Mechanism of complement regulator Factor H mediated pneumococcal uptake by human cells
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2242 Abstracts / Molecular Immunology 47 (2010) 2198–2294 tion amongst the tested strains. In addition, we observed that the complement inhibitor C4BP functions as a bridging molecule facili- tating pneumococcal adherence to the host epithelial cells, thereby likely promoting initial steps of colonization during the course of infection. doi:10.1016/j.molimm.2010.05.136 214 Factor H Tyr402 binds more efficiently to Leptospira than Factor H His402 but both variants exhibit similar cofactor activities A.S. Silva, M.M. Castiblanco Valencia, A.S. Barbosa, L. Isaac Instituto de Ciências Biomédicas, Universidade de São Paulo and Insti- tuto Butantan, Brazil Leptospirosis is one of the most important worldwide zoonoses and is caused by Leptospira, spread mostly through the urine from contaminated rodents. The immune response is essential for host protection and the activation of the complement system is impor- tant for bacteria killing. Recently, it has been demonstrated that nonpathogenic Leptospira strains are rapidly killed by the comple- ment system, whereas pathogenic strains are able to resist serum bactericidal activity. However, limited knowledge is available con- cerning the mechanisms employed by these bacteria to circumvent this major line of defense. Recently, it has been demonstrated that pathogenic leptospiral strains are able to bind to this com- plement regulator and that surface-bound FH acts as a cofactor for the cleavage of C3b by Factor I. This indicates that acquisition of this complement regulator may contribute to leptospiral serum resistance. Two forms of human FH have been intensively stud- ied: FH Tyr402 and FH His402 and it has been shown that this last variant is highly associated with the development of age-related macular degeneration. In this study, we assessed the relevance of the FH Tyr402His polymorphism with respect to FH binding to Leptospira. Both FH variants were purified from human plasma by chromatography and incubated with the virulent strain Lep- tospira interrogans serovar Pomona. Surface-bound proteins were evaluated by an enzyme immunoassay using whole Leptospira. We observed a marked difference in the interaction of these two FH forms with the bacteria: FH Tyr402 bound more efficiently to Lep- tospira than FH His402. Despite binding to Leptospira with different affinities, both variants exhibited similar cofactor activities against C3b, thus indicating that they are equally efficient in regulating the complement cascade. doi:10.1016/j.molimm.2010.05.137 215 Mechanism of complement regulator Factor H mediated pneu- mococcal uptake by human cells Vaibhav Agarwal a,b,1 , Tauseef Asmat a , Shanshan Luo c , Peter F. Zipfel c,d , Sven Hammerschmidt a,b,∗ a Department Genetics of Microorganisms, Institute for Genetics and Functional Genomics, Ernst Moritz Arndt University of Greifswald, Greifswald, Germany b Max von Pettenkofer-Institute for Hygiene and Medical Microbiology, Ludwig-Maximilians-University München, München, Germany c Department of Infection Biology, Leibniz Institute for Natural Product Research and Infection Biology, Hans-Knoell-Institute, Jena, Germany d Friedrich Schiller University, Jena, Germany The Gram-positive bacterium Streptococcus pneumoniae (pneu- mococci) is an important human pathogen causing severe disease. Pneumococci are able to recruit human complement regulator Factor H to the bacterial cell surface via an interaction of PspC (Pneumococcal surface protein C) with the short consensus repeats (SCR) 8-11 and SCR19-20 of Factor H. In this study we demon- strate how bacterial-bound Factor H is exploited to promote pneumococcal adherence to and uptake by epithelial cells or polymorphonuclear leukocytes (PMNs). Glycosaminoglycans such as heparin and dermatan sulfate significantly reduced Factor H- mediated pneumococcal adherence to epithelial cells without affecting binding of Factor H to the bacteria. In addition, adher- ence of pneumococci to human epithelial cells was inhibited by mAbs recognizing SCR19-20 of Factor H suggesting that the C ter- minal glycosaminoglycan-binding region of Factor H mediates the bacterial contact to human cells. Blocking of the CR3 receptor, i.e. CD11b and CD18, of PMNs or CR3-expressing epithelial cells reduced significantly pneumococcal association with PMNs and uptake by epithelial cells, respectively. Similar, Pra1, a secreted Candida albicans protein and CR3 ligand, blocked the formation of pneumococcal-PMNs associates. Strikingly, Pra1 inhibited also pneumococcal uptake by lung epithelial cells, but did not affect adherence. In addition, inhibition experiments suggested that the dynamics of host-cell actin microfilaments and activity of pro- tein tyrosine kinases and phosphatidylinositol-kinase (PI3K) are requird for invasion of pneumococci via Factor H. In conclusion we demonstrate that pneumococcal entry into host cells via Factor H occurs by a two-step mechanism. The initial contact is medi- ated through an interaction of pneumococcal-bound Factor H with host cell glycosaminoglycans while pneumococcal uptake depends on integrins, host cell actin cytoskeleton dynamics and signalling molecules such as PI3K. doi:10.1016/j.molimm.2010.05.138 1 Present address: Department of Laboratory Medicine, University Hospital Malmö, Lund University, Malmö, Sweden.
