Host transcriptome sequencing of FACS-sorted human induced pluripotent stem cell-derived brain endothelial cells infected with Neisseria meningitidis
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ABSTRACT:
ORGANISM(S): Homo sapiens Neisseria meningitidis MC58
PROVIDER: GSE126449 | GEO | 2019/02/13
REPOSITORIES: GEO
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Project description:Neisseria meningitidis is a Gram-negative bacterium that asymptomatically colonises the nasopharynx of humans. For an unknown reason, N. meningitidis can cross the nasopharyngeal barrier and invade the bloodstream where it becomes one of the most harmful extracellular bacterial pathogen. This infectious cycle involves the colonisation of two different environments. (a) In the nasopharynx, N. meningitidis grow on the top of mucus-producing epithelial cells surrounded by a complex microbiota. To survive and grow in this challenging environment, the meningococcus expresses specific virulence factors such as polymorphic toxins and MDA?. (b) Meningococci have the ability to survive in the extra cellular fluids including blood and cerebrospinal fluid. The interaction of N. meningitidis with human endothelial cells leads to the formation of typical microcolonies that extend overtime and promote vascular injury, disseminated intravascular coagulation, and acute inflammation. In this review, we will focus on the interplay between N. meningitidis and these two different niches at the cellular and molecular level and discuss the use of inhibitors of piliation as a potent therapeutic approach.
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Project description:The human-specific, Gram-negative bacterium Neisseria meningitidis (Nm) is a leading cause of bacterial meningitis world-wide. It has been described that Nm can enter the central nervous system via the blood-cerebrospinal fluid barrier (BCSFB), which is constituted by the epithelial cells of the choroid plexus. Using a recently established in vitro model of the human BCSFB based on human malignant choroid plexus papilloma (HIBCPP) cells we investigated the cellular response of HIBCPP cells challenged with the meningitis-causing Nm strain MC58. In comparison we analysed the answer to the closely related unencapsulated carrier isolate Nm α14. Transcriptome analysis revealed a stronger transcriptional response after infection with strain MC58, in particular with its capsule deficient mutant MC58siaD-, which correlated with bacterial invasion levels. Expression evaluation and Gene Set Enrichment Analysis pointed to a NF-κB-mediated pro-inflammatory immune response involving up-regulation of the transcription factor IκBζ. Consistent with this, infected cells secreted significant levels of pro-inflammatory chemokines and cytokines, among others, IL8, CXCL1-3 and the IκBζ target gene product IL6. Expression profile of pattern recognition receptors in HIBCPP cells and the response to specific agonists indicates that TLR2 rather than TLR4 is involved in the cellular reaction following Nm infection.
2014-09-16 | GSE42870 | GEO
Project description:The complement system is an essential component of the innate and acquired immune system, and consists of a series of proteolytic cascades that are initiated by the presence of microorganisms. In health, activation of complement is precisely controlled through membrane-bound and soluble plasma-regulatory proteins including complement factor H (fH; ref. 2), a 155 kDa protein composed of 20 domains (termed complement control protein repeats). Many pathogens have evolved the ability to avoid immune-killing by recruiting host complement regulators and several pathogens have adapted to avoid complement-mediated killing by sequestering fH to their surface. Here we present the structure of a complement regulator in complex with its pathogen surface-protein ligand. This reveals how the important human pathogen Neisseria meningitidis subverts immune responses by mimicking the host, using protein instead of charged-carbohydrate chemistry to recruit the host complement regulator, fH. The structure also indicates the molecular basis of the host-specificity of the interaction between fH and the meningococcus, and informs attempts to develop novel therapeutics and vaccines.
