Pneumococcal Interaction with Human Dendritic Cells: Phagocytosis, Survival, and Induced Adaptive Immune Response Are Manipulated by PavA This information is current as of September 27, 2021. Nadja Noske, Ulrike Kämmerer, Manfred Rohde and Sven Hammerschmidt J Immunol 2009; 183:1952-1963; Prepublished online 1 July 2009; doi: 10.4049/jimmunol.0804383 Downloaded from http://www.jimmunol.org/content/183/3/1952 References This article cites 52 articles, 24 of which you can access for free at: http://www.jimmunol.org/ http://www.jimmunol.org/content/183/3/1952.full#ref-list-1 Why The JI? Submit online. • Rapid Reviews! 30 days* from submission to initial decision • No Triage! Every submission reviewed by practicing scientists by guest on September 27, 2021 • Fast Publication! 4 weeks from acceptance to publication *average Subscription Information about subscribing to The Journal of Immunology is online at: http://jimmunol.org/subscription Permissions Submit copyright permission requests at: http://www.aai.org/About/Publications/JI/copyright.html Email Alerts Receive free email-alerts when new articles cite this article. Sign up at: http://jimmunol.org/alerts The Journal of Immunology is published twice each month by The American Association of Immunologists, Inc., 1451 Rockville Pike, Suite 650, Rockville, MD 20852 Copyright © 2009 by The American Association of Immunologists, Inc. All rights reserved. Print ISSN: 0022-1767 Online ISSN: 1550-6606. The Journal of Immunology Pneumococcal Interaction with Human Dendritic Cells: Phagocytosis, Survival, and Induced Adaptive Immune Response Are Manipulated by PavA1 Nadja Noske,*†‡ Ulrike Ka¨mmerer,§ Manfred Rohde,¶ and Sven Hammerschmidt2*†‡ Dendritic cells (DCs) ingest and process bacteria for presenting their Ags to T cells. PavA (pneumococcal adherence and virulence factor A) is a key virulence determinant of pneumococci under in vivo conditions and was shown to modulate adherence of pneumococci to a variety of nonprofessional phagocytic host cells. Here, we demonstrated the role of PavA for the interaction of human DCs with live pneumococci and analyzed the induced host cell responses upon ingestion of viable pneumococci. Expression of PavA protected pneumococci against recognition and actin cytoskeleton-dependent phagocytosis by DCs compared with iso- genic pavA mutants. A major proportion of internalized pneumococci were found in membrane-bound phagosomes. Pneumococcal Downloaded from phagocytosis promotes maturation of DCs, and both wild-type pneumococci and PavA-deficient pneumococci triggered production of proinflammatory cytokines such as IL-1␤, IL-6, IL-8, IL-12, and TNF-␣ and antiinflammatory IL-10. However, cytokine production was delayed and reduced when DCs encounter pneumococci lacking PavA, which also results in a less efficient activation of the adaptive immune response. Strikingly, purified PavA reassociates to pneumococci but not DCs and reduced phagocytosis of the pavA mutant to levels similar to those of wild-type pneumococci. Additionally, pavA mutants covered with exogenously provided PavA protein induced a DC cytokine profile similar to wild-type pneumococci. In conclusion, these results http://www.jimmunol.org/ suggest that PavA is key factor for live pneumococci to escape phagocytosis and to induce optimal cytokine productions by DCs and adaptive immune responses as well. The Journal of Immunology, 2009, 183: 1952–1963. treptococcus pneumoniae (pneumococci) are commensals nation of immune defenses upon stimulation in response to micro- of the human respiratory tract and colonize up to 70% of bial signals (5). Immature DCs efficiently phagocytose or macropi- S the individuals without causing clinical symptoms. How- nocytose bacteria and process them into cell surface-presentable ever, these apparently harmless colonizers are also well known as Ags. During this process, the immature DCs mature and convert serious human pathogens that transmigrate into the lungs, enter the into potent APCs. Maturation of DCs is characterized by changes bloodstream, and cross the blood-brain barrier (1). As a conse- in surface expression of MHC, adhesion and costimulatory mole- by guest on September 27, 2021 quence, this versatile pathogen causes infections ranging from cules, and cytokine production as well (6). Upon maturation, DCs severe local infections, such as otitis media and sinusitis, to migrate from the place of Ag uptake into tissue-draining lymphoid life-threatening infections, including pneumonia, sepsis, and organs such as lymph nodes or spleen. Maturation of DCs dimin- meningitis (2). In healthy individuals, the mucosal surfaces with ishes their capacity to internalize and process Ags but greatly en- their epithelial cells and the secreted mucus constitute