Microorganisms live in communities in nature. The structure of microbial communities is shaped by the interactions with neighboring organisms. These interactions are often mediated by small molecules called natural products. Biosynthesis of these natural products is tightly regulated and can be induced by several factors including microbial interactions, quorum sensing, and the presence of chemical inducers such as antimicrobial compounds, or specific metabolic precursors required for biosynthesis. Description of circumstances that activate natural product biosynthesis are crucial to unravel mechanisms of induction at the molecular level and to determine their ecological functions. The overall objectives of our research program is to understand how natural product biosynthesis is activated by specific microbial and chemical environments as well as to mimic these environments in vitro enabling discovery of new natural products and their biosynthetic pathways. We will utilize mucus-dwelling bacteria from human airways, and marine corals, both of which are sources of unique natural products to investigate natural product- mediated interspecies interactions. In the project period, we will develop and apply mass spectrometry-based workflows to 1) elucidate the role of natural products and quorum sensing in interspecies interactions in host- relevant chemical and physical environment via the use of synthetic human mucus, 2) understand how chemical inducers such as antibiotics shape microbial community structure of mucus microbiomes, and 3) characterize natural product chemical space of mucus-dwelling bacteria derived from corals, an understudied resource of bioactive natural products. The methods described in this proposal enhance detection and characterization of natural product chemical space. Thus, this work will identify new natural products with antibiotic potential, which are needed in the face of increasing antibiotic resistance and declining rate of discovery of natural products- based antibiotics. The observations from this study will illuminate the mechanistic basis for natural product-mediated microbial interactions and natural-product mediated structuring of microbial communities.