Signaltransduktion während der Invasion des zentralen Nervensystems durch Listeria monocytogenes - die Rolle von Wirtszellfaktoren mit Fokus auf CD44.
Kinder- und Jugendmedizin
Zusammenfassung der Projektergebnisse
Listeria monocytogenes (L. monocytogenes) can invade the central nervous system (CNS) by crossing either the blood-brain barrier (BBB) or the blood-cerebrospinal fluid barrier (BCSFB) and lead to brain infection resulting in meningitis. We investigated the role of signal transduction pathways involved during brain infection by L. monocytogenes with a special focus on CD44, a co-receptor of the host cell receptor MET. MET is targeted by the listerial virulence factor Internalin B (InlB) during host cell invasion. Analyzing the role of another Internalin of L. monocytogenes, InlF, which targets surface vimentin on host cells for CNS invasion, we found that InlF is not required for invasion in in vitro models of the human BCSFB (HIBCPP cells, do not express vimentin) or BBB (HBMEC, express vimentin). Surprisingly, treatment with the surface vimentin modulator Withaferin A reduced invasion of L. monocytogenes into HBMEC, pointing to an InlF independent function of vimentin. Previously published results indicated a role of cell surface vimentin in mediating extracellular matrix stiffnessdependent adhesion of L. monocytogenes. We propose that this mechanism is involved during CNS invasion by L. monocytogenes at the BBB. To characterize the role of CD44/CD44v6 at the BBB during infection with L. monocytogenes in more detail, we used CRISPR/Cas9 technology to generate the cell lines HBMECΔCd44, HBMECΔCd44v6, HBMECΔIcam-1, and HBMECΔCd44ΔIcam-1. Using these cell lines as well as CD44v6 blocking peptides a clear function of CD44v6 during invasion of InlB-coated latex beads or L. monocytogenes was demonstrated, which coincided with the activation of MET and of downstream targets as Akt, GSK3α/β and Erk. Importantly, we found that deletion of ICAM-1 also partially decreases the activation of MET, whereas the deletion of both Cd44 and ICAM-1 completely abrogated MET activation, pointing to a direct role during host cell invasion of L. monocytogenes at the BBB. Transcriptome analysis of wild-type HBMECs and HBMEC ΔCD44v6 challenged with L. monocytogenes or in absence of bacteria demonstrated a clear impact of CD44v6 in the dampening of expression of genes related to the immune response that is largely independent of listerial infection. Investigation of HBMEC ΔCD44 (all CD44 isoforms removed) revealed a major impact on expression of immune related genes, with a particular positive impact on neutrophil activation. We hypothesize that CD44/CD44v6 supports escape of L. monocytogenes from attack by the immune system in the BBB microenvironment. The function of CD44 during infection with L. monocytogenes was investigated in vivo in mouse models. We did not observe significant differences in the loads of bacteria in mouse organs in C56BL/6JRj mice treated or not treated with a CD44v6 blocking peptide. Infection of mice with an inducible knockout of CD44 or CD44v6 in the BBB and BCSFB unfortunately did not result in reproducible brain infection under induced and control conditions. Analyzing the role of CD44/CD44v6 in these KO mice would require optimization of the in vivo infection protocol.
Projektbezogene Publikationen (Auswahl)
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BeadNet: deep learning-based bead detection and counting in low-resolution microscopy images. Bioinformatics, 36(17), 4668-4670.
Scherr, Tim; Streule, Karolin; Bartschat, Andreas; Böhland, Moritz; Stegmaier, Johannes; Reischl, Markus; Orian-Rousseau, Véronique & Mikut, Ralf
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Potential Roles and Functions of Listerial Virulence Factors during Brain Entry. Toxins, 12(5), 297.
Banović, Franjo; Schroten, Horst & Schwerk, Christian
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Different Involvement of Vimentin during Invasion by Listeria monocytogenes at the Blood–Brain and the Blood–Cerebrospinal Fluid Barriers In Vitro. International Journal of Molecular Sciences, 23(21), 12908.
Banovic, Franjo; Schulze, Sandrin; Abu, Mraheil Mobarak; Hain, Torsten; Chakraborty, Trinad; Orian-Rousseau, Véronique; Moroniak, Selina; Weiss, Christel; Ishikawa, Hiroshi; Schroten, Horst; Adam, Rüdiger & Schwerk, Christian
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Signal transduction during CNS invasion by Listeria monocytogenes and the role of CD44 in the invasion. PhD thesis. (2022)
Banovic, F.
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The role of CD44 in Listeria monocytogenes infection of the brain endothelium. PhD thesis. (2022)
Streule, K. L.