a physical hances their ability to prime naive T cells. As a consequence, DC barrier that prevents pathogens to gain access into deeper tissues. responses upon bacterial infections initiate adaptive immune re- Here, the pathogens are also faced by the production of antimi- sponses (7). However, pathogens have evolved various strategies crobial agents, such as defensins (3, 4). Additionally, mucosal tis- to escape host immune responses. Regarding DCs, their subversion sues are scattered with sentinel professional phagocytes and APCs by pathogens and exploitation as a Trojan horse to disseminate including dendritic cells (DCs).3 DCs are involved in the coordi- within the host—as recently shown for HIV and Chlamydia—is under debate (8–10). Pneumococci are encased by a capsular polysaccharide (CPS), *Department Genetics of Microorganisms, Institute for Genetics and Functional Genomics, University of Greifswald, Greifswald, Germany; †Max von Pettenkofer and the CPS is recognized as a sine qua non for invasive diseases Institute, Ludwig-Maximilians University Mu¨nchen, Mu¨nchen, Germany; ‡Research (11). The capsule protects pneumococci against uptake into pro- Center for Infectious Diseases and §Department of Obstetrics and Gynecology, Uni- fessional phagocytes and complement-mediated opsonophagocy- versity of Wu¨rzburg, Wu¨rzburg, Germany; and ¶Department of Microbial Pathogen- esis, Helmholtz Center for Infection Research, Braunschweig, Germany tosis (12, 13). Another potent virulence factor of pneumococci Received for publication December 31, 2008. Accepted for publication May 28, 2009. interfering with both eukaryotic cell function and the immune sys- tem is the pore-forming cytolysin pneumolysin (14, 15). TLR4 The costs of publication of this article were defrayed in part by the payment of page charges. This article must therefore be hereby marked advertisement in accordance recognizes pneumolysin as a pathogen-associated molecular pat- with 18 U.S.C. Section 1734 solely to indicate this fact. tern, thereby providing protection against pneumococcal infections 1 This work was supported in part by the Bundesministerium fu¨r Bildung und For- (16). Moreover, pneumolysin production enhances mucosal clear- schung (CAPNETZ C8; to S.H.) and the Deutsche Forschungsgemeinschaft (Sonder- ance of pneumococci by stimulating neutrophil recruitment and forschungsbereich 479, Teilprojekt A7, DFG HA 3125/2-1 and 4-1). 2 Address correspondence and reprint requests to Dr. Sven Hammerschmidt, De- partment Genetics of Microorganisms, Institute for Genetics and Functional charide; FESEM, field emission electron microscopy; iDC, immature dendritic cell; Genomics, Ernst Moritz Arndt University Greifswald, Friedrich Ludwig Jahn Lamp1, lysosomal-associated membrane protein 1; MBP, maltose-binding protein; Strasse 15a, D-17487 Greifswald, Germany. E-mail address: sven.hammerschmidt@ MOI, multiplicity of infection; OxMi, oxidative mitogenesis; PavA, pneumococcal uni-greifswald.de adherence and virulence factor A; TEM, transmission electron microscopy. 3 Abbreviations used in this paper: DC, dendritic cell; BMDC, bone marrow-derived dendritic cell; CLSM, confocal laser scanning microscopy; CPS, capsular polysac- Copyright © 2009 by The American Association of Immunologists, Inc. 0022-1767/09/$2.00 www.jimmunol.org/cgi/doi/10.4049/jimmunol.0804383 The Journal of Immunology 1953 promotes bacterial Ag delivery to the nasal-associated lymphoid mary Abs used for DCs were: human CD11c (IgG1), CD25 (IgG1), CD83 tissue (17, 18). The cell wall of pneumococci is decorated with (IgG2), CD86 (IgG2), or HLA-DR (IgG1) for MHC-class II. Nonspecific numerous virulence factors that execute their function in different binding of Abs was calculated by incubating DCs with the appropriate isotype anti-IgG2 or anti-IgG1 (Caltag Laboratories, BD Pharmingen, or host niches (19, 20). Remarkably, atypical surface proteins, lack- Miltenyi Biotec). The Abs were conjugated with PE or FITC. Surface ing a leader peptide or even a motif for membrane anchoring such marker expression was quantified using a FACSCalibur (BD Biosciences). as the PavA protein (pneumococcal adherence and virulence factor Infection of human DCs A), have been found to be key virulence factors of S. pneumoniae. Deficiency in PavA impairs adherence of pneumococci to host For our antibiotic protection assays DCs were seeded into 96-well cell ϫ 5 epithelial and endothelial cells and attenuates virulence of pneu- culture plates (Greiner Bio-One) at a density of 1 10 cells/well. To perform immunofluorescence staining or electron microscopy, DCs were mococci in mice infection models (21, 22). Although pneumococci seeded on glass coverslips (diameter, 12 mm) in 24-well cell culture plates lacking PavA produce