SecA2-dependent effectors of Streptococcus pneumoniae and characterization of their role during infection

• Applicant: Professor Dr. Sven Hammerschmidt
• Subject Area: Medical Microbiology and Mycology, Hygiene, Molecular Infection Biology
• Term: since 2024
• Project identifier: Deutsche Forschungsgemeinschaft (DFG) - Project number 545493092

Protein secretion is an essential physiological process that needs to be tightly regulated to ensure bacterial survival under different conditions and to control infection. Streptococcus pneumoniae is both a natural colonizer of the human upper respiratory tract and a virulent pathogen. As such, the initial interactions with host cells will be crucial to establish carriage or disease and the proteins secreted at this time will play an important role in the cross talk with the host. Therefore, strain-specific protein factors are crucial for host infection, but their identity and how they are regulated for secretion is currently unknown. Although most housekeeping proteins are secreted using the SecA system, several bacteria, including the pneumococcus have conserved accessory Sec systems, whose role is poorly defined. In this project we propose to identify the bacterial proteins exported through this system and characterize its function during infection and for bacterial physiology. The three partners of the project, all experts on pneumococci but with complementary expertise, have divided the project in three main tasks and each partner will coordinate 1 of them. In Task 1 we will decipher the molecular workings and interaction network between SecA2, SecY2 and proteins of the canonical Sec pathway. Furthermore, the localization of the SecA2 complex will be visualized and the impact on the pneumococcal cell cycle investigated. Task 2 aims to identify the proteins secreted in a SecA2 dependent manner by using state of the art proteomics. This will be done both using an unbiased, and a targeted approach, in which we will address the role of SecA2 in the secretion of proteins known to be important for infection. These proteins, such as pneumolysin, LytC, and enolase, all have in common a lack of a signal peptide or cell wall anchoring motif suggestive of a non-canonical secretion. Task 3 aims to characterize the pathophysiological role of the SecA2 secretion system in the interplay with host cells and under experimental in vivo conditions. The phenotype and pathophysiology of our secA2-mutants constructed in our three selected serotypes will be assessed in a Galleria mellonella infection model as well as in an intranasal mouse infection model. Importantly, throughout Tasks 2 and 3, particular attention will be given to strain specificities as three different S. pneumoniae serotypes having three different genetic backgrounds will be used to increase the impact of our results. By combining molecular biology, cell imaging, proteomics and structural biology approaches, the knowledge generated from this proposal will allow the discovery of a molecular process delivering proteins important for colonization and/or infection, conserved in different species of Gram-positive bacteria, and which have the potential to become targets for the development of new antibacterial treatments.

• DFG Programme: Research Grants
• International Connection: France
• Partner Organisation: Agence Nationale de la Recherche / The French National Research Agency
• Cooperation Partners: Christophe Grangeasse, Ph.D.; Mélanie Hamon, Ph.D.