Structural and Mechanistic Studies of Bacterial Chemotaxis to Complex Effectors in Two Gastrointestinal Pathogens

Kailie Franco

Bacterial chemotaxis is a widely conserved sensory system that enables directed motility along physical or chemical gradients, and many of the world's most concerning multidrug-resistant pathogens rely on it to navigate the host and establish infection. Within a host, the cues these pathogens encounter are rarely simple. They include mixtures of canonically opposing effectors, such as dietary nutrient attractants alongside repellent metabolic byproducts of host commensal bacteria, as well as reactive and short-lived species, like the reactive oxygen species (ROS) generated during host inflammation. Although chemotaxis toward individual, purified effectors such as sugars and amino acids is well characterized, much work remains to be done to understand how bacterial pathogens use their internal sense of direction to locate beneficial niches and circumvent host immunity within these complex chemical landscapes.This dissertation focuses on two gastrointestinal pathogens: Salmonella enterica serovar Typhimurium, a Gram-negative, chemotactic, intracellular bacterium that preferentially colonizes the large intestine and is the leading cause of salmonellosis; and Helicobacter pylori, a Gram-negative, chemotactic, carcinogenic bacterium that infects the human stomach, causing gastric ulcers and, in many cases, stomach cancer. Both are designated World Health Organization priority pathogens owing to rising rates of antimicrobial resistance, and together they cause substantial morbidity and mortality worldwide. Using these organisms as model systems for chemotactic pathogens, this work investigates how such bacteria resolve mixtures of competing effectors, detect reactive effectors, and structurally transduce these signals into directed movement. In the second chapter, an intestinal explant system and live imaging are used to examine how S. Typhimurium navigates conflicting fecal cues. Although the microbiota-derived compound indole is repellent in isolation, human fecal matter, despite its high indole content, is shown to be strongly attractive, because the chemoreceptor Tsr simultaneously senses indole and the attractant L-serine and directs bacteria toward the highest accessible ratio of attractant to repellent. This dual sensing is conserved across diverse Enterobacteriaceae and produces a spectrum of intermediate behaviors, termed here chemohalation. In the third chapter, the sensing of reactive effectors is examined. The chemosensory zinc-binding (CZB) chemoreceptor McpA is shown to mediate attraction of S. Typhimurium not only to hypochlorous acid but also to peroxides at picomolar-to-nanomolar concentrations. Rather than relying solely on oxidation of the conserved CZB cysteine, this response is found to depend on redox-driven changes in Zn²⁺ availability, redefining CZB domains as redox-responsive zinc switches and pointing to an ancient, broadly conserved mode of redox sensing. In the fourth chapter, the first intact structures of a soluble chemoreceptor, H. pylori TlpD, are presented in complex with its Zn²⁺ ligand. Captured in multiple conformations, these structures reveal how small changes at the CZB ligand-binding site propagate over 140 Å to the distal kinase-interacting tips, providing a high-resolution framework for long-range allostery in a widespread class of chemoreceptors important to bacterial pathogenesis. Together, these studies elucidate how chemotactic pathogens interpret complex and reactive host environments, from the ecological scale of competing chemical gradients to the atomic scale of signal transduction and advance a more translational understanding of how these agents find the niches that favor infection. With this knowledge, it may one day be possible to disable bacterial navigational systems and prevent disease caused by pathogens such as these through non-antibiotic means.

Structural and Mechanistic Studies of Bacterial Chemotaxis to Complex Effectors in Two Gastrointestinal Pathogens

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