多组学技术揭示葡萄叶片响应灰葡萄孢菌侵染的抗性机制
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摘要
以葡萄灰霉病高抗品种‘申丰’叶片为试验材料,采用基于液相色谱质谱联用的非标记定量蛋白质组学和非靶向定量代谢组学技术,比较了叶片在灰葡萄孢菌侵染胁迫3 d后体内蛋白质和代谢物的差异变化水平。试验结果表明,葡萄叶片中有1 374个蛋白质和33种小分子代谢物在病菌侵染后发生了1.5倍以上的差异表达(P<0.05)。功能注释和代谢通路富集等生物信息学分析发现,灰葡萄孢菌侵染对叶绿体蛋白表达影响最大,且主要集中在植病互作、植物激素与生物碱合成3条信号路径上。基于多组学数据的联合分析进一步表明,水杨酸合成与信号转导通路中的分支酸、水杨酸、异分支酸丙酮酸裂解酶pchB、转录因子TGA和病程相关蛋白PR-1表达水平显著上调。水杨酸介导的抗病信号通路的全面激活是葡萄叶片抵御灰葡萄孢菌侵染的有效手段。本研究发现为后续深入揭示葡萄灰霉病菌互作分子机制及葡萄抗病新品种选育奠定了理论研究基础。
Liquid chromatography and mass spectrometry based label-free proteomics and non-target metabolomics technology were used to study the proteome and metabolome change of disease-resistant grape cultivar ‘Shenfeng’infected with Botrytis cinerea. There were 1 374 proteins and 33 metabolites showing more than 1.5-fold changes in‘Shenfeng’leaves infected with B. cinerea, respectively. The differentially expressed proteins and metabolites were analyzed by gene ontology annotation and bioinformatics. The results showed that B.cinerea infection changed the expression level of chloroplast proteins, and mainly focused on plant and pathogen interaction, synthesis pathways of plant hormones and alkaloids. Multi-omics analysis further showed that there was a consistent increase in the expression levels of chorismic acid, salicylic acid, isochorismic pyruvate lyase pch B,transcription factor TGA and pathogenesis-related protein PR-1 in the salicylic acid-mediated disease-resistant signal transduction pathway. The full activation of salicylic acid-mediated disease resistance signaling pathway is an effective means for grape leaves to resist B. cinerea infection. The result is of great benefit to further deeply reveal the molecular mechanisms of plant-pathogen interactions and the breeding of pathogen-resistant grape varieties.
Liquid chromatography and mass spectrometry based label-free proteomics and non-target metabolomics technology were used to study the proteome and metabolome change of disease-resistant grape cultivar ‘Shenfeng’infected with Botrytis cinerea. There were 1 374 proteins and 33 metabolites showing more than 1.5-fold changes in‘Shenfeng’leaves infected with B. cinerea, respectively. The differentially expressed proteins and metabolites were analyzed by gene ontology annotation and bioinformatics. The results showed that B.cinerea infection changed the expression level of chloroplast proteins, and mainly focused on plant and pathogen interaction, synthesis pathways of plant hormones and alkaloids. Multi-omics analysis further showed that there was a consistent increase in the expression levels of chorismic acid, salicylic acid, isochorismic pyruvate lyase pch B,transcription factor TGA and pathogenesis-related protein PR-1 in the salicylic acid-mediated disease-resistant signal transduction pathway. The full activation of salicylic acid-mediated disease resistance signaling pathway is an effective means for grape leaves to resist B. cinerea infection. The result is of great benefit to further deeply reveal the molecular mechanisms of plant-pathogen interactions and the breeding of pathogen-resistant grape varieties.
引文
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[2]WILLIAMSON B,TUDZYNSKI B,TUDZYNSKI P,et al.Botrytis cinerea:the cause of grey mould disease.Molecuar Plant Pathology,2007,8(5):561-580.
[3]陈宇飞,文景芝,李立军.葡萄灰霉病研究进展.东北农业大学学报,2006,37(5):693-699.CHEN Y F,WEN J Z,LI L J.Research progress of grape grey mildew.Journal of Northeast Agricultural University,2006,37(5):693-699.(in Chinese with English abstract)
[4]FANG X P,CHEN J P,DAI L Y,et al.Proteomic dissection of plant responses to various pathogens.Proteomics,2015,15(9):1525-1543.
[5]MUKHERJEE A K,CARP M J,ZUCHMAN R,et al.Proteomics of the response of Arabidopsis thaliana to infection with Alternaria brassicicola.Journal of Proteomics,2010,73(4):709-720.
[6]YAO Y A,WANG J,MA X,et al.Proteomic analysis of Mninduced resistance to powdery mildew in grapevine.Journal of Experimental Botany,2012,63(14):5155-5170.
[7]FIEHN O.Metabolomics:the link between genotypes and phenotypes.Plant Molecular Biology,2002,48:155-171.
