology with the Escherichia coli transcriptional regulators PhoB and OmpR, which control the expression of loci in response to different environmental stimuli. The psiD locus, which is regulated by phosphate availability, was found to be under the control of the phoP gene product. Sequences homologous to phoP were found in several Gram-negative species and in the yeast Saccharomyces cerevisiae.
| S-EPMC297997 | biostudies-other
Project description:The PhoP/PhoQ two-component system controls the extracellular magnesium deprivation response in Salmonella enterica. In addition, several virulence-associated genes that are mainly required for intramacrophage survival during the infection process are under the control of its transcriptional regulation. Despite shared Mg(2+) modulation of the expression of the PhoP-activated genes, no consensus sequence common to all of them could be detected in their promoter regions. We have investigated the transcriptional regulation and the interaction of the response regulator PhoP with the promoter regions of the PhoP-activated loci phoPQ, mgtA, slyB, pmrD, pcgL, phoN, pagC, and mgtCB. A direct repeat of the heptanucleotide sequence (G/T)GTTTA(A/T) was identified as the conserved motif recognized by PhoP to directly control the gene expression of the first five loci, among which the first four are ancestral to enterobacteria. On the other hand, no direct interaction of the response regulator with the promoter of phoN, pagC, or mgtCB was apparent by either in vitro or in vivo assays. These loci are Salmonella specific and were probably acquired by horizontal DNA transfer. Besides, sequence analysis of pag promoters revealed the presence of a conserved PhoP box in 6 out of the 12 genes analyzed. Our results strongly suggest that the expression of a set of Mg(2+)-controlled genes is driven by PhoP via unknown intermediate regulatory mechanisms that could also involve ancillary factors.
| S-EPMC219391 | biostudies-other
Project description:BACKGROUND:S. Typhi, a human-restricted Salmonella enterica serovar, causes a systemic intracellular infection in humans (typhoid fever). In comparison, S. Typhimurium causes gastroenteritis in humans, but causes a systemic typhoidal illness in mice. The PhoP regulon is a well studied two component (PhoP/Q) coordinately regulated network of genes whose expression is required for intracellular survival of S. enterica. METHODOLOGY/PRINCIPAL FINDINGS:Using high performance liquid chromatography mass spectrometry (HPLC-MS/MS), we examined the protein expression profiles of three sequenced S. enterica strains: S. Typhimurium LT2, S. Typhi CT18, and S. Typhi Ty2 in PhoP-inducing and non-inducing conditions in vitro and compared these results to profiles of phoP(-)/Q(-) mutants derived from S. Typhimurium LT2 and S. Typhi Ty2. Our analysis identified 53 proteins in S. Typhimurium LT2 and 56 proteins in S. Typhi that were regulated in a PhoP-dependent manner. As expected, many proteins identified in S. Typhi demonstrated concordant differential expression with a homologous protein in S. Typhimurium. However, three proteins (HlyE, STY1499, and CdtB) had no homolog in S. Typhimurium. HlyE is a pore-forming toxin. STY1499 encodes a stably expressed protein of unknown function transcribed in the same operon as HlyE. CdtB is a cytolethal distending toxin associated with DNA damage, cell cycle arrest, and cellular distension. Gene expression studies confirmed up-regulation of mRNA of HlyE, STY1499, and CdtB in S. Typhi in PhoP-inducing conditions. CONCLUSIONS/SIGNIFICANCE:This study is the first protein expression study of the PhoP virulence associated regulon using strains of Salmonella mutant in PhoP, has identified three Typhi-unique proteins (CdtB, HlyE and STY1499) that are not present in the genome of the wide host-range Typhimurium, and includes the first protein expression profiling of a live attenuated bacterial vaccine studied in humans (Ty800).
