A novel Phytophthora-specific putative lipid-binding protein triggers plant immunity.

Abstract

Plants recognize conserved pathogen-derived molecules to activate pattern-triggered immunity (PTI), yet many secreted proteins from oomycete pathogens remain functionally uncharacterized. Here, we identified PcCF16, a previously uncharacterized secreted protein from Phytophthora capsici, through activity-guided fractionation of the pathogen secretome. PcCF16 induced cell death and PTI-associated gene expression in Nicotiana benthamiana, and purified recombinant PcCF16 triggered reactive oxygen species production across multiple plant species. PcCF16-induced cell death was independent of the co-receptors BAK1 and SOBIR1, while defense gene activation remained SOBIR1-independent but was partially dependent on BAK1. Recombinant PcCF16 pretreatment enhanced resistance to P. capsici, while P. capsici lines constitutively expressing PcCF16 exhibited reduced virulence. Structure-function analysis identified residues 202-291 as the shortest fragment tested that was sufficient for cell death induction, whereas N-glycosylation enhanced PcCF16 accumulation and activity but was not strictly required. Structural analyses revealed similarity between PcCF16 and members of the tubular lipid-binding protein (TULIP) superfamily and predicted internal hydrophobic cavities. Consistent with these features, PcCF16 preferentially associated with phosphatidic acid in vitro. Despite substantial sequence divergence, PcCF16 homologs retained similar predicted protein folds but differed in cell death-inducing activity, with surrounding protein context influencing the activity of the conserved C-terminal region. PcCF16 was preferentially expressed during the necrotrophic phase of infection, suggesting a biological function distinct from that of typical early-stage immune elicitors. Together, these findings identify PcCF16 as a Phytophthora-specific secreted immune elicitor with potential lipid-associated functions and reveal an unusual link between late-stage pathogen biology and plant immune activation.