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Project description:Neisseria meningitidis requires iron, and in the absence of iron alters its gene expression to increase iron acquisition and to make the best use of the iron it has. During different stages of colonization and infection available iron sources differ, particularly the host iron-binding proteins haemoglobin, transferrin, and lactoferrin. This study compared the transcriptional responses of N. meningitidis, when grown in the presence of these iron donors and ferric iron, using microarrays.Specific transcriptional responses to the different iron sources were observed, including genes that are not part of the response to iron restriction. Comparisons between growth on haemoglobin and either transferrin or lactoferrin identified changes in 124 and 114 genes, respectively, and 33 genes differed between growth on transferrin or lactoferrin. Comparison of gene expression from growth on haemoglobin or ferric iron showed that transcription is also affected by the entry of either haem or ferric iron into the cytoplasm. This is consistent with a model in which N. meningitidis uses the relative availability of host iron donor proteins as niche indicators.Growth in the presence of haemoglobin is associated with a response likely to be adaptive to survival within the bloodstream, which is supported by serum killing assays that indicate growth on haemoglobin significantly increases survival, and the response to lactoferrin is associated with increased expression of epithelial cell adhesins and oxidative stress response molecules. The transferrin receptor is the most highly transcribed receptor and has the fewest genes specifically induced in its presence, suggesting this is the favoured iron source for the bacterium. Most strikingly, the responses to haemoglobin, which is associated with unrestricted growth, indicates a low iron transcriptional profile, associated with an aggressive phenotype that may be adaptive to access host iron sources but which may also underlie the lethal features of meningococcal septicaemia, when haemoglobin may become a major source of iron.
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Project description:The zur regulon in Neisseria meningitidis was elucidated in the strain MC58 using a zur knockout strain and conditions which activate Zur ( zinc supplementation in the medium)
2012-10-05 | GSE38033 | GEO
Project description:The goal of this experiment was to evaluate the expression profile of cardiomyocytes freshly isolated from zebrafish embryos. Here we report mRNA abundance measurements for zebrafish cardiomyocytes at 20 hours post fertilization (hpf). We found that cardiomyocytes isolated from zebrafish embyros are enriched for orthologues of genes abundant in mammalian iPS-derived cardiomyocytes and other in vitro models of cardiomyocytes. It was also enriched with genes required for cardiac development in zebrafish and mammalian models. Using a rapid-throughput CRISPR-Cas9 based G0 screening pipeline, we tested 50 abundant transcription factors derived from this profile, rediscoved transcription factors known to be required for cardiac development, and found a novel role for zbtb16a in cardiac development.
2018-07-01 | GSE115263 | GEO
Project description:As mediators of adhesion, autoaggregation and bacteria-induced plasma membrane reorganization, type IV pili are at the heart of Neisseria meningitidis infection. Previous studies have proposed that two minor pilins, PilV and PilX, are displayed along the pilus structure and play a direct role in mediating these effects. In contrast with this hypothesis, combining imaging and biochemical approaches we found that PilV and PilX are located in the bacterial periplasm rather than along pilus fibers. Furthermore, preventing exit of these proteins from the periplasm by fusing them to the mCherry protein did not alter their function. Deletion of the pilV and pilX genes led to a decrease in the number, but not length, of pili displayed on the bacterial surface indicating a role in the initiation of pilus biogenesis. By finely regulating the expression of a central component of the piliation machinery, we show that the modest reductions in the number of pili are sufficient to recapitulate the phenotypes of the pilV and pilX mutants. We further show that specific type IV pili-dependent functions require different ranges of pili numbers.
