AG Hammerschmidt - Infektionsbiologie
Infektionen mit pathogenen Erregern verursachen über 25 % der weltweiten Todesfälle. Die Arbeitsgruppe von Prof. Hammerschmidt erforscht die molekularen Mechanismen bakterieller Infektionen und die Mechanismen der Immunabwehr Gram-positiver Bakterien.
Molekulare und zelluläre Infektionsbiologie: Streptococcus pneumoniae
Streptococcus pneumoniae (Pneumokokken) sind Gram-positive Bakterien und gehören zur Normalflora des Respirationstraktes bei gesunden Erwachsenen und Kindern. Auf der anderen Seite sind Pneumokokken die Erreger schwerer lokaler Entzündungen und respiratorischer Erkrankungen. Die Ausbreitung der Pneumokokken im Menschen verursacht neben der Mittelohrentzündung auch lebensbedrohliche Erkrankungen. So sind Pneumokokken die häufigsten Erreger einer ambulant erworbenen Pneumonie (AEP) und häufige Erreger einer bakteriellen Meningitis und Sepsis. Die Mechanismen und Faktoren, die eine Umwandlung vom Kommensalen in ein pathogenes Bakterium begünstigen, sind wenig erforscht. Wir wollen in unseren Projekten die bakteriellen Faktoren identifizieren, die an der Pathogenese der Pneumokokken-Erkrankungen beteiligt sind. Des Weiteren wollen wir die molekularen Mechanismen der Erreger-Wirt-Interaktion in Infektionen in der Zellkultur und unter physiologischen und in vivo relevanten Bedingungen untersuchen.
Adherence molecules are key players in pathogen-host interactions. These bacterial components are usually surface exposed structures which facilitate adherence to host cells, or they target host serum proteins of the extracellular matrix. Pneumococci produce at least three classes of typical surface proteins: lipoproteins, LPXTG-anchored surface proteins and choline-binding proteins (CBPs). The CBPs have received considerable attention because of their versatility, and their sophisticated role in the interaction with host proteins.
PspC (also referred to as CbpA or SpsA) is one of the CBPs and was identified as the adhesin for the cellular polymeric Ig receptor (pIgR). The pIgR is a transmembrane protein produced by nasopharyngeal epithelial cells and the ectodomain, also known as secretory component (SC), consists of five Ig-like ectodomains.
Molecular analysis and the use of peptide array technology identified the hexapeptide YRNYPT in the amino-terminal portion of PspC as the minimal binding motif for SC. Identification of this motif, which is highly conserved in PspC proteins among different serotypes, might contribute towards a new peptide based vaccine strategy.
We have shown that the PspC-SC interaction is a human specific trait and identified the hexapeptide YRNYPT in PspC as the key co-factor of PspC for interaction with the ectodomains 3 and 4 of pIgR. The pathogenic significance of this interaction is underlined by the fact that PspC provides a mechanism for adherence to mucosal host cells and for invasion. Our current studies focus on how pneumococci and PspC engage the pIg receptor for invasion and which host cellular signal transduction cascades are induced upon host cell invasion.
A prerequisite to initiate severe and invasive infections is the ability of the bacteria to adhere to host cells. The attachment to host cells can be mediated directly via adhesion-receptor interaction such as the PspC-pIgR or indirectly by engaging a host protein as a molecular bridge. Gram-positive bacteria have evolved unique strategies to interact with host extracellular matrix (ECM) and plasma proteins for colonization, invasion, and survival. In the recent years we have identified several pneumococcal adhesins which interacts with components of the ECM by unique binding sites. The protein-protein cross-talks are able to promote the initial phase of the infection or facilitate survival in host niches. These interactions, therefore, might represent important virulence mechanisms for pneumococci which will be elucidated.
One striking example is the interaction of S. pneumoniae and other Gram-positive bacteria with the matricellular host protein human thrombospondin-1 (TSP1). TSP1 has key roles in interactions between human cells and components of the extracellular matrix. Our experiments indicated a novel role for the lectin TSP1 in pathogen-host interactions. Our studies indicated that Streptococcus pneumoniae and other Gram-positive pathogens including S. pyogenes, Staphylococcus aureus, and Listeria monocytogenes interact specifically with human TSP1. We were also able to show for the first time that host cell bound TSP1 promotes adherence of Gram-positive pathogens to human epithelial and endothelial cell lines. Attachment of S. pneumoniae and other Gram-positive pathogens to host cells via TSP1 was blocked by soluble peptidoglycan, indicating recognition of bacterial peptidoglycan by TSP1. In this scenario, peptidoglycan functions as adhesin and TSP1 acts as a molecular bridge linking Gram-positive bacteria with receptors on the host cell.
