Mechanisms of Bacterial Virulence
Modifying Enzymes as Molecular Determinants of Bacterium-Host Cell Interactions
Tech Area / Field
- BIO-SFS/Biosafety and BioSecurity/Biotechnology
- MED-DID/Diagnostics & Devices/Medicine
3 Approved without Funding
Gamalei Institute of Epidemiology and Microbiology, Russia, Moscow
- Northwestern University Medical School, USA, IL, Chicago\nInstitut Pasteur, France, Paris
Project summaryOne of the most important initial steps in bacterium-host communications is the interaction of microbes with phagocytic eukaryotic cell. Normally, such contact is followed by ingestion of intruding bacteria and results in the killing of phagocytosed microorganisms. However, certain (i.e. pathogenic) bacteria have elaborated special strategies to resist such killing, either by active averting of engulfing or, in contrast, by active induction of phagocytosis with subsequent transformation of a phagocyte into a “niche” for replication. Bacteria of the first group are known as “extracellular pathogens” (e.g. Clostridia), while representatives of the second group are “intracellular pathogens” (e.g. Legionella).
Representatives of the two groups differ in many aspects (physiology, metabolism, composition of cell wall, pathogenesis of the diseases etc.). However, one feature is common to both types of infectious agents – in many examples targets of their molecular attack include cytoskeleton (actin, actin-related proteins, G-proteins). Thus, comparative analysis of bacterial enzymes (ADP-ribosyltransferas–es, UDP-glycosyltransferases, deaminases etc.), modifying actin machinery components, could represent a fruitful approach to investigations into molecular mechanisms of bacterial virulence and, in particular, into molecular mechanisms of microorganisms-host cell interactions.
The group submitting the project has been occupied with the problem of molecular mechanisms of bacteria-phagocytic cell interactions for more than 10 years. During this period a panel of proteins in Legionella, Listeria, Francisella tularensis has been investigated. These products include two modifying enzymes in Legionella – ADP-ribosyltransferase and UDP-glycosyltransferase. Data on the latter enzymes have been obtained by the authors for the first time. Thorough characterization of these proteins, produced by intracellular pathogen Legionella and their comparison to toxins of extracellular pathogens Clostridia, represents key points of the project.
The goal of the current project includes investigations into modifying enzymes of intracellular and extracellular pathogens and comparative analysis of the action of these enzymes upon actin apparatus of eukaryotic cell. This type of research could be interesting for an understanding of the virulence mechanisms of pathogenic bacteria and, on the other hand, it could facilitate deeper knowledge of the function of actin machinery. The project is scheduled for 2 years of research.
The main goal of the project is pided into 5 tasks: (1) purification of ADP-ribosyltransferase and UDP-glycosyltransferase of L.pneumophila; (2) Molecular cloning of their genes; (3) Construction of recombinant strains, hyperproducing these proteins; (4) Determination of eukaryotic substrates for Legionella enzymes; (5) Comparative investigation of Legionella proteins and clostridial toxins, modifying components of cytoskeleton upon cellular actin dynamics and their influence on intracellular survival of vaccine F.tularensis strains.
This project will result in new basic knowledge on molecular mechanisms of microorganisms-host cell interactions. This information will be important for an understanding of structure-function interrelationships of bacterial enzymes, and will aid investigations into genetic control of protein expression and secretion etc. Pure recombinant proteins and their cDNAs could be used as components of diagnostic test-systems and for development of new drugs and vaccines. Purified proteins (bacterial enzymes and their eukaryotic targets) and antisera could be used by cell biologists as probes during research on actin rearrangements in a cell. Comparative investigations into biological activities of clostridial toxins and Legionella enzymes will enable a deeper knowledge of the molecular mechanism of the action of modifying enzymes as virulence factors of extracellular and intracellular parasites.
The project is in good accordance with the aims of the ISTC, since it will facilitate the integration of Russian researchers into the international scientific community and will resolve definite problems of basic research in the field of virulence mechanisms of intracellular and extracellular pathogens. Results obtained during project implementation could create the basis for a new international scientific program in the field of molecular mechanisms of bacterial virulence.
Participation of foreign collaborators in the project is necessary for executing cooperative stages of research, scientific discussions, joint publications and commentary on reports and for monitoring overall project management.