Plant-pathogen interaction is a multifaceted process that involves two distinct forms of chemical communication referred as recognition and defense. Plant extracellular matrix (ECM) dictates early interaction between plant and pathogen and acts as regulatory hub of many biological responses and signaling networks that perceives and transmits patho-stress signal. Blast disease caused by hemibiotrophic fungus Magnoporthe oryzae is a major impediment for global crop productivity. To elucidate the role of ECM in imparting resistance against blast disease, temporal changes of ECM proteome and phosphoproteome was studied in a resistant rice cultivar upon M. oryzae infection.
Patho-stress was imposed on three-leaf-stage seedlings and tissues were harvested at early, middle and later phases of Magnaporthe attack. Temporal proteome and phosphoproteome were developed with ECM enriched fraction using TMT labeling and peptide fractionation followed by TiO2-based enrichment of phosphopeptides. Proteins were identified using LC-MS/MS analysis. Integrated global network was built to identify immunity related pathways.
Multiplex quantification of patho-stressed rice ECM proteoforms led to the identification of 336 immune-responsive proteins (IRPs) and 23 phosphopeptides mapped to 17 immune-responsive phosphoproteins (IRPPs) related to cell wall remodeling, extension, hydration and acidification linked to ROS and extracellular calcium signaling. Furthermore, the data highlighted protein phosphorylation as a critical regulatory mechanism controlling anisotropic growth and remodeling of the wall imparting immunity. Cluster and network analyses identified significant proteoform groups and hubs pointing towards the onset and context of disease signaling.
Integrated proteome and phosphoproteome analyses for the first time provides an useful insight into the complex regulatory network operating in the ECM. A comprehensive analysis not only helps to unravel the mechanism of blast resistance, but also enlist novel biomarkers for targeted genetic manipulation for food and nutrition security.