Function of Legionella pneumophila Mip in bacterial protein secretion, human lung tissue infection and colonization of nematodes

• Antragsteller: Professor Dr. Michael Steinert
• Fachliche Zuordnung: Parasitologie und Biologie der Erreger tropischer Infektionskrankheiten
• Förderung: Förderung von 2013 bis 2017
• Projektkennung: Deutsche Forschungsgemeinschaft (DFG) - Projektnummer 232674996

The macrophage infectivity potentiator (Mip) protein is a major virulence factor of Legionella pneumophila, an aquatic bacterium and the causative agent of Legionnaires´ disease. Mip exhibits peptidyl-prolyl-cis/trans-isomerase (PPIase) activity, binds to the extracellular matrix (ECM) component collagen IV and contributes to the bacterial transmigration across the lung epithelial barrier by a yet unknown mechanism. Our previous analyses revealed that the collagen-derived peptide P290 binds to a specific hydrophobic cavity of Mip, thus inhibiting bacterial epithelial transmigration in vitro. Moreover, we demonstrated that the secretome profile of L. pneumophila is strongly influenced by Mip. Therefore, we aim to answer three fundamental questions. (i) How does Mip influence protein secretion of L. pneumophila? (ii) What is the crucial contribution of Mip during human lung infection? (iii) Did Mip initially adapt to collagen of simple environmental metazoans such as nematodes? To address these questions, the Mip-dependent secretome of L. pneumophila and the structural, enzymatic and regulatory functions of Mip during L. pneumophila protein secretion will be analyzed using a proteomic approach, substrate zymography, specific secretion mutants and functionally varied inhibitors. To link Mip-dependent alterations of the extracellular milieu with tissue destruction and bacterial replication, we will analyze the Mip-dependent histopathology during ex vivo infections of explanted human lung tissue sections from tumor patients. Direct and indirect effects of Mip will be analyzed by specific inhibition assays with PPIase inhibitors (FK506, rapamycin), the collagen binding inhibitor P290, specific protease inhibitors, in situ zymography and confocal laser scanning microscopy (CLSM). Since preliminary environmental data and in vitro infections strongly suggest that L. pneumophila colonizes nematodes, we will also analyze the Mip-dependent infection of the collagen IV-rich intestinal tract of Caenorhabditis elegans. At an aquatic sampling site, where L. pneumophila and nematodes are both abundant, we will also address the question whether or not nematodes are real hosts of L. pneumophila. The proof of a natural interaction between Legionella and nematodes would open a new chapter in the research of Legionella ecology, evolution and virulence.

• DFG-Verfahren: Sachbeihilfen