Characterization of Zur-dependent genes and direct Zur targets in Yersinia pestis

<p>Abstract</p> <p>Background</p> <p>The zinc uptake regulator Zur is a Zn²⁺-sensing metalloregulatory protein involved in the maintenance of bacterial zinc homeostasis. Up to now, regulation of zinc homeostasis by Zur is poorly understood in <it>Y. pestis</it>.</p> <p>Results</p> <p>We constructed a <it>zur </it>null mutant of <it>Y. pestis </it>biovar <it>microtus </it>strain 201. Microarray expression analysis disclosed a set of 154 Zur-dependent genes of <it>Y. pestis </it>upon exposure to zinc rich condition. Real-time reverse transcription (RT)-PCR was subsequently used to validate the microarray data. Based on the 154 Zur-dependent genes, predicted regulatory Zur motifs were used to screen for potential direct Zur targets including three putative operons <it>znuA, znuCB </it>and <it>ykgM</it>-<it>RpmJ2</it>. The LacZ reporter fusion analysis verified that Zur greatly repressed the promoter activity of the above three operons. The subsequent electrophoretic mobility shift assay (EMSA) demonstrated that a purified Zur protein was able to bind to the promoter regions of the above three operons. The DNase I footprinting was used to identify the Zur binding sites for the above three operons, verifying the Zur box sequence as predicted previously in γ-Proteobacteria. The primer extension assay was further used to determine the transcription start sites for the above three operons and to localize the -10 and -35 elements. Zur binding sites overlapped the -10 sequence of its target promoters, which was consistent with the previous observation that Zur binding would block the entry of the RNA polymerase to repress the transcription of its target genes.</p> <p>Conclusion</p> <p>Zur as a repressor directly controls the transcription of <it>znuA, znuCB </it>and <it>ykgM</it>-<it>RpmJ2 </it>in <it>Y. pestis </it>by employing a conserved mechanism of Zur-promoter DNA association as observed in γ-Proteobacteria. Zur contributes to zinc homeostasis in <it>Y. pestis </it>likely through transcriptional repression of the high-affinity zinc uptake system ZnuACB and two alternative ribosomal proteins YkgM and RpmJ2.</p

Genome Characterization and Potential Risk Assessment of the Novel SARS-CoV-2 Variant Omicron (B.1.1.529)

As the novel coronavirus SARS-CoV-2 spread around the world, multiple waves of variants emerged, thus leading to local or global population shifts during the pandemic. A new variant named Omicron (PANGO lineage B.1.1.529), which was first discovered in southern Africa, has recently been proposed by the World Health Organization to be a Variant of Concern. This variant carries an unusually large number of mutations, particularly on the spike protein and receptor binding domain, in contrast to other known major variants. Some mutation sites are associated with enhanced viral transmission, infectivity, and pathogenicity, thus enabling the virus to evade the immune protective barrier. Given that the emergence of the Omicron variant was accompanied by a sharp increase in infection cases in South Africa, the variant has the potential to trigger a new global epidemic peak. Therefore, continual attention and a rapid response are required to decrease the possible risks to public health

Identification and characterization of PhoP regulon members in Yersinia pestis biovar Microtus

<p>Abstract</p> <p>Background</p> <p>The transcription regulator PhoP has been shown to be important for <it>Y. pestis </it>survival in macrophages and under various <it>in vitro </it>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 <it>Y. pestis</it>. A series of biochemical experiments were done herein to study members of the PhoP regulon of <it>Y. pestis </it>biovar <it>Microtus</it>.</p> <p>Results</p> <p>By 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 <it>Y. pestis</it>. Data presented here providing a good basis for modeling PhoP-promoter DNA interactions that is crucial to the PhoP-mediated transcriptional regulation.</p> <p>Conclusion</p> <p>The 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 <it>Y. pestis </it>survives the antibacterial strategies employed by host macrophages.</p

Yersinia pestis and Plague: Some Knowns and Unknowns

Since its first identification in 1894 during the third pandemic in Hong Kong, there has been significant progress in understanding the lifestyle of Yersinia pestis , the pathogen that is responsible for plague. Although we now have some understanding of the pathogen’s physiology, genetics, genomics, evolution, gene regulation, pathogenesis and immunity, there are many unknown aspects of the pathogen and its disease development. Here, we focus on some of the knowns and unknowns related to Y. pestis and plague. We notably focus on some key Y. pestis physiologic and virulence traits that are important for its mammal-flea-mammal life cycle, but also its emergence from the enteropathogen, Yersinia pseudotuberculosis . Some aspects of the genetic diversity of Y. pestis , the distribution and ecology of plague, as well as the medical countermeasures to protect our population are also provided. Lastly, we present some biosafety and biosecurity information related to Y. pestis and plague

