Host cell targets of bacterial infections: combining systems biology and mechanistic approaches – HostTarget

The emergence of multi-drug-resistant mechanisms among major pathogenic bacteria represents a rising threat on human health. The lack of new therapeutic options requires an urgent effort of the scientific community to identify new microbial targets and envision alternative strategies. A promising approach consists of targeting the host instead of the pathogen. During infection, bacteria subvert key host factors and cellular responses to their own benefit. Interfering with these cellular factors may perturb pathogenic colonization or modulate the immune response in a way that, eventually, has a positive impact on the outcome of infection. As infection results from countless molecular interactions between microbial components and host factors, identifying critical host targets of bacteria requires addressing these cross-talks on a systems level.
With the HostTarget proposal, we propose an ambitious project which aims at identifying and characterizing new host targets of bacterial infection in the perspective of anti-infective treatments. Using the infection with the enteroinvasive bacterium Shigella flexneri as main model of infection, we apply systems biology approaches including large scale loss-of-function screens and phosphoproteomics to map the molecular processes involved in the bacterial invasion of epithelial cells, survival of infected cells and control of inflammation. Selected key host/pathogen interactions will be characterized in depth to design optimum perturbation strategies for treatment.
The first objective consists in identifying key host components involved in the entry process of S. flexneri into non-phagocytic epithelial cells. Primary genome wide RNAi and microRNA mimics screens have already been performed and revealed numerous candidates. Secondary screens monitoring different steps of the entry process will be used to define their roles and select promising targets for in depth molecular characterization and mechanistic modeling. To start, the role of TM9SF2, the first validated hit will be investigated in molecular details.
The second goal will be the characterization of the cellular pathways controlling the survival of infected epithelial cells used as replication niche. In particular, we will investigate how the secreted S. flexneri effector IpgD activates AKT and promotes host cell survival. Using LC-MS/MS-based phosphoproteomics, we showed that the mTOR pathway is highly enriched in the phosphoproteome of infected cells and that AKT activation is mTOR complex 2-dependent (mTORC2). As mTORC2 is a central player of both bacterial and viral infection and has a broad impact in health and disease in general, we will investigate its mode of activation during infection. In particular, we will use an original protein delivery tool to explore the implication of the proteins PI3K, IPMK and the PI5P and PIP3 phospholipids.
Finally, we will investigate the control of inflammation during infection of the intestinal epithelium. We recently reported a gap junction-mediated mechanism of communication between infected and non-infected bystander cells amplifying the expression of IL-8 during infection by enteroinvasive bacteria. We now aim at identifying the proteins involved in this phenomenon, as well as the small diffusing molecules. We performed a genome wide RNAi screen and identified around 200 genes affecting bystander IL-8 production. We validated the roles of TRAF6, TIFA and ATP1A1 and will investigate how these proteins affect the control of inflammation in vitro and in the mouse pulmonary model of S. flexneri infection.
This project has a strong expected value in basic and medical research. It will allow the identification of new host targets of pathogenic bacteria for treatment. It will also bring new insights into important signaling pathways including mTOR, AKT, NF-kB and the MAP kinases that are critical, beyond infection, in cancer, metabolic or inflammatory disorders.