Dataset Information

Transcriptome analysis of human brain microvascular endothelial cells response to Neisseria meningitidis and its antigen MafA using RNA-seq.

ABSTRACT: Interaction of Neisseria meningitidis (NM) with human brain microvascular endothelial cells (hBMECs) initiates of multiple cellular processes, which allow bacterial translocation across the blood-brain barrier (BBB). NM is equipped with several antigens, which interacts with the host cell receptors. Recently we have shown that adhesin MafA (UniProtKB-X5EG71), relatively less studied protein, is one of those surface exposed antigens that adhere to hBMECs. The present study was designed to comprehensively map the undergoing biological processes in hBMECs challenged with NM or MafA using RNA sequencing. 708 and 726 differentially expressed genes (DEGs) were identified in hBMECs exposed to NM and MafA, respectively. Gene ontology analysis of the DEGs revealed that several biological processes, which may alter the permeability of BBB, were activated. Comparative analysis of DEGs revealed that MafA, alike NM, might provoke TLR-dependent pathway and augment cytokine response. Moreover, both MafA and NM were able to induce genes involved in cell surface modifications, endocytosis, extracellular matrix remodulation and anoikis/apoptosis. In conclusion, this study for the first time describes effect of NM on the global gene expression in hBMECs using high-throughput RNA-seq. It also presents ability of MafA to induce gene expression, which might aid NM in breaching the BBB.

SUBMITTER: Kanova E

PROVIDER: S-EPMC6904618 | biostudies-literature | 2019 Dec

REPOSITORIES: biostudies-literature

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Transcriptome analysis of human brain microvascular endothelial cells response to Neisseria meningitidis and its antigen MafA using RNA-seq.

Káňová Evelína E Tkáčová Zuzana Z Bhide Katarína K Kulkarni Amod A Jiménez-Munguía Irene I Mertinková Patrícia P Drážovská Monika M Tyagi Punit P Bhide Mangesh M

Scientific reports 20191210 1

Interaction of Neisseria meningitidis (NM) with human brain microvascular endothelial cells (hBMECs) initiates of multiple cellular processes, which allow bacterial translocation across the blood-brain barrier (BBB). NM is equipped with several antigens, which interacts with the host cell receptors. Recently we have shown that adhesin MafA (UniProtKB-X5EG71), relatively less studied protein, is one of those surface exposed antigens that adhere to hBMECs. The present study was designed to compreh ...[more]

PMID: 31822804

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Project description:The pili and outer membrane proteins of Neisseria meningitidis (meningococci) facilitate bacterial adhesion and invasion into host cells. In this context expression of meningococcal PilC1 protein has been reported to play a crucial role. Intracellular calcium mobilization has been implicated as an important signaling event during internalization of several bacterial pathogens. Here we employed time lapse calcium-imaging and demonstrated that PilC1 of meningococci triggered a significant increase in cytoplasmic calcium in human brain microvascular endothelial cells, whereas PilC1-deficient meningococci could not initiate this signaling process. The increase in cytosolic calcium in response to PilC1-expressing meningococci was due to efflux of calcium from host intracellular stores as demonstrated by using 2-APB, which inhibits the release of calcium from the endoplasmic reticulum. Moreover, pre-treatment of host cells with U73122 (phospholipase C inhibitor) abolished the cytosolic calcium increase caused by PilC1-expressing meningococci demonstrating that active phospholipase C (PLC) is required to induce calcium transients in host cells. Furthermore, the role of cytosolic calcium on meningococcal adherence and internalization was documented by gentamicin protection assay and double immunofluorescence (DIF) staining. Results indicated that chelation of intracellular calcium by using BAPTA-AM significantly impaired PilC1-mediated meningococcal adherence to and invasion into host endothelial cells. However, buffering of extracellular calcium by BAPTA or EGTA demonstrated no significant effect on meningococcal adherence to and invasion into host cells. Taken together, these results indicate that meningococci induce calcium release from intracellular stores of host endothelial cells via PilC1 and cytoplasmic calcium concentrations play a critical role during PilC1 mediated meningococcal adherence to and subsequent invasion into host endothelial cells.

Project description:To extend our knowledge of target proteins in endothelial cells infected with the meningitis-causing pathogen Neisseria meningitidis, we characterized the interaction between the bacterial and human brain microvascular endothelial cell (HBMEC) monolayers. By use of human cDNA microarrays, transcriptional analysis revealed distinct responses to 4 and 8 h of infection. We also addressed the question of whether the major virulence factor of meningococci, i.e., the capsule, influences the host cell response. Of the 1,493 (at 4 h postinfection) and 1,246 (at 8 h postinfection) genes with altered expression upon bacterial contact, about 49.4% and 45%, respectively, depended on capsule expression. In particular, we identified an increase of expression for genes encoding proteins involved in bacterial adhesion and invasion. High levels of apoptosis-related gene (bad, bak, asp, and immediate-early response gene 1) expression could also be detected in infected cells. Further analyses confirmed that HBMECs displayed several hallmarks of apoptosis in response to N. meningitidis infection, namely, phosphatidylserine translocation and activation of caspase 3 and AMP-activated protein kinase alpha. Moreover, several differentially regulated genes not previously known to respond to meningococcal infection were identified. Of these, genes encoding cell adhesion proteins (CD44, CD98, and CD99), genes involved in downstream signaling of integrins (integrin-linked kinase, mitogen-activated protein kinase kinase 1, and mitogen-activated protein kinase kinase kinase 10) as well as negative regulators of these pathways (dual-specificity phosphatases 1, 5, and 14 and G protein pathway suppressor 2), and genes involved in cytoskeleton reorganization (those encoding Arp2/3, p34-arc, actinin alpha 1, vasodilatator-stimulated protein, and Wiskott-Aldrich syndrome protein) were the most prominent. This global transcriptional analysis creates a new platform for further molecular and cellular analysis of the interaction between N. meningitidis and target cells.

Dataset Information

Transcriptome analysis of human brain microvascular endothelial cells response to Neisseria meningitidis and its antigen MafA using RNA-seq.

Publications

Transcriptome analysis of human brain microvascular endothelial cells response to Neisseria meningitidis and its antigen MafA using RNA-seq.

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