Functional characterization of an MPEG1 homologue from Paralichthys olivaceus

MPEG1 (macrophage-expressed gene 1) is a membrane attack complex/perforin (MACPF) superfamily protein implicated in antimicrobial defense, yet its functional mechanisms in teleost fish remain poorly understood. In this study, we identified and characterized an MPEG1 homologue (PoMPEG1) from Paralichthys olivaceus (Japanese flounder). PoMPEG1 encodes a 710-amino-acid protein comprising a conserved MACPF-P2 ectodomain, a transmembrane helix, and a short cytosolic tail, and localizes to lysosomal compartments. PoMPEG1 was ubiquitously expressed in flounder tissues and strongly induced upon bacterial infection. The cytosolic tail of PoMPEG1 contains a cationic segment with potent bactericidal activity against Vibrio harveyi. Structural and functional analyses revealed that this activity depends on an intact α-helical conformation and is inhibited by an adjacent anionic segment, suggesting a regulatory mechanism controlling its activation. In contrast, the ectodomain of PoMPEG1 likely mediated defense against the intracellular pathogen Edwardsiella piscicida. This is supported by the observation that PoMPEG1 knockdown increased the intracellular bacterial load, whereas the introduction of the intact protein, but not its ectodomain-deficient mutants, reduced it. Together, these findings demonstrate that PoMPEG1 functions as a dual-action immune effector employing distinct mechanisms to combat extracellular and intracellular pathogens, providing new insights into MPEG1-mediated host defense in teleost fish.