%0 Generic
%A Walch, Philipp Darius Konstantin
%C Heidelberg
%D 2021
%F heidok:30028
%R 10.11588/heidok.00030028
%T Dissecting the host-pathogen interface during Salmonella infection
%U https://archiv.ub.uni-heidelberg.de/volltextserver/30028/
%X Facultatively intracellular pathogens adapt to and rewire host defenses to induce infection and promote survival and proliferation. Salmonella enterica serovar Typhimurium (STm) injects effector proteins via two Type 3 secretion systems into the host cytoplasm to usurp host cell machineries. Hence, investigating protein-protein interactions at the host-pathogen interface is essential for gaining a deeper insight into the interdependency between Salmonella and their host during infection. While more than 30 translocated effectors have been described, knowledge about their function during infection remains incomplete, and the number of proteins that are secreted during infection is likely underestimated.    In this work, l demonstrate the identification of interaction partners of known translocated effectors, maintaining the infection context and without relying on ectopic expression. The Affinity Purification Quantitative Mass Spectrometry (AP/QMS) workflow that I describe in this thesis bridges the gap between large-scale proteomics and high physiological relevance. In the process, I highlight, validate and characterize interactions occurring at endogenous levels of effector secretion. Many bacterial effector proteins displayed promiscuity and multifunctionality by targeting different host processes. Additionally, I assess effector convergence on distinct host processes, as well as effector cooperation and physical effector- effector interaction. These concepts are of groundbreaking importance to better understanding the reprograming of host pathways in different host backgrounds.    I identify cholesterol transport as a convergence point for multiple effector proteins and assess the impact of the involved effectors and targeted host proteins on cholesterol accumulation at the Salmonella-containing vacuole. In addition, I demonstrate that the Salmonella effector SteC is able to directly bind and phosphorylate formin-like proteins, thereby providing a missing link regarding the method by which Salmonella induces cytoskeletal rearrangements during infection. Furthermore, I describe an arrayed STm knockout screen in an infection context, relying on high-throughput microscopy and unbiased image feature extraction. This is used to showcase bacterial processes that are essential for infection and to predict a novel secreted effector protein, YebF, based on its feature fingerprint.    In conclusion, the work presented in this thesis provides the infection biology research community with a rich dataset of novel effector-target interactions, an adaptable and validated AP/QMS workflow, and a well-characterized strain collection for the identification of protein- protein interactions at the pathogen-host interface. Additionally, a database of unbiased phenotypic fingerprints obtained from high-throughput microscopy will be invaluable for the prediction of novel effector proteins and the dissection of host-pathogen interconnectivity. The multifaceted systems biology approach to Salmonella infection presented in this study will fuel hypothesis-driven research and provides a decisive step towards a holistic understanding of the host-pathogen interface.
%Z Author affiliations:  European Molecular Biology Laboratory (EMBL), Genome Biology Unit, Heidelberg, Germany  Joint PhD degree from EMBL and Heidelberg University, Faculty of Biosciences