Leveraging Ecological Interactions to Unearth Cryptic Metabolites and Regulatory Pathways from Algal and Plant Microbiomes - Project Summary Bacterial natural products are essential for various industries, particularly those essential for human health. However, bacteria only synthesize a small portion of their metabolome under typical laboratory conditions due to the absence of environmental triggers that activate their production. Consequently, traditional natural product discovery methods uncover only a fraction of the metabolic potential encoded in bacterial genomes. Cryptic metabolite activators are often environmental molecules that organisms have evolved to respond to. Thus, using these niche-specific molecules in laboratory settings could potentially turn on the production of cryptic metabolites. I have demonstrated that niche-specific molecules elicit cryptic ceramide lipids in algal-microbiome bacteria, using an ecology-guided high-throughput elicitor screening (HiTES) approach, which we term Eco- HiTES. In this proposal, I will uncover the molecular mechanism for ceramide elicitation while additionally using the Eco-HiTES platform to discover cryptic metabolites from land plant root microbiomes. My preliminary work demonstrated that inhibiting bacterial respiration with the algal flavonoid myricetin in Roseovarius tolerans elicits ceramide biosynthesis, suggesting that respiratory stress directly regulates secondary metabolism. In Aim 1, I will explore respiration as a metabolic switch for cryptic metabolite production and determine the molecular mechanism for this in R. tolerans. More broadly, I will investigate whether this is a conserved mechanism for ceramide production across bacteria, while also accessing whether respiratory modulation can trigger the production of novel secondary metabolites beyond ceramides. This work will provide a mechanistically guided approach to eliciting cryptic metabolites in bacteria that experience frequent respiratory stress in their natural environments, such as plant and algal microbiomes. Plant rhizospheres offer striking examples of microbial chemical symbioses. To date, the secondary metabolomes of several plant-associated bacteria have been investigated, however the absence of environmental triggers has hampered the discovery of natural products. In Aim 2, I will apply Eco-HiTES to investigate plant-elicited cryptic secondary metabolites from the Arabidopsis root microbiome. My preliminary results show that plant metabolites elicit the production of cryptic natural products in root-associated bacteria, supporting the feasibility and promise of this approach. I will isolate and determine the structures of several cryptic metabolites from plant microbiomes and assess their pharmaceutical and agricultural potential. By bridging metabolic regulation and ecology-driven discovery, this research will provide fundamental insights into bacterial secondary metabolism while unlocking novel bioactive metabolites with potential applications in drug discovery, microbiome engineering, and sustainable agriculture. These findings will serve as a foundation for future independent research, expanding Eco-HiTES to new microbiomes and developing respiration-based strategies for natural product discovery.