Microorganisms from all of the three empires bacteria, fungi and parasites have been convicted to interact in a specific manner with components of the fibrinolytic pathways. Pathogenic bacteria are capable to subvert the function of proteases, activators or inhibitors for their own benefits including dissemination within the host and evasion of host inflammatory immune response. The plasminogen activation system is part of the fibrinolysis which is tightly regulated and protected against dysfunction by various activators and inhibitors.
In the recent years we have analyzed the molecular and basic principles of plasminogen recruitment by pneumococci and other pathogenic bacteria including Neisseria meningitidis and Bifidobacteria. In addition, we have elucidated some of the benefits for the pathogens. The alfa-enolase of pneumococci was identified as the major plasmin(ogen)-binding protein on the surface of the bacteria. This is remarkable since alfa-enolases are essential glycolytic enzymes and they are usually found in the cytoplasm. Pneumococcal alfa-enolase is unique by its surface association thereby mediating the binding of plasmin(ogen) which was indicated by immunoelectron microscopy. Binding experiments and immunoelectron microscopy suggested the reassociation of secreted enolase to the bacterial cell surface.
Binding of human plasminogen to S. pneumoniae and its subsequent activation promotes penetration of bacteria through reconstituted basement membranes. For pneumococci, efficient plasmin-mediated degradation of host extracellular matrix (ECM) and transmigration through a fibrin matrix is correlated with the functional activity of the nonameric FYDKERKVY plasminogen binding region of the enolase. The impaired dissolution of fibrin by pneumococcal enolase mutants and their attenuation in an intranasal mouse infection model underlined the role of the nonameric peptide as the key co-factor in plasmin-mediated invasion of pneumococci. Nevertheless, it remains unclear how pneumococci engage the host fibrinolytic system to facilitate bacterial invasion and dissemination under in vivo conditions.
PspC is a key virulence factor and multifunctional protein that mediates adhesion to host cells but also immune evasion of the host complement. PspC has also been shown to bind components of the innate immune system such as complement protein C3 and factor H. PspC binds the host immune and complement regulator factor H, which is composed of 20 short consensus repeats (SCR). This interaction contributes to pneumococcal virulence. Our molecular analysis demonstrated two separate PspC binding regions within the factor H. The binding epitopes of PspC were localized to SCR8–11 and SCR19–20, by using recombinant factor H deletion constructs. A detailed analysis of binding epitopes in these SCR by peptide spot arrays identified several linear binding regions within the sequences of SCR8–11 and SCR19–20. Remarkably, Factor H attached to the surface of pneumococci via PspC significantly enhanced pneumococcal adherence to host epithelial and endothelial cells. This adhesion was specific and was blocked with a truncated N-terminal factor H-binding fragment of PspC. The role of Factor H recruitment during invasive pneumococcal disease and the impact on pneumococci induced host responses during uptake by host cells is currently under investigation.
- DFG | German Research Foundation
- BMBF | Federal Ministry of Education and Research - InfectControl 2020
- Federal State of Mecklenburg-Vorpommern | KoInfekt
- DAAD | German Academic Exchange Service
- Prof. Stefan Hippenstiel, Prof. Andreas Hocke, Prof. Norbert Suttorp, Department of Internal Medicine, Infectious and Respiratory Diseases, Universitätsmedizin Berlin
- Prof. Dr. Juan Hermoso, CSIC, Madrid, Spanien
- Prof. Dr. Uwe Koedel, Neurologie, LMU München
- Prof. Dr. Manfred Rohde, Helmholtz-Zentrum für Infektionsforschung, Braunschweig
- Prof. Dr. Jan Maarten van Dijl, Groningen, Niederlande
- Dr. Nicolas Gisch, Research Center Borstel
- Prof. Peter F. Zipfel, Hans-Knöll Institut, Jena
- PD Dr. Wolfgang Eisenreich, Biochemie, TU München