TRYPTOPHAN DERIVATIVES IN EHEC PATHOGENESIS

The human gastrointestinal (GI) tract is inhabited by trillions of commensal bacteria that play crucial roles in human physiology. Chemical communication between microbes and their hosts underlies the basis of their associations The GI pathogen enterohemorrhagic E. coli (EHEC), responsible for outbreaks of bloody diarrhea worldwide, exploits cell-to-cell signaling between the gastrointestinal microbial flora and the hos as a means to gage and recognize the host environment. This signaling is predicated upon signaling molecules that are tryptophan derivatives in the colon such as the host neurotransmitter serotonin and the microbiota produced indole. Serotonin is a neurotransmitter that is primarily synthesized in the GI tract Serotonin is released into the lamina propria to activate peristalsis, secretion, and vasodilation. However, it has been recently reported that serotonin is also secreted into the lumen, but its function in this compartment remains unclear. Many functional GI disorders are associated with alterations in serotonin signaling, but the effect of serotonin signaling on bacterial-mediated GI disorders remains unknown. Although a number of bacteria and viruses have been show to influence serotonin signaling, no study has investigated the effect that serotonin has on bacterial pathogenesis. The bacterially produced tryptophan derivative indole is known to influence intestinal barrier function, and has also been shown to regulate virulence gene expression in EHEC. We identified the bacterial membrane bound histidine sensor kinase (HK) CpxA as a sensor of serotonin and indole. CpxA is the HK that upon autophosphorylation, phosphotranfers to the response regulator (RR) CpxR, which is a transcriptional factor. CpxA also dephosphorylates this RR, and the phosphorylation state of CpxR defines which sites it binds to on its DNA targets. Through CpxA, both serotonin and indole decrease expression of the locus of enterocyte effacement (LEE) in EHEC, which encodes a type three secretion system (T3SS), effectors and an adhesin necessary for EHEC to form lesions on enterocytes leading to intestinal disease. Both serotonin and indole inhibit CpxA's autophosphorylation, consequently decreasing the expression of its targets. Altogether, these results suggest that CpxA is an important small molecule receptor crucial for cell-to-cell signaling and inhibition of virulence of GI pathogens. Accordingly, the specific aims of this application are: 1) In vitro mechanistic studies on virulence gene regulation by tryptophan derivatives. 2) Serotonin and indole signaling at the microbiota, pathogen and host cell interface. 3) Serotonin regulation of bacterial virulence in the murine GI tract.