| Tobacco mosaic virus（TMV）is a single stranded RNA virus belonging to Tobamovirus,which infects tobacco and various plants of the Solanaceae family.Quorum sensing（QS）is a term used to describe cell-to-cell communications.QS regulates bacterial behavior through sensing signal molecules produced by bacteria.Recent studies have found that QS signal molecules can be detected by microbial eukaryotic hosts to establish inter-kingdom communications.The quorum sensing molecules of AHL produced by bacteria affect the growth,development and immune response of plants.As biocontrol bacteria,photosynthesis bacterium is widely used in agricultural production due to its special physiological function and oligonucleotides,complex factors and disease resistant proteins.Rhodopseudomonas palustris,a typical purple non-sulfur photosynthetic bacterium,can use an acyl-HSL synthase to produce p-coumaroyl-HSL by using environmental p-coumaric acid.This novel AHL has rarely been reported in the study of the plant antiviral diseases.In this study,after the plants infiltrated with pC-HSL,they were inoculated again with TMV.We discussed the molecular mechanism of pC-HSL in response to TMV infection from transcriptome,proteome and Post-translational modifications,and combined with physiological and biochemical and molecular tests.The main content and conclusions are as follow:1.Rhodopseudomonas palustris quorum sensing molecule pC-HSL induces systemic resistance to TMV infection via up-regulation of Nb SIPK/Nb WIPK expressions in Nicotiana benthamiana.We investigated the effect of pC-HSL on plant immunity and have found that this QS molecule can induce a systemic resistance to TMV infection in N.benthamiana.The results show that pC-HSL treatment can prolong the activation of two mitogen-associated protein kinase（MAPK）genes（i.e.,Nb SIPK and Nb WIPK）and enhance the expressions of transcription factor WRKY8 as well as resistance gene Nb PR1 and Nb PR10,leading to an increased accumulation of reactive oxygen species（ROS）in the TMV infected plants.Our results also show that pC-HSL treatment can increase activities of two ROS-scavenging enzymes,POD and SOD.Knockdown of Nb SIPK and Nb WIPK expression in N.benthamiana plants through VIGS has shown that the pC-HSL-mediated defense response relies on the activity of Nb SIPK and Nb WIPK.At the same time,pC-HSL pre-treated also showed a strong induction of kinas activities of Nb SIPK and Nb WIPK post TMV inoculation.2.The transcriptome analysis that pC-HSL is involved in response to TMV infection in Nicotiana benthamiana.Transcriptome results show that compared with dd H2O the pC-HSL treatment,in the plant-pathogen interaction pathway the genes expression levels changed in PTI（CNGCs and Ca M/CML）and ETI（PBS1 and PIK）.The MAPK signaling pathway-plant,the expressions of SIPK/WIPK and VIP1 were similar,both have the up-regulate and down-regulate genes,and PR1,ACS6 and WRKY29 were up-regulated.Plant hormone signal transduction pathway,a total 8 plant hormones receptor genes were identified.3.The proteome analysis that pC-HSL is involved in response to TMV infection in Nicotiana benthamiana.Transcriptome analysis shows that a total of 5253 quantifiable proteins were obtained in four groups of treatments.The system leaves of plants treatmented with dd H2O+TMV/dd H2O,pC-HSL+TMV/dd H2O+TMV,pC-HSL+TMV/pC-HSL,pC-HSL/dd H2O have 577,39,592,17 up-regulated genes and 491,80,494,33down-regulated genes respectively.The GO enrichment analysis showed that after pC-HSL treatment,the accumulation of nuclear components,nucleic acid metabolism and phosphatase hydrolase activity increased,while the accumulation of transferase activity decreased.After TMV infection,intracellular and extracellular signal transduction related proteins and cytoskeleton proteins related to cell synthesis were accumulated and enhanced.In addition,Proteins related to energy metabolism such as ornithine exchange factor were also significantly accumulated.KEGG metabolic pathway enrichment showed that 32 metabolic pathways were enriched in pC-HSL+TMV treatment compared with dd H2O+TMV.Among them,relating to resistance to disease:the main up-regulate protein enrichment are Nitrogen metabolism,Photosynthesis,Phagosome,Pyruvate metabolism,Endocytosis,Pentose and glucuronate interconversions,Carbon fixation in photosynthetic organisms and Oxidative phosphorylation.The main down-regulate protein enrichment are Flavonoid biosynthesis,plant-pathogen interaction,Phenylpropanoid biosynthesis,Amino sugar and nucleotide sugar metabolism,MAPK signaling pathway.Transcriptional factor and protein interaction network analysis indicated that EF1αand Bonzai 1 May be involved in the response to TMV infection after pC-HSL pretreatment.pC-HSL pretreatment enhanced the interaction between ribosomal associated proteins and then interaction between HSP70 and HSP90/PA.pC-HSL pretreatment promoted the energy metabolism of plants after TMV infection.4.The 2-hydroxyisobutyrylation analysis that pC-HSL is involved in response to TMV infection in Nicotiana benthamiana.We identified 9823 2-hydroxyisobutyryl lysine sites on 3383 proteins in tobacco leaves,representing the first lysine 2-hydroxyisobutyrylome dataset in tobacco plants.The bioinformatics analysis indicated that 2-hydroxyisobutyrylated proteins were mostly located in the chloroplast（39.11%）and cytoplasm（29.06%）and nucleus（16.35%）and with conserved motifs flanking the 2-hydroxyisobutyryl sites.GO and KEGG pathway enrichment analysis has revealed that 2-hydroxyisobutyryl modified proteins were involved in a wide range of molecular functions and cellular processes,such as photosynthesis,glycolysis,citrate acid cycle,and response to biotic stimulus etc.Furthermore,insights into pathogen-induced changes of 2-hydroxyisobutyrylation proteome were gained from treatments with TMV virus.Exposure to TMV virus led to the decrease of 2-hydroxyisobutyryl modification level of a large number of proteins,while pC-HSL pretreatment resulted in 2-hydroxyisobutyryl modification level of many proteins increased significantly,especially the histone modification level which might promote the expression of disease resistance proteins in tobacco.5.Transgenic pC-HSL synthase gene（rpa I）of tobacco/tomato and its induced resistance to TMV.pC-HSL synthase gene（rpa I）was transferred into N.benthamiana/tomato by agrobacterium tumefaciens mediated genetic transformation.The transgenic plants were verified and the systemic resistance of the transgenic plants to TMV was analyzed.The results show that the transgenic plants showed significant anti-TMV activity compared with the control,and the expression level of pC-HSL in transgenic tobacco leaves was significantly higher than that in transgenic tomato leaves. |
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