Uncovering aryl polyene biology to identify new drug targets in Gram-negative bacterial pathogens

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ABSTRACT We are rapidly running out of options to combat infectious diseases in an era that is marked by a rise in antibiotic resistant bacterial pathogens. The discovery rate of novel antibiotics is not keeping up with this pressing need and thus we have to identify alternative drug targets and develop complementary elimination strategies. This is a particularly challenging task for infections caused by Gram-negative bacteria because their outer membrane is impermeable to many compounds. We discovered an aryl polyene (APE) virulence factor, widespread among Gram-negative pathogens, and here propose to characterize its biosynthesis and use it as a target for the development of elimination strategies. APEs are the product of the most abundant family of biosynthetic gene clusters (BGCs) in our global in silico analysis of bacterial genomes. They are present in several multidrug- resistant pathogens, and our preliminary data shows that APEs are esterified to a yet unidentified anchor molecule in the Gram-negative outer membrane. Furthermore, we discovered that APEs serve as virulence factors that protect their producers from host innate immune clearance mechanisms. Our long-term goal is to understand APE biology on a molecular, functional and evolutionary level. The current proposal is centered on the hypothesis that APE biosynthesis and cell envelope localization provide targets for specific antivirulence strategies against Gram-negative pathogens. We will test this by first characterizing the function of individual genes within the APE outer membrane localization pathway and identifying the APE-containing anchor molecule. Next, we will determine the environmental conditions that control regulation of APE expression using a forward genetics approach. Finally, we will develop interference strategies to inhibit APE production, using a targeted inhibitor of the initiation enzyme in the biosynthetic pathway. Our proposed project will yield mechanistic insights into the biology of APEs, as well as identify small molecule tool compounds that can be used to study APE biology and enable alternative pathogen eradication strategies.