Necrotrophic fungal pathogen Cochliobolus miyabeanus causes brown spot disease in rice leaves upon infection, resulting in critical rice yield loss. To better understand the rice-C. miyabeanus interaction, we employed proteomic approaches to establish differential proteomes of total and secreted proteins from the inoculated leaves. The 2DE approach after PEG-fractionation of total proteins coupled with MS (MALDI-TOF/TOF and nESI-LC-MS/MS) analyses led to identification of 49 unique proteins out of 63 differential spots. SDS-PAGE in combination with nESI-LC-MS/MS shotgun approach was applied to identify secreted proteins in the leaf apoplast upon infection and resulted in cataloging of 501 unique proteins, of which 470 and 31 proteins were secreted from rice and C. miyabeanus, respectively. Proteins mapped onto metabolic pathways implied their reprogramming upon infection. The enzymes involved in Calvin cycle and glycolysis decreased in their protein abundance, whereas enzymes in the TCA cycle, amino acids, and ethylene biosynthesis increased. Differential proteomes also generated distribution of identified proteins in the intracellular and extracellular spaces, providing a better insight into defense responses of proteins in rice against C. miyabeanus. Established proteome of the rice-C. miyabeanus interaction serves not only as a good resource for the scientific community but also highlights its significance from biological aspects.
Necrotrophic pathogen Cochliobolus miyabeanus (C. miyabeanus) causes rice brown leaf spot disease and drastically affects the yield and quality of rice grains. However, the molecular mechanism of rice-C. miyabeanus remains poorly understood due to the limited research conducted on this pathosystem. To elucidate the molecular mechanism of rice-C. miyabeanus, a transcriptome analysis was conducted from in vitro and in planta grown C. miyabeanus. This analysis led to the identification of a total of 24,060 genes of which 426 in vitro and 57 in planta expressed genes were predicted to encode for secretory proteins. As these 57 genes were specifically expressed in planta and were predicted to be secretory in nature, these were consider as putative effectors, highlighting their possible roles in the fungal pathogenicity. Notably, among these putative effectors, CmXyn1 which encodes a glycosyl hydrolase 11 displayed the highest expression level under in planta conditions and was thus selected for further functional characterization. Interestingly, the extracellular expression of CmXyn1 transiently induced cell death in Nicotiana benthamiana leaves, while intracellular expression was comparatively lesser effective. In addition, transcriptome analysis on rice leaves during C. miyabeanus infection and comparing it to the rice leaf transcriptome data obtained during hemibiotrophic pathogen Magnaporthe oryzae infection led to the discovery of 18 receptors/receptor-like kinases that were commonly expressed in response to both pathogens, indicating their key roles in rice defense response. Taken together, our findings provide new insights into rice-C. miyabeanus interaction as well as the unique and common defense responses of rice against hemibiotroph and necrotroph model systems.
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