Cell Membrane Is Impaired, Accompanied by Enhanced Type III Secretion System Expression in Yersinia pestis Deficient in RovA Regulator

BACKGROUND: In the enteropathogenic Yersinia species, RovA regulates the expression of invasin, which is important for enteropathogenic pathogenesis but is inactivated in Yersinia pestis. Investigation of the RovA regulon in Y. pestis at 26 °C has revealed that RovA is a global regulator that contributes to virulence in part by the direct regulation of psaEFABC. However, the regulatory roles of RovA in Y. pestis at 37 °C, which allows most virulence factors in mammalian hosts to be expressed, are still poorly understood. METHODOLOGY/PRINCIPAL FINDINGS: The transcriptional profile of an in-frame rovA mutant of Y. pestis biovar Microtus strain 201 was analyzed under type III secretion system (T3SS) induction conditions using microarray techniques, and it was revealed that many cell-envelope and transport/binding proteins were differentially expressed in the ΔrovA mutant. Most noticeably, many of the T3SS genes, including operons encoding the translocon, needle and Yop (Yersinia outer protein) effectors, were significantly up-regulated. Analysis of Yop proteins confirmed that YopE and YopJ were also expressed in greater amounts in the mutant. However, electrophoresis mobility shift assay results demonstrated that the His-RovA protein could not bind to the promoter sequences of the T3SS genes, suggesting that an indirect regulatory mechanism is involved. Transmission electron microscopy analysis indicated that there are small loose electron dense particle-like structures that surround the outer membrane of the mutant cells. The bacterial membrane permeability to CFSE (carboxyfluorescein diacetate succinimidyl ester) was significantly decreased in the ΔrovA mutant compared to the wild-type strain. Taken together, these results revealed the improper construction and dysfunction of the membrane in the ΔrovA mutant. CONCLUSIONS/SIGNIFICANCE: We demonstrated that the RovA regulator plays critical roles in the construction and functioning of the bacterial membrane, which sheds considerable light on the regulatory functions of RovA in antibiotic resistance and environmental adaptation. The expression of T3SS was upregulated in the ΔrovA mutant through an indirect regulatory mechanism, which is possibly related to the altered membrane construction in the mutant

Characterization of an aspartate aminotransferase encoded by YPO0623 with frequent nonsense mutations in Yersinia pestis

Yersinia pestis, the causative agent of plague, is a genetically monomorphic bacterial pathogen that evolved from Yersinia pseudotuberculosis approximately 7,400 years ago. We observed unusually frequent mutations in Y. pestis YPO0623, mostly resulting in protein translation termination, which implies a strong natural selection. These mutations were found in all phylogenetic lineages of Y. pestis, and there was no apparent pattern in the spatial distribution of the mutant strains. Based on these findings, we aimed to investigate the biological function of YPO0623 and the reasons for its frequent mutation in Y. pestis. Our in vitro and in vivo assays revealed that the deletion of YPO0623 enhanced the growth of Y. pestis in nutrient-rich environments and led to increased tolerance to heat and cold shocks. With RNA-seq analysis, we also discovered that the deletion of YPO0623 resulted in the upregulation of genes associated with the type VI secretion system (T6SS) at 26°C, which probably plays a crucial role in the response of Y. pestis to environment fluctuations. Furthermore, bioinformatic analysis showed that YPO0623 has high homology with a PLP-dependent aspartate aminotransferase in Salmonella enterica, and the enzyme activity assays confirmed its aspartate aminotransferase activity. However, the enzyme activity of YPO0623 was significantly lower than that in other bacteria. These observations provide some insights into the underlying reasons for the high-frequency nonsense mutations in YPO0623, and further investigations are needed to determine the exact mechanism

An Overdispersed Black-Box Variational Bayesian–Kalman Filter with Inaccurate Noise Second-Order Statistics

Aimed at the problems in which the performance of filters derived from a hypothetical model will decline or the cost of time of the filters derived from a posterior model will increase when prior knowledge and second-order statistics of noise are uncertain, a new filter is proposed. In this paper, a Bayesian robust Kalman filter based on posterior noise statistics (KFPNS) is derived, and the recursive equations of this filter are very similar to that of the classical algorithm. Note that the posterior noise distributions are approximated by overdispersed black-box variational inference (O-BBVI). More precisely, we introduce an overdispersed distribution to push more probability density to the tails of variational distribution and incorporated the idea of importance sampling into two strategies of control variates and Rao–Blackwellization in order to reduce the variance of estimators. As a result, the convergence process will speed up. From the simulations, we can observe that the proposed filter has good performance for the model with uncertain noise. Moreover, we verify the proposed algorithm by using a practical multiple-input multiple-output (MIMO) radar system