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Transcript of Mechanism of complement regulator Factor H mediated pneumococcal uptake by human cells
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242 Abstracts / Molecular Imm
ion amongst the tested strains. In addition, we observed that theomplement inhibitor C4BP functions as a bridging molecule facili-ating pneumococcal adherence to the host epithelial cells, therebyikely promoting initial steps of colonization during the course ofnfection.
oi:10.1016/j.molimm.2010.05.136
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actor H Tyr402 binds more efficiently to Leptospira than FactorHis402 but both variants exhibit similar cofactor activities
.S. Silva, M.M. Castiblanco Valencia, A.S. Barbosa, L. Isaac
Instituto de Ciências Biomédicas, Universidade de São Paulo and Insti-uto Butantan, Brazil
Leptospirosis is one of the most important worldwide zoonosesnd is caused by Leptospira, spread mostly through the urine fromontaminated rodents. The immune response is essential for hostrotection and the activation of the complement system is impor-ant for bacteria killing. Recently, it has been demonstrated thatonpathogenic Leptospira strains are rapidly killed by the comple-ent system, whereas pathogenic strains are able to resist serum
actericidal activity. However, limited knowledge is available con-erning the mechanisms employed by these bacteria to circumventhis major line of defense. Recently, it has been demonstratedhat pathogenic leptospiral strains are able to bind to this com-lement regulator and that surface-bound FH acts as a cofactor forhe cleavage of C3b by Factor I. This indicates that acquisition ofhis complement regulator may contribute to leptospiral serumesistance. Two forms of human FH have been intensively stud-ed: FH Tyr402 and FH His402 and it has been shown that this lastariant is highly associated with the development of age-relatedacular degeneration. In this study, we assessed the relevance of
he FH Tyr402His polymorphism with respect to FH binding toeptospira. Both FH variants were purified from human plasmay chromatography and incubated with the virulent strain Lep-ospira interrogans serovar Pomona. Surface-bound proteins werevaluated by an enzyme immunoassay using whole Leptospira. Webserved a marked difference in the interaction of these two FHorms with the bacteria: FH Tyr402 bound more efficiently to Lep-ospira than FH His402. Despite binding to Leptospira with different
ffinities, both variants exhibited similar cofactor activities against3b, thus indicating that they are equally efficient in regulating theomplement cascade.oi:10.1016/j.molimm.2010.05.137
gy 47 (2010) 2198–2294
215
Mechanism of complement regulator Factor H mediated pneu-mococcal uptake by human cells
Vaibhav Agarwal a,b,1, Tauseef Asmat a, Shanshan Luo c, Peter F.Zipfel c,d, Sven Hammerschmidt a,b,∗
a Department Genetics of Microorganisms, Institute for Genetics andFunctional Genomics, Ernst Moritz Arndt University of Greifswald,Greifswald, Germanyb Max von Pettenkofer-Institute for Hygiene and Medical Microbiology,Ludwig-Maximilians-University München, München, Germanyc Department of Infection Biology, Leibniz Institute for Natural ProductResearch and Infection Biology, Hans-Knoell-Institute, Jena, Germanyd Friedrich Schiller University, Jena, Germany
The Gram-positive bacterium Streptococcus pneumoniae (pneu-mococci) is an important human pathogen causing severe disease.Pneumococci are able to recruit human complement regulatorFactor H to the bacterial cell surface via an interaction of PspC(Pneumococcal surface protein C) with the short consensus repeats(SCR) 8-11 and SCR19-20 of Factor H. In this study we demon-strate how bacterial-bound Factor H is exploited to promotepneumococcal adherence to and uptake by epithelial cells orpolymorphonuclear leukocytes (PMNs). Glycosaminoglycans suchas heparin and dermatan sulfate significantly reduced Factor H-mediated pneumococcal adherence to epithelial cells withoutaffecting binding of Factor H to the bacteria. In addition, adher-ence of pneumococci to human epithelial cells was inhibited bymAbs recognizing SCR19-20 of Factor H suggesting that the C ter-minal glycosaminoglycan-binding region of Factor H mediates thebacterial contact to human cells. Blocking of the CR3 receptor,i.e. CD11b and CD18, of PMNs or CR3-expressing epithelial cellsreduced significantly pneumococcal association with PMNs anduptake by epithelial cells, respectively. Similar, Pra1, a secretedCandida albicans protein and CR3 ligand, blocked the formationof pneumococcal-PMNs associates. Strikingly, Pra1 inhibited alsopneumococcal uptake by lung epithelial cells, but did not affectadherence. In addition, inhibition experiments suggested that thedynamics of host-cell actin microfilaments and activity of pro-tein tyrosine kinases and phosphatidylinositol-kinase (PI3K) arerequird for invasion of pneumococci via Factor H. In conclusionwe demonstrate that pneumococcal entry into host cells via FactorH occurs by a two-step mechanism. The initial contact is medi-ated through an interaction of pneumococcal-bound Factor H with
host cell glycosaminoglycans while pneumococcal uptake dependson integrins, host cell actin cytoskeleton dynamics and signallingmolecules such as PI3K.doi:10.1016/j.molimm.2010.05.138
1 Present address: Department of Laboratory Medicine, University HospitalMalmö, Lund University, Malmö, Sweden.
