ITRAQ Protein Profile Analysis Provides Integrated Insight Into Tolerance Mechanisms Of Tobacco Responding To TMV Infection

With the wide host rang, tobacco mosaic virus(TMV) is always focus of attention due to its great harm to agricultural production, especially Solanaceae plants. Though science researchers had conducted extensive studies on gene engineering and biogenic pesticides control, so far application effect was not perfect. The prevention and control of TMV base on the mechanism of TMV and host interaction. So using absolute quantification i TRAQ technology, we compared analysis proteome of a susceptible cultivar NC89 and its tolerance line Yuyan 8, in leaf tissue, before and after the inoculation with TMV in this study, to investigate the tolerance mechanism of Yuyan 8 responding to TMV(The leaves were harvested at 0 hour and 24 hour post inoculation from NC 89 and Yuyan 8, designated as N/Nd and Y/Yd, respectively.). The main results are as follows:1、Compared analysis proteome of four samples using i TRAQ technology, a total of 2125 differentially expressed proteins were identified. Among these identified proteins, 641, 842, 612 differentially expressed proteins were defined in Y vs N, Yd vs Y, Nd vs N respectively. Subsequently, we performed Gene Ontology(GO) analysis to these proteins in Y vs N, Yd vs Y and Nd vs N. “virion part” only found in Yd and Nd indicating that TMV has transferred to upper leaves neighboring inoculated leave at 24 hour post TMV inoculation. The induced changes of molecular function category mainly involve in “transcription regulator”, “molecular transducer” and “enzyme regulator”. The induced changes of biological process category mainly reflect on “growth”, “locomotion” and “rhythmic process”.2、Venn diagram analysis of the differentially expressed proteins identified in Y vs N、Yd vs Y and Nd vs N. 260 unique proteins induced in Yuyan 8 post TMV inoculation, involving in “Epstein-Barr virus infection” and “Carbon fixation in photosynthetic organisms”; 183 unique proteins induced in NC 89 post TMV inoculation, involving in “Ribosome”, “Photosynthesis” and “Oxidative phosphorylation”; 150 proteins induced in Yuyan 8 and NC 89 post TMV inoculation, involving in “Cysteine and methionine metabolism”,“ Cyanoamino acid metabolism” and “Photosynthesis- antenna proteins”. 285 proteins changed by TMV induced in Yuyan 8 post TMV inoculation, involving in “Epstein-Barr virus infection”,“Methane metabolism”,“Pentose phosphate pathway”; 132 proteins changed by TMV induced in NC 89 post TMV inoculation, involving in “Ribosome”,“ Carbon fixation pathways in prokaryotes”.3、The accuracy of i TRAQ database was validated by detecting the changed expression of D1 and Hsp 70 using Western blotting and the enzyme activity of POD and CAT. Meanwhile, 6 differentially expressed proteins involving photosynthesis and plant defense were selected for q RT-PCR, the expression levels of all these genes matched well with our i TRAQ data, indicating that the activities of these proteins are regulated at the transcriptional level.Summing up, the tolerance mechanism of Yuyan 8 depends on degree of damange of whole photosynthesis system, antioxidant capacity, lignin biosynthesis capacity and abundance of potential resistance genes. It provides important theoretical basis for screening resistance-related proteins as marker character to new resisted flue-cured tobacco breeding.