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Project description:Neisseria meningitidis is a commensal of human nasopharynx. In some circumstances, this bacteria can invade the bloodstream and, after crossing the blood brain barrier, the meninges. A filamentous phage, designated MDAΦ for Meningococcal Disease Associated, has been associated with invasive disease. In this work we show that the prophage is not associated with a higher virulence during the bloodstream phase of the disease. However, looking at the interaction of N. meningitidis with epithelial cells, a step essential for colonization of the nasopharynx, we demonstrate that the presence of the prophage, via the production of viruses, increases colonization of encapsulated meningococci onto monolayers of epithelial cells. The analysis of the biomass covering the epithelial cells revealed that meningococci are bound to the apical surface of host cells by few layers of heavily piliated bacteria, whereas, in the upper layers, bacteria are non-piliated but surrounded by phage particles which (i) form bundles of filaments, and/or (ii) are in some places associated with bacteria. The latter are likely to correspond to growing bacteriophages during their extrusion through the outer membrane. These data suggest that, as the biomass increases, the loss of piliation in the upper layers of the biomass does not allow type IV pilus bacterial aggregation, but is compensated by a large production of phage particles that promote bacterial aggregation via the formation of bundles of phage filaments linked to the bacterial cell walls. We propose that MDAΦ by increasing bacterial colonization in the mucosa at the site-of-entry, increase the occurrence of diseases.
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Project description:DNA processing chain A (DprA) is a DNA binding protein which is ubiquitous in bacteria and required for DNA transformation. However, the interaction of DprA with competence proteins and recombination is poorly understood. Therefore, the proteomes of whole Neisseria meningitidis (Nm) wildtype and dprA mutant cells were compared. Such a comparative proteomic analysis increases our understanding of the interactions of DprA with other Nm components, and may elucidate its potential role beyond DNA processing in transformation. Using label-free quantitative proteomics, a total of 1057 unique Nm proteins were identified out of which 100 were quantified as differentially abundant (P ≤ 0.05 and fold change ≥ |2|) in the dprA null mutant. Proteins involved in homologous recombination (RecA, UvrD and HolA), pilus biogenesis (PilG, PilT1, PilT2, PilM, PilO, PilQ, PilF and PilE), and cell division, including core energy metabolism and response to oxidative stress were downregulated in dprA null mutant Nm. Immunoblotting was employed to validate the differences observed in two selected proteins. The analysis revealed reduced amounts of PilG in the dprA null mutant and reduced amounts of DprA in the Nm pilG null mutant. In conclusion, DprA interacts with proteins essential for Nm DNA recombination in transformation, pilus biogenesis and other functions associated with the inner membrane. Inverse downregulation of Nm DprA and PilG expression in the corresponding mutants indicate a link between DNA processing and pilus biogenesis demonstrated for the first time.
2017-04-26 | PXD006121 | Pride
Project description:Neisseria meningitidis serogroup B is a pathogen that can infect diverse sites within the human host. According to the N. meningitidis genomic information and experimental observations glucose can be completely catabolized through the Entner-Doudoroff pathway and the pentose phosphate pathway. The Embden-Meyerhof-Parnas pathway is not functional, because the gene for phosphofructokinase is not present. The phylogenetic distribution of phosphofructokinase indicates that in most obligate aerobic organisms PFK is lacking. We conclude that this is because of the limited contribution of PFK to the energy supply in aerobically grown organisms in comparison with the energy generated through oxidative phosphorylation. Under anaerobic or microaerobic conditions the available energy is limiting and PFK provides an advantage, which explains the presence of PFK in many (facultative) anaerobic organisms. In accordance with this, in silico flux balance analysis predicted an increase of biomass yield as a result of PFK expression. However, analysis of a genetically engineered N. meningitidis strain that expresses a heterologous phosphofructokinase showed that the yield of biomass on substrate decreased in comparison with a pfkA deficient control strain, which was associated mainly with an increase in CO2 production, whereas production of by-products was comparable between the two strains. This might explain why the pfkA gene has not been obtained by horizontal gene transfer, since it is initially unfavourable for biomass yield. No large effects related to heterologous expression of pfkA were observed in the transcriptome. Although our results suggest that introduction of PFK does not contribute to a more efficient strain in terms of biomass yield, achievement of a robust, optimal metabolic network that enables a higher growth rate or a higher biomass yield, might be possible after adaptive evolution of the strain, which remains to be investigated.
2009-11-11 | GSE18951 | GEO