Role of Phytophthora infestans protease inhibitors and their target tomato proteases in disease
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Series/Report no.:Plant Pathology. Graduate student poster competition, 2006
The oomycete Phytophthora infestans causes late blight, a reemerging and ravaging disease of potato and tomato. Data mining of genomic and cDNA sequences revealed that P. infestans evolved 18 extracellular protease inhibitor genes belonging to two major structural classes: (i) Kazal-like serine protease inhibitors (EPI1-14) and (ii) cystatin-like cysteine protease inhibitors (EPIC1-4). We hypothesize that P. infestans secretes proteins that inhibit host proteases and facilitate infection by protecting secreted P. infestans proteins from proteolytic degradation and/or by perturbing host defense signaling cascades that include proteolytic steps. Previous biochemical studies showed that both recombinant EPI1 and EPI10 inhibit and interact with the pathogenesis-related (PR) P69B subtilisin-like serine protease of tomato. We extended our biochemical analyses to EPIC1 and EPIC2 and found that EPIC1 and EPIC2B were unstable in tomato apoplastic fluids and were degraded by tomato P69B but EPI1 protected both proteins from degradation. Affinity purified P69B was sufficient to degrade EPIC1 and EPIC2B but not EPI1a, suggesting selectivity in degradation by P69B. Coimmunoprecipitation experiments revealed that EPIC2 interacts with a novel papain-like extracellular cysteine protease, termed Phytophthora Interacting Protein 1 (PIP1). The interaction was further confirmed by coimmunoprecipitation using in planta expressed PIP1 protein. Altogether, our findings suggest that a cascade of inhibition of host proteases initiated by EPI1 occurs in the tomato apoplast during infection by P. infestans. In addition, this study provides biochemical evidence for a direct contribution of the PR protein P69B subtilase to defense through the degradation of proteins from invading pathogens.
National Research Initiative of the USDA Cooperative State Research, Education and Extension Service, grant number # 2005-35319-15305.
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