Dysregulation of Cytosolic c-di-GMP in Edwardsiella piscicida Promotes Cellular Non-Canonical Ferroptosis

Programmed cell death plays an important role in modulating host immune defense and pathogen Infection. Ferroptosis is a type of inflammatory cell death induced by intracellular iron-dependent accumulation of toxic lipid peroxides. Although Ferroptosis has been associated with Cancer and other sterile diseases, very little is known about the role of Ferroptosis in modulating host-pathogen interactions. We show that accumulation of the secondary messenger bis-(3',5')-cyclic dimeric GMP (c-di-GMP) in the pathogenic bacterium Edwardsiella piscicida (E. piscicida) triggers a non-canonical Ferroptosis pathway in infected HeLa cells. Moreover, we observed that the dysregulation of c-di-GMP in E. piscicida promotes iron accumulation, mitochondrial dysfunction, and production of Reactive Oxygen Species, all of which that can be blocked by iron chelator. Importantly, unlike classical Ferroptosis that is executed via excess lipid peroxidation, no lipid peroxidation was detected in the infected cells. Furthermore, lipoxygenases inhibitors and lipophilic antioxidants are not able to suppress morphological changes and cell death induced by E. piscicida mutant producing excess c-di-GMP, and this c-di-GMP dysregulation attenuates Bacterial virulence in vivo. Collectively, our results reveal a novel non-canonical Ferroptosis pathway mediated by Bacterial c-di-GMP and provide evidence for a role of Ferroptosis in the regulation of pathogen Infection.