A previously uncharacterized two-component signaling system in uropathogenic Escherichia coli coordinates protection against host-derived oxidative stress with activation of hemolysin-mediated host cell pyroptosis

Uropathogenic Escherichia coli (UPEC) deploy an array of virulence factors to successfully establish urinary tract infections. Hemolysin is a pore-forming toxin, and its expression correlates with the severity of UPEC infection. Two-component signaling systems (TCSs) are a major mechanism by which bacteria sense environmental cues and respond by initiating adaptive responses. Here, we began this study by characterizing a novel TCS (C3564/C3565, herein renamed orhK / orhR for o xidative r esistance and h emolysis k inase/ r egulator) that is encoded on a UPEC pathogenicity island, using bioinformatic and biochemical approaches. A prevalence analysis indicates that orhK / orhR is highly associated with the UPEC pathotype, and it rarely occurs in other E . coli pathotypes tested. We then demonstrated that OrhK/OrhR directly activates the expression of a putative methionine sulfoxide reductase system (C3566/C3567) and hemolysin (HlyA) in response to host-derived hydrogen peroxide (H 2 O 2 ) exposure. OrhK/OrhR increases UPEC resistance to H 2 O 2 in vitro and survival in macrophages in cell culture via C3566/C3567. Additionally, OrhK/OrhR mediates hemolysin-induced renal epithelial cell and macrophage death via a pyroptosis pathway. Reducing intracellular H 2 O 2 production by a chemical inhibitor impaired OrhK/OrhR-mediated activation of c3566-c3567 and hlyA . We also uncovered that UPEC links the two key virulence traits by cotranscribing the c3566-c3567 and hlyCABD operons. Taken together, our data suggest a paradigm in which a signal transduction system coordinates both bacterial pathogen defensive and offensive traits in the presence of host-derived signals; and this exquisite mechanism likely contributes to hemolysin-induced severe pathological outcomes.