Pseudomonas aeruginosa infection: analysis of antigenic proteins of the virulence-associated type VI secretion system

Interaction between microbes and human hosts can range from a benign, even commensal collaboration to a competition that may turn fatal resulting in the death of the host, the bacteria or both. The outcome of the interaction is balanced by a communication/signalling game in between the host and the pathogen. Among the arsenal of communication tools used by bacteria to influence host response are secretion systems (SS), which deliver toxins to the host. The SS and the transported toxins are key players in bacterial pathogenesis. Recent studies have seen the emergence of a novel type of SS, named T6SS. Bacteria harbouring T6SS all have a life style where they live in intimate contact with a host.
Pseudomonas aeruginosa is an opportunistic pathogen. It causes urinary tract infections, respiratory system infections, dermatitis, soft tissue infections, bacteremia, bone and joint infections, gastrointestinal infections and a variety of systemic infections, particularly in immuno-suppressed patients. P. aeruginosa infection is a serious problem in patients hospitalized with cancer, cystic fibrosis, and burns. The case fatality rate in these patients is 50 percent. P. aeruginosa is the third most commonly-isolated nosocomial pathogen accounting for 10% of all hospital-acquired infections.
P. aeruginosa possesses 3 T6SS. The goal of our study is to understand the molecular mechanisms, which are involved in the T6SS functioning, knowledge applicable to other T6SS-containing bacterial pathogens. We will further use this knowledge to develop antimicrobials, which will help fighting human morbidity and mortality due to persistent bacterial infections.

The secretion process is crucial for supporting communication between the bacteria and its environment or its host. Secretion systems are considered as types in gram-negative bacteria according to their structure and mode of functioning. Recent studies have seen the emergence of a novel type of bacterial secretion machinery, the type VI secretion system (T6SS). The T6SS components are mostly original, not found in other secretion systems and they have been proposed to promote secretion of two types of proteins, namely Hcp and Vgr, whose function and host targets are still unclear.
The T6SS components are encoded within cluster of genes initially named IAHP for IcmF-associated homologous proteins, since they were systematically found associated with a gene encoding an IcmF-like component. Bacteria harbouring T6SS are all gram-negative bacteria and they all have a life style where they live in intimate contact with eukaryotic cells. The T6SS is involved in bacterial pathogenesis, usually for colonisation, virulence, cytotoxicity or intracellular multiplication. The expression of T6SS genes is induced in vivo while the bacteria colonize the host. The precise role and mode of action of the T6SS is still unknown.
Our model organism, P. aeruginosa, is found in multiple human infections and harbours 3 T6SS clusters (HSI-I to HSI-III). Whereas several T6SS components have putative function, others have not. Among these, the HsiBC and HsiFGH homologues are antigenic and produced during infection since HsiBC and HsiFG antibodies are found in sera of infected hosts. This indicates that these proteins may be secreted or cell surface exposed upon activation of the T6SS.
Our project focuses on the characterization of the antigenic P. aeruginosa T6SS proteins HsiBCFGH, and aims at unravelling their role in the T6SS function. In order to gain understanding in the function, on the one hand we will mutagenize the genes and analyse the impact on host interaction, on the other hand we will perform cristallography studies to determine whether folds in these unknown proteins could be recognized and give hints on existence of putative homologues.