The interplay between autophagy and S. aureus infection

• Applicant: Professor Dr. Ingo Schmitz
• Subject Area: Medical Microbiology and Mycology, Hygiene, Molecular Infection Biology ; Immunology
• Term: from 2012 to 2022
• Project identifier: Deutsche Forschungsgemeinschaft (DFG) - Project number 213312245

Autophagy is a catabolic mechanism that is important for many biological processes such as cell homeostasis, development and immunity. On the molecular level, autophagy is executed by so-called AuTophaGy-related (ATG) proteins. At the center of the autophagy pathway and crucial for efficient autophagic flux are two ubiquitin conjugation systems including the ATG5 conjugation system. However, signaling pathways regulating the activity of essential autophagy mediators are still poorly defined. During the previous funding period, we characterized a Gadd45β-MEKK4-p38 MAPK signaling cascade that leads to inhibition of autophagy. Specifically, p38 MAPK activated by the Gadd45β-MEKK4 module translocates to autophagosomes where it targets the ATG5 complex. Here, the region surrounding threonine 75 of ATG5 appears to be an important interaction site that directs p38 MAPK towards phosphorylation of ATG12. In addition, we showed that infection with Staphylococcus aureus induces selective autophagy. Intracellular S. aureus gets associated with ubiquitin and recruited via autophagy receptors such as SQSTM1/p62 into autophagosomes. However, S. aureus evades autophagic degradation by activating p38 MAPK, degradation of the autophagosomal membranes and escape into the cytoplasm. We believe that this is an important mechanism how S. aureus can persist in the host. Therefore, we apply for a continuation of this project to investigate the interplay between S. aureus infection and autophagy. First, we want to analyze by which mechanisms S. aureus induces autophagy upon invasion of the host cell. Thus, we will analyze the contribution of pattern recognition receptors as well as metabolic stress to the induction of autophagy. Second, we will clarify how p38 MAPK is activated upon S. aureus infection. Next to the analysis of prime candidates such as TLR2 and Nod2 as well as the bacterial kinase PknB, we will use proteomics and dual RNA-seq to identify bacterial factors that are upregulated in autophagy-sufficient host cells. Third, we will analyze which factors allow S. aureus to escape from the autophagosome. Prime candidates are virulence factors that are described to have pore-forming or lipase activities. Thus, bacterial strains deficient in these factors will be employed in infection experiments. Additionally, the dual RNA-seq analysis that we plan to perform in autophagy-sufficient and -deficient cells will help us to decipher genes that are involved in this process. For all the questions we want to address, we will use a combination of knockout host cells and gene-deficient bacteria. Thus, combining innate immunology, cellular microbiology, genomics and proteomics we will identify and characterize the virulence factors that help S. aureus to evade autophagy as well as the autophagy components that are targeted by the bacteria.

• DFG Programme: Research Grants