黄萎病菌侵染对感病棉花品种叶片蛋白质组的影响

doi:10.11963/issn.1002-7807.201502009

摘要/Abstract

摘要： 为进一步探索黄萎病菌与棉花的相互作用，以感黄萎病棉花品种冀棉11为实验材料，在接种大丽轮枝菌V991 3 d后提取叶片蛋白，运用双向电泳和质谱分析技术研究黄萎病菌侵染后叶片蛋白质组学的变化。分析发现，与对照相比差异表达量在2倍以上的蛋白点有22个，其中上调表达的蛋白点10个，下调表达的蛋白点12个。质谱鉴定发现，上调表达的蛋白包括ATP合成酶β亚基，RuBisCO活化酶1，RuBisCO活化酶α2以及果糖-1，6-二磷酸醛缩酶，下调表达的蛋白包括质体叶绿素AB结合蛋白，核酮糖二磷酸羧化酶大链以及核酮糖二磷酸羧化酶小链。分别对下调表达的蛋白rubisco小亚基基因（RBCS）和叶绿素AB结合蛋白基因（cab）、上调表达的蛋白RuBisCO活化酶1基因（RCA1）进行荧光定量PCR，发现接种黄萎病菌后3个基因在mRNA水平与蛋白水平上的变化是一致的。通过分析推测，黄萎病菌通过与参与光合能量代谢的蛋白相互作用，破坏植物的光合作用，近而引起棉花叶片的萎蔫与凋零。

关键词: 棉花叶片; 黄萎病菌; 蛋白质组; 实时荧光定量

Abstract: To explore the interaction mechanism between Gossypium hirsutum and Verticillium dahliae, total protein of Jimian 11 leaves inoculated with V. dahliae for 3 days was extracted to analyze proteomic changes. Twenty-two protein spots were found showing a change of more than 2-fold between the inoculated and the mock-inoculated samples, of which 10 were up-regulated and 12 were down-regulated. The up-regulated proteins included subunit beta of ATP synthase, RuBisCO activase 1, RuBisCO activase alpha 2 and fructose-1,6-bisphosphate aldolase. The down-regulated protein spots included chloroplast chlorophyll A-B binding protein, ribulose bisphosphate carboxylase big chain and ribulose bisphosphate carboxylase small chain. Real-time polymerase chain reaction of RBCS, cab and RCA1 genes showed that expression changes of the three genes were consistent at the mRNA and protein levels. From the analyses conducted, we hypothesized that the pathogen might interact with proteins participating in photosynthetic energy metabolism, disrupting cotton photosynthesis and causing wilting and withering of cotton leaves.

Key words: cotton leaves; Verticillium dahliae; proteomics; qRT-PCR

中图分类号:

S435.621

沈凡瑞, 张文蔚, 张华崇, 刘凯, 简桂良. 黄萎病菌侵染对感病棉花品种叶片蛋白质组的影响[J]. 棉花学报, 2015, 27(2): 159-165.

Shen Fanrui, Zhang Wenwei, Zhang Huachong, Liu Kai, Jian Guiliang. Changes in the Proteome of Susceptible Cotton Leaves after Inoculation of Verticillium dahliae[J]. Cotton Science, 2015, 27(2): 159-165.

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链接本文: https://journal.cricaas.com.cn/Jweb_mhxb/CN/10.11963/issn.1002-7807.201502009