| S-EPMC2736619 | biostudies-literature
Project description:Salmonella must rapidly adapt to various niches in the host during infection. Relevant virulence factors must be appropriately induced, and systems that are detrimental in a particular environment must be turned off. Salmonella infects intestinal epithelial cells using a type 3 secretion system (T3SS) encoded on Salmonella pathogenicity island 1 (SPI1). The system is controlled by three AraC-like regulators, HilD, HilC, and RtsA, which form a complex feed-forward loop to activate expression of hilA, encoding the main transcriptional regulator of T3SS structural genes. This system is tightly regulated, with many of the activating signals acting at the level of hilD translation or HilD protein activity. Once inside the phagosomes of epithelial cells, or in macrophages during systemic stages of disease, the SPI1 T3SS is no longer required or expressed. Here, we show that the PhoPQ two-component system, critical for intracellular survival, appears to be the primary mechanism by which Salmonella shuts down the SPI1 T3SS. PhoP negatively regulates hilA through multiple distinct mechanisms: direct transcriptional repression of the hilA promoter, indirect transcriptional repression of both the hilD and rtsA promoters, and activation of the small RNA (sRNA) PinT. Genetic analyses and electrophoretic mobility shift assays suggest that PhoP specifically binds the hilA promoter to block binding of activators HilD, HilC, and RtsA as a mechanism of repression.IMPORTANCESalmonella is one of the most common foodborne pathogens, causing an estimated 1.2 million illnesses per year in the United States. A key step in infection is the activation of the bacterial invasion machinery, which induces uptake of the bacterium into epithelial cells and leads to induction of inflammatory diarrhea. Upon entering the vacuolar compartments of host cells, Salmonella senses an environmental transition and represses the invasion machinery with a two-component system relevant for survival within the vacuole. This adaptation to specific host niches is an important example of how signals are integrated for survival of the pathogen.
| S-EPMC6657598 | biostudies-literature
Project description:The sigma factor RpoS regulates the expression of many stress response genes and is required for virulence in several bacterial species. We now report that RpoS accumulates when Salmonella enterica serovar Typhimurium is growing logarithmically in media with low Mg(2+) concentrations. This process requires the two-component regulatory system PhoP/PhoQ, which is specifically activated in low Mg(2+). We show that PhoP controls RpoS protein turnover by serving as a transcriptional activator of the iraP (yaiB) gene, which encodes a product that enhances RpoS stability by interacting with RssB, the protein that normally delivers RpoS to the ClpXP protease for degradation. Mutation of the phoP gene rendered Salmonella as sensitive to hydrogen peroxide as an rpoS mutant after growth in low Mg(2+). In Escherichia coli, low Mg(2+) leads to only modest RpoS stabilization, and iraP is not regulated by PhoP/PhoQ. These findings add the sigma factor RpoS to the regulatory proteins and two-component systems that are elevated in a PhoP/PhoQ-dependent fashion when Salmonella face low Mg(2+) environments. Our data also exemplify the critical differences in regulatory circuits that exist between the closely related enteric bacteria Salmonella and E. coli.
| S-EPMC1557385 | biostudies-literature
Project description:The response regulator SsrB activates expression of genes encoded within and outside of a pathogenicity island (SPI-2), which is required for systemic infection of Salmonella. SsrB binds upstream of the sifA, sifB, and sseJ effector genes and directly regulates transcription. SsrB also relieves gene silencing by the nucleoid protein H-NS. Single molecule experiments with magnetic tweezers demonstrated that SsrB displaces H-NS from DNA only when it is bound in a polymerization (stiffening) mode and not when H-NS is bound to DNA in the bridging mode. Thus, in contrast to previous views, the polymerization binding mode of H-NS is the relevant form for counter-silencing by SsrB. Our results reveal that response regulators can directly activate transcription and also relieve H-NS silencing. This study adds to the repertoire of mechanisms by which NarL/FixJ subfamily members regulate transcription. Because SsrB-dependent promoters are diversely organized, additional mechanisms of transcriptional activation at other loci are likely.
| S-EPMC3023485 | biostudies-literature