Transcriptional regulation of the waaAE-coaD operon by PhoP and RcsAB in Yersinia pestis biovar Microtus.
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ABSTRACT:
SUBMITTER: Liu L
PROVIDER: S-EPMC4259886 | biostudies-literature |
REPOSITORIES: biostudies-literature
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Project description:BackgroundThe transcription regulator PhoP has been shown to be important for Y. pestis survival in macrophages and under various in vitro stresses. However, the mechanism by which PhoP promotes bacterial intracellular survival is not fully understood. Our previous microarray analysis suggested that PhoP governed a wide set of cellular pathways in Y. pestis. A series of biochemical experiments were done herein to study members of the PhoP regulon of Y. pestis biovar Microtus.ResultsBy using gel mobility shift assay and quantitative RT-PCR, a total of 30 putative transcription units were characterized as direct PhoP targets. The primer extension assay was further used to determine the transcription start sites of 18 PhoP-dependent promoters and to localize the -10 and -35 elements. The DNase I footprinting was used to identify the PhoP-binding sites within 17 PhoP-dependent promoters, enabling the identification of PhoP box and matrix that both represented the conserved signals for PhoP recognition in Y. pestis. Data presented here providing a good basis for modeling PhoP-promoter DNA interactions that is crucial to the PhoP-mediated transcriptional regulation.ConclusionThe proven direct PhoP targets include nine genes encoding regulators and 21 genes or operons with functions of detoxification, protection against DNA damages, resistance to antimicrobial peptides, and adaptation to magnesium limitation. We can presume that PhoP is a global regulator that controls a complex regulatory cascade by a mechanism of not only directly controlling the expression of specific genes, but also indirectly regulating various cellular pathways by acting on a set of dedicated regulators. These results help us gain insights into the PhoP-dependent mechanisms by which Y. pestis survives the antibacterial strategies employed by host macrophages.
| S-EPMC2322996 | biostudies-literature
Project description:BACKGROUND: Yersinia pestis is the causative agent of plague. The two transcriptional regulators, PhoP and RovA, are required for the virulence of Y. pestis through the regulation of various virulence-associated loci. They are the global regulators controlling two distinct large complexes of cellular pathways. METHODOLOGY/PRINCIPAL FINDINGS: Based on the LacZ fusion, primer extension, gel mobility shift, and DNase I footprinting assays, RovA is shown to recognize both of the two promoters of its gene in Y. pestis. The autoregulation of RovA appears to be a conserved mechanism shared by Y. pestis and its closely related progenitor, Y. pseudotuberculosis. In Y. pestis, the PhoP regulator responds to low magnesium signals and then negatively controls only one of the two promoters of rovA through PhoP-promoter DNA association. CONCLUSIONS/SIGNIFICANCE: RovA is a direct transcriptional activator for its own gene in Y. pestis, while PhoP recognizes the promoter region of rovA to repress its transcription. The direct regulatory association between PhoP and RovA bridges the PhoP and RovA regulons in Y. pestis.
| S-EPMC3180457 | biostudies-literature
Project description:Yersinia pestis has been historically divided into three biovars: antiqua, mediaevalis, and orientalis. On the basis of this study, strains from Microtus-related plague foci are proposed to constitute a new biovar, microtus. Based on the ability to ferment glycerol and arabinose and to reduce nitrate, Y. pestis strains can be assigned to one of four biovars: antiqua (glycerol positive, arabinose positive, and nitrate positive), mediaevalis (glycerol positive, arabinose positive, and nitrate negative), orientalis (glycerol negative, arabinose positive, and nitrate positive), and microtus (glycerol positive, arabinose negative, and nitrate negative). A 93-bp in-frame deletion in glpD gene results in the glycerol-negative characteristic of biovar orientalis strains. Two kinds of point mutations in the napA gene may cause the nitrate reduction-negative characteristic in biovars mediaevalis and microtus, respectively. A 122-bp frameshift deletion in the araC gene may lead to the arabinose-negative phenotype of biovar microtus strains. Biovar microtus strains have a unique genomic profile of gene loss and pseudogene distribution, which most likely accounts for the human attenuation of this new biovar. Focused, hypothesis-based investigations on these specific genes will help delineate the determinants that enable this deadly pathogen to be virulent to humans and give insight into the evolution of Y. pestis and plague pathogenesis. Moreover, there may be the implications for development of biovar microtus strains as a potential vaccine.
