DAO 130:117-129 (2018)  -  DOI: https://doi.org/10.3354/dao03255

ABSTRACT: EseN is a type III secretion system (T3SS) effector that is encoded on the Edwardsiella ictaluri chromosome and is homologous to a family of T3SS effector proteins with phosphothreonine lyase (PTL) activity, including OspF from Shigella and SpvC from Salmonella. A yeast-2-hybrid system was used to identify the major vault protein (MVP) as a specific host-cell binding partner for EseN, and the proximity ligation assay (PLA) confirmed the interaction. Similar to other pathogens, E. ictaluri invasion activates extracellular signal-regulated kinases 1 and 2 (ERK1/2) early in the infection, which are subsequently inactivated by EseN. Structurally, EseN contains a highly conserved docking motif that is required for specific binding to mitogen-activated protein kinases, such as ERK1/2, and a motif that is essential for PTL activity. Immunoblotting and immunofluorescence analyses indicate that EseN inactivates ERK1/2 by dephosphorylation in vivo in the head kidney of infected fish and ex vivo in head kidney derived macrophages. Interaction of EseN with phosphorylated ERK1/2 (pERK1/2) was also confirmed using PLA, suggesting that MVP serves as a signaling scaffold for ERK1/2 and EseN. Channel catfish Ictalurus punctatus infected with E. ictaluri strains lacking the eseN gene had reduced numbers of E. ictaluri in the tissues following infection and reduced mortality compared to fish infected with the wild-type. Our results indicate that eseN encodes a PTL domain that interacts with MVP as a possible scaffold protein and inactivates pERK1/2 to ERK1/2, resulting in increased proliferation of E. ictaluri and, ultimately, death of the host.

KEY WORDS: Edwardsiella ictaluri · Type III secretion system effectors · EseN · ERK1/2 · MVP protein · Channel catfish · Ictalurus punctatus

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Cite this article as: Dubytska LP, Thune RL (2018) Edwardsiella ictaluri type III secretion system (T3SS) effector EseN is a phosphothreonine lyase that inactivates ERK1/2. Dis Aquat Org 130:117-129. https://doi.org/10.3354/dao03255 Export citation Share:    Facebook - - linkedIn