https://journal.cricaas.com.cn/Jweb_mhxb/CN/Y2015/V27/I2/159

参考文献

[1] 马存. 棉花枯萎病和黄萎病的研究[M]. 北京: 中国农业出版社, 2007. Ma Cun. Study on cotton Fusarium wilt and Verticillium wilt[M]. Beijing: China Agriculture Press, 2007. [2] 郭春燕, 詹克慧. 蛋白质组学技术研究进展及应用[J]. 云南农业大学学报, 2010, 25 (4): 583-591. Guo Chunyan, Zhan Kehui. Research progress and application in technology of proteome[J]. Journal of Yunnan Agricultural University, 2010, 25(4): 583-591. [3] Wasinger V C, Cordwell S J, Cerpa-Poljak A, et al. Progress with gene-product mapping of the mollicutes: Mycoplasma genitalium[J]. Electrophoresis, 1995, 16(1): 1090-1094. [4] 梁宇, 荆玉祥, 沈世华. 植物蛋白质组学研究进展[J]. 植物生态学报, 2004, 28(1): 114-125. Liang Yu, Jing Yuxiang, Shen Shihua. Advances in plant proteomics[J]. Chinese Journal of Plant Ecology, 2004, 28(1): 114- 125. [5] Wang Wei, Vignani R, Scali M, et al. A universal and rapid protocol for protein extraction from recalcitrant plant tissues for proteomic analysis[J]. Electrophoresis, 2006, 27(13): 2782-2786. [6] Wang Fuxin, Ma Yinping, Yang Chunlin, et al. Proteomic analysis of the sea-island cotton roots infected by wilt pathogen Verticillium dahliae[J]. Proteomics, 2011, 11(22): 4296-4309. [7] 朱龙付, 涂礼莉, 曾范昌, 等. 一种适合于cDNA文库构建的高质量棉花RNA的简单抽提法[J]. 作物学报, 2005, 31(12): 1657-1659. Zhu Longfu, Tu Lili, Zeng Fanchang, et al. An improved simple protocol for isolation of high quality RNA from Gossypium spp. suitable for cDNA library construction[J]. Acta Agronomica Sinica, 2005, 31(12): 1657-1659. [8] Schmittgen T D, Livak K J. Analyzing real-time PCR data by the comparative CT method[J]. Nature Protocols, 2008, 3(6): 1101- 1108. [9] Huffaker R C. Structure, biochemistry and physiology of leaf proteins[M] // Boulter D. Nucleic acids and proteins in plants I. Heidelberg: Springer, 1982: 370-400. [10] 赵淑芳, 胡东维. 白粉病菌侵染诱导的大麦叶肉细胞变化的超微结构与细胞化学[J]. 植物病理学报, 2003, 33(5): 444- 448. Zhao Shufang, Hu Dongwei. Ultrastructure and cytochemistry of barley mesophyll attacked by Blumeria graminis f. sp. Hordei[J]. Acta Phytopathologica Sinica, 2003, 33(5): 444-448. [11] 崔宏伟, 孙剑萍, 刘春芝, 等. 烟草叶片接种野火病菌后蛋白质表达差异[J]. 植物保护, 2012, 38(6): 7-11. Cui Hongwei, Sun Jianping, Liu Chunzhi, et al. Protein differential expression of tobacco leaves induced with Pseudomonas syringae pv. tabaci[J]. Plant Protection, 2012, 38(6): 7-11. [12] 张月娟, 赵廷昌, 杨玉文. 利用酵母双杂交系统筛选感病番茄中蔬四号与无毒蛋白AvrPto, AvrPtoB互作的蛋白[J]. 中国农业科学, 2012, 44(23): 4939-4944. Zhang YueJuan, Zhao Tingchang, Yang Yuwen. Screening of the interactive proteins of a virulent proteins AvrPto or AvrPtoB in susceptible tomato cultivar Zhongshusihao by yeast two-hybrid system[J]. Scientia Agricultura Sinica, 44 (23): 4939-4944. [13] 郑立敏. 苎麻花叶病毒编码NSP与寄主RBCS1的互作研究[D]. 福州: 福建农林大学, 2011. Zheng Limin. Interactional analysis of plant host factor RBCS1 with RaMoV NSP[D]. Fuzhou: Fujian Agricultural and Forest University, 2011. [14] 蒋德安, 陆庆, 翁晓燕, 等. 水稻剑叶衰老期RuBisCO活化酶对RuBisCO活力和光合速率的调节[J]. 浙江大学学报: 农业与生命科学版, 2000, 26(2):119-124. Jiang De'an, Lu Qing, Weng Xiaoyan, et al. Regulation of rubisco carboxylation activity and photosynthetic rate by rubisco activase during leaf senescence in rice[J]. Journal of Zhejiang University: Agriculture & Life Science, 2000, 26 (2): 119-124. [15] 李达, 李强, 关西贞, 等. 白粉菌侵染Pm21近等基因系的早期蛋白组差异研究[J]. 华北农学报, 2012, 27 (1) : 5-11. Li Da, Li Qiang, Guan Xizhen, et al. Proteomic analyses of the early Pm21 wheat near isogenic linesand their parents affected by powdery mildew[J]. Acta Agriculture Boreali-Sinica, 2012, 27(1): 5-11. [16] 李卫芳, 王忠, 韩鹰, 等. 小麦RuBisCO活化酶的纯化及其活性特性[J]. 中国农业科学, 2002, 35(8): 929-933. Li Weifang, Wang Zhong, Han Ying, et al. Purification and activity characteristics of rubisco activase from wheat leaves[J]. Scientia Agricultura Sinica, 2002, 35(8): 929-933. [17] 毛伟华, 龚亚明, 宋兴舜, 等. CMV侵染胁迫下黄瓜重要功能基因表达及代谢响应的研究[J]. 中国农业科学, 2008, 41(11): 3691-3697. Mao Weihua, Gong Yaming, Song Xingshun, et al. Expression analysis of important functional genes and pathways in cucumber leaves in response to CMV stress[J]. Scientia Agricultura Sinica, 2008, 41(11): 3691-3697. [18] 倪张林, 魏家绵. ATP合酶的结构与催化机理[J]. 植物生理与分子生物学学报, 2003, 29(5): 367-374. Ni Zhanglin, Wei Jiamian. Structure and catalytic mechanism of ATP synthetase[J]. Journal of Plant Physiology and Molecular Biology, 2003, 29(5): 367-374. [19] 龙瑞才, 杨青川, 康俊梅, 等. 紫花苜蓿果糖-1, 6-二磷酸醛缩酶基因全长克隆及分析[J]. 西北植物学报, 2010, 30(6): 1075- 1082. Long Ruicai, Yang Qingchuan, Kang Junmei, et al. Cloning and characterization of a fructose-1, 6-bisphosphate aldolase Gene in Medicago sativa L.[J]. Acta Botanica Boreali-Occidentalia Sinica, 2010, 30(6): 1075-1082. [20] Michelis R, Gepstein S. Identification and characterization of a heat-induced isoform of aldolase in oat chloroplast[J]. Plant Molecular Biology, 2000, 44(4): 487-498. [21] Umeda M, Uchimiya H. Differential transcript levels of genes associated with glycolysis and alcohol fermentation in rice plants (Oryza sativa L.) under submergence stress[J]. Plant Physiology, 1994, 106(3): 1015-1022. [22] Subramanian B, Bansal V K, Kav N N V. Proteome-level investigation of Brassica carinata-derived resistance to Leptosphaeria maculans[J]. Journal of Agricultural and Food Chemistry, 2005, 53(2): 313-324.