Plasticity of the cytotoxic Nep1-like protein enables promiscuity in binding to its lipid receptor glycosylinositol phosphorylceramides

Interactions between plants and pathogens pose a major challenge for plant health and agriculture. One type of threat comes from widespread cytolysins necrosis- and ethylene-inducing peptide 1–like proteins (NLPs) secreted by microbial pathogens. NLPs disrupt plant membranes through interactions with glycosylinositol phosphorylceramides (GIPCs), but the specificity of these interactions remains unclear. We studied binding and pore formation of NLPs in model membranes with GIPCs derived from dicotyledonous or monocotyledonous plants. Using the cytotoxic NLP 2 from fungus Moniliophthora perniciosa (MpNEP2), we show that membrane binding and damage extend beyond previously studied oomycetal NLPs. Furthermore, NLP_Pya from the oomycete Pythium aphanidermatum exhibits promiscuous binding with a marked preference for branched GIPC series B lipids. Molecular dynamics simulations demonstrated that the structural plasticity of NLP_Pya provides a rational basis for its interaction with a variety of GIPC headgroups. These results clarify the molecular basis of NLP cytotoxicity and emphasize their role in broad-spectrum pathogenicity.