[8]BATOVSKA D I,TODOROVA I T,NEDELCHEVA D V,et al.Preliminary study on biomarkers for the fungal resistance in Vitis vinifera leaves.Journal of Plant Physiology,2008,165(8):791-795.
[9]VON S V,ZHANG W,KANAWATI B,et al.The Arabidopsis glucosyltransferase UGT76B1 conjugates isoleucic acid and modulates plant defense and senescence.The Plant Cell,2011,23(11):4124-4145.
[10]WINTER D,SEIDLER J,ZIV-LEHRMAN S,et al.Simultaneous identification and quantification of proteins by differential16O/18O labeling and UPLC-MS/MS applied to mouse cerebellar phosphoproteome following irradiation.Anticancer Research,2009,29(12):4949-4958.
[11]NIEHL A,ZHANG Z J,KUIPER M,et al.Label-free quantitative proteomic analysis of systemic responses to local wounding and virus infection in Arabidopsis thaliana.Journal of Proteome Research,2013,12(6):2491-2503.
[12]CAMARIES G,SCALSCHI L,VICEDO B,et al.An untargeted global metabolomic analysis reveals the biochemical changes underlying basal resistance and priming in Solanum lycopersicum,and identifies 1-methyltryptophan as a metabolite involved in plant responses to Botrytis cinerea and Pseudomonas syringae.The Plant Journal,2015,84(1):125-139.
[13]SADE D,SHRIKI O,CUADROS-INOSTROZA A,et al.Comparative metabolomics and transcriptomics of plant response to tomato yellow leaf curl virus infection in resistant and susceptible tomato cultivars.Metabolomics,2015,11(1):81-97.
[14]HAO L,WANG J X,PAGE D,et al.Comparative evaluation of MS-based metabolomics software and its application to preclinical Alzheimer’s disease.Scientific Reports,2018,8(1):9291-9300.
[15]DURSO G,PIZZA C,PIACENTE S,et al.Combination of LC-MS based metabolomics and antioxidant activity for evaluation of bioactive compounds in Fragaria vesca leaves from Italy.Journal of Pharmaceutical and Biomedical Analysis,2018,150:233-240.
[16]MAHESH H B,SUBBA P,ADVANI J,et al.Multi-omics driven assembly and annotation of the sandalwood(Santalum album)genome.Plant Physiology,2018,176:2772-2788.
[17]BLANCO-MIGUEZ A,FDEZ-RIVEROLA F,SANCHEZ B,et al.Resources and tools for the high-throughput,multi-omic study of intestinal microbiota.Briefings in Bioinformatics,2017,18(6):156-157.
[18]LEI R,JIANG H S,HU F,et al.Chlorophyll fluorescence lifetime imaging provides new insight into the chlorosis induced by plant virus infection.Plant Cell Reports,2017,36(2):327-341.
[19]LIU J,YANG J,BI H P,et al.Why mosaic?Gene expression profiling of African cassava mosaic virus-infected cassava reveals the effect of chlorophyll degradation on symptom development.Journal of Integrative Plant Biology,2014,56(2):122-132.
[20]CLEMENTE-MORENO M J,DIAZ-VIVANCOS P,RUBIOM,et al.Chloroplast protection in plum pox virus-infected peach plants by L-2-oxo-4-thiazolidine-carboxylic acid treatments:effect in the proteome.Plant Cell Environment,2013,36(3):640-654.
[21]RAGHVENDRA A S.Photosynthesis:a Aomprehensive Treatise.London,UK:Cambridge University Press,1998:72-86.
[22]SARRY J E,KUHN L,DUCRUIX C,et al.The early responses of Arabidopsis thaliana cells to cadmium exposure explored by protein and metabolite profiling analyses.Proteomics,2006,6(7):2180-2198.
[23]NAKABAYASHI R,SAITO K.Integrated metabolomics for abiotic stress responses in plants.Current Opinion in Plant Biology,2015,24:10-16.
[24]FEUSSNER I,POLLE A.What the transcriptome does not tell:proteomics and metabolomics are closer to the plants’patho-phenotype.Current Opinion in Plant Biology,2015,26:26-31.
[25]BEDNAREK P,OSBOURN A.Plant-microbe interactions:chemical diversity in plant defense.Science,2009,324:746-748.
[26]HUFFAKER A,KAPLAN F,VAUGHAN M M,et al.Novel acidic sesquiterpenoids constitute a dominant class of pathogeninduced phytoalexins in maize.Plant Physiology,2011,156:2082-2097.
[27]WILDERMUTH M C,DEWDNEY J,WU G,et al.Isochorismate synthase is required to synthesize salicylic acid for plant defence.Nature,2001,414(6863):562-565.
[28]KINKEMA M,FAN W H,DONG X N.Nuclear localization of NPR1 is required for activation of PR gene expression.The Plant Cell,2000,12(12):2339-2350.
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