| S-EPMC451627 | biostudies-literature
Project description:Yersinia pestis is the etiological agent of the notorious plague that has claimed millions of deaths in history. Of the four known Y. pestis biovars (Antiqua, Medievalis, Orientalis, and Microtus), Microtus strains are unique for being highly virulent in mice but avirulent in humans. Here, human peripheral lymphocytes were infected with the fully virulent 141 strain or the Microtus strain 201, and their transcriptomes were determined and compared. The most notable finding was that robust responses in the pathways for cytokine-cytokine receptor interaction, chemokine signaling pathway, Toll-like receptor signaling and Jak-STAT signaling were induced at 2 h post infection (hpi) in the 201- but not the 141-infected lymphocytes, suggesting that human lymphocytes might be able to constrain infections caused by strain 201 but not 141. Consistent with the transcriptome results, much higher IFN-γ and IL-1β were present in the supernatants from the 201-infected lymphocytes, while inflammatory inhibitory IL-10 levels were higher in the 141-infected lymphocytes. The expressions of CSTD and SLC11A1, both of which are functional components of the lysosome, increased in the 201-infected human macrophage-like U937 cells. Further assessment of the survival rate of the 201 bacilli in the U937 cells and murine macrophage RAW 264.7 cells revealed no viable bacteria in the U937 cells at 32 hpi.; however, about 5-10% of the bacteria were still alive in the RAW264.7 cells. Our results indicate that human macrophages can clear the intracellular Y. pestis 201 bacilli more efficiently than murine macrophages, probably by interfering with critical host immune responses, and this could partially account for the host-specific pathogenicity of Y. pestis Microtus strains.
| S-EPMC6491462 | biostudies-literature
Project description:Extremely virulent organism that causes plague
| PRJNA10638 | ENA
Project description:Biofilm formation is critical for blocking flea foregut and hence for transmission of Y. pestis by flea biting. In this study, we identified the regulatory role of the AraC-family transcriptional regulator BfvR (YPO1737 in strain CO92) in biofilm formation and virulence of Yersinia pestis biovar Microtus. Crystal violet staining, Caenorhabditis elegans biofilm assay, colony morphology assay, intracellular c-di-GMP concentration determination, and BALB/c mice challenge were employed to reveal that BfvR enhanced Y. pestis biofilm formation while repressed its virulence in mice. Further molecular biological assays demonstrated that BfvR directly stimulated the expression of hmsHFRS, waaAE-coaD, and hmsCDE, which, in turn, affected the production of exopolysaccharide, LPS, and c-di-GMP, respectively. In addition, BfvR directly and indirectly repressed psaABC and psaEF transcription, respectively. We concluded that the modulation of biofilm- and virulence-related genes by BfvR led to increased biofilm formation and reduced virulence of Y. pestis biovar Microtus.
| S-EPMC6176095 | biostudies-literature
Project description:We have investigated the physical and genetic structure and regulation of the Yersinia pestis yscBCDEF region, previously called lcrC. DNA sequence analysis showed that this region is homologous to the corresponding part of the ysc locus of Yersinia enterocolitica and suggested that the yscBCDEF cistrons belong to a single operon on the low-calcium response virulence plasmid pCD1. Promoter activity measurements of ysc subclones indicated that yscBCDEF constitutes a suboperon of the larger ysc region by revealing promoter activity in a clone containing the 3' end of yscD, intact yscE and yscF, and part of yscG. These experiments also revealed an additional weak promoter upstream of yscD. Northern (RNA) analysis with a yscD probe showed that operon transcription is thermally induced and downregulated in the presence of Ca2+. Primer extension of operon transcripts suggested that two promoters, a moderate-level constitutive one and a stronger, calcium-downregulated one, control full-length operon transcription at 37 degrees C. Primer extension provided additional support for the proposed designation of a yscBCDEF suboperon by identifying a 5' end within yscF, for which relative abundances in the presence and absence of Ca2+ revealed regulation that is distinct from that for transcripts initiating farther upstream. YscB and YscC were expressed in Escherichia coli by using a high-level transcription system. Attempts to express YscD were only partially successful, but they revealed interesting regulation at the translational level.
