Increasing antibiotic resistance urgently calls for the rational design of novel antibacterial approaches. Building on unique biochemical mass spectrometry data, the present proposal aims to develop novel antimicrobial therapies and vaccines and to gain fundamental knowledge about the role of cellular infection in the development of a rare cerebrovascular disorder called Moyamoya. This will be achieved by multidisciplinary analysis of infection models with Listeria monocytogenes and Mycobacterium tuberculosis, two major types of intracellular bacterial pathogens that cause human disease. A first objective aims to develop nanobodies against Interferon Stimulated Gene 15 (ISG15), a ubiquitin like modification of the immune system that will be explored as therapeutic target for infectious and autoimmune diseases. In a second objective, multi-omics analysis of Moyamoya patient-derived samples will be used to investigate whether Moyamoya could originate from potential defects in the ISG15-mediated immune response to infection. Finally, we plan to develop novel mRNA vaccines against tuberculosis based on bacterial antigens identified by immunopeptidome analysis.