Functional Characterization of Putative Extracellular Cystatins, PpmEPICs, in Pathogenicity of Phytophthora palmivora on Papaya

Abstract

Phytophthora palmivora is an economically important oomycete pathogen that infects many tropical and subtropical plant species including papaya. Papaya latex is rich in cysteine proteases, especially papain. Proteolysis of non-self proteins has been established as an important aspect of host defense. Due to the high abundance of papain in papaya latex and its broad specificity in protein degradation, it is likely that P. palmivora has developed ways to interact with and inhibit papain to facilitate infection. Four putative extracellular protease inhibitors (PpmEPICs) were identified from P. palmivora next-generation sequencing data. The PpmEPICs contain the conserved domains of cysteine protease inhibitors. Phylogenetic analysis determined that PpmEPIC8 is unique to the P. palmivora genome and may be the result of evolutionary arms race between plant and pathogen. PpmEPIC8 expression is induced during infection and recombinant PpmEPIC8 inhibits papain enzymatic activity. These results suggest that PpmEPIC8 is a functional cystatin that targets papain to facilitate P. palmivora infection on papaya. Transgenic lines that overexpress PpmEPIC8 and a construct designed to silence PpmEPIC8 in P. palmivora were generated. These will be used in future research to determine if overexpression and/or silencing of PpmEPIC8 affect P. palmivora virulence on papaya.