| S-EPMC206281 | biostudies-other
Project description:The cyclic AMP receptor protein (CRP) is a bacterial regulator that controls more than 100 promoters, including those involved in catabolite repression. In the present study, a null deletion of the crp gene was constructed for Yersinia pestis bv. microtus strain 201. Microarray expression analysis disclosed that at least 6% of Y. pestis genes were affected by this mutation. Further reverse transcription-PCR and electrophoretic mobility shift assay analyses disclosed a set of 37 genes or putative operons to be the direct targets of CRP, and thus they constitute the minimal CRP regulon in Y. pestis. Subsequent primer extension and DNase I footprinting assays mapped transcriptional start sites, core promoter elements, and CRP binding sites within the DNA regions upstream of pla and pst, revealing positive and direct control of these two laterally acquired plasmid genes by CRP. The crp disruption affected both in vitro and in vivo growth of the mutant and led to a >15,000-fold loss of virulence after subcutaneous infection but a <40-fold increase in the 50% lethal dose by intravenous inoculation. Therefore, CRP is required for the virulence of Y. pestis and, particularly, is more important for infection by subcutaneous inoculation. It can further be concluded that the reduced in vivo growth phenotype of the crp mutant should contribute, at least partially, to its attenuation of virulence by both routes of infection. Consistent with a previous study of Y. pestis bv. medievalis, lacZ reporter fusion analysis indicated that the crp deletion resulted in the almost absolute loss of pla promoter activity. The plasminogen activator encoded by pla was previously shown to specifically promote Y. pestis dissemination from peripheral infection routes (subcutaneous infection [flea bite] or inhalation). The above evidence supports the notion that in addition to the reduced in vivo growth phenotype, the defect of pla expression in the crp mutant will greatly contribute to the huge loss of virulence of this mutant strain in subcutaneous infection.
| S-EPMC2573370 | biostudies-literature
Project description:Yersinia pestis, the agent of plague, forms a biofilm in its flea vector to enhance transmission. Y. pestis biofilm development is positively regulated by hmsT and hmsD, encoding diguanylate cyclases (DGCs) involved in synthesis of the bacterial second messenger c-di-GMP. rcsA, encoding an auxiliary protein in Rcs phosphorelay, is nonfunctional in Y. pestis, while in Yersinia pseudotuberculosis, rcsA is functional and represses biofilms. Previously we showed that Rcs phosphorelay negatively regulates transcription of hmsT in Y. pestis and its ancestor Yersinia pseudotuberculosis. In this study, we show that Rcs positively regulates hmsCDE operon (encoding HmsD) in Y. pestis; while in the presence of functional rcsA, Rcs represses hmsCDE operon in Y. pseudotuberculosis. Loss of rcsA's function in Y. pestis not only causes derepression of hmsT but also causes activation of hmsD, which may account for the increased biofilm formation in Y. pestis. In addition, differential regulation of the two DGCs, HmsT and HmsD by Rcs may help Y. pestis to adapt to different environment.
| S-EPMC4325325 | biostudies-literature
Project description:human peripheral lymphocytes were infected with the full virulent strain 141 or human avirulent Microtus strain 201, and their transcriptomes were determined and compared. The most impressive finding is that robust responses of lysosome, TLR signaling and FcR mediated phagocytosis pathways were provoked at 2 hpi in 201- but not in 141-infected lymphocytes, suggesting that human lymphocytes might be able to constrain infection caused by strain 201 but not 141.
2019-05-15 | GSE121084 | GEO