瓜类褪绿黄化病毒编码的P6蛋白亚细胞定位及致病特征分析  

Characterization of the Subcellular Localization and Pathogenicity of P6 Encoded by Cucurbit Chlorotic Yellows Virus

作　　者：涂丽琴 干射香 吴淑华[1] 任春梅[1] 程兆榜[1] 章松柏[3] 朱月林[2] 周益军[1] 季英华[1] TU Liqin;GAN Shexiang;WU Shuhua;REN Chunmei;CHENG Zhaobang;ZHANG Songbai;ZHU Yuelin;ZHOU Yijun;JI Yinghua(Institute of Plant Protection,Jiangsu Academy of Agricultural Sciences,Nanjing 210014,China;College of Horticulture,Nanjing Agricultural University,Nanjing 210095,China;College of Agriculture,Yangtze University,Jingzhou,Hubei 434025,China)

机构地区：[1]江苏省农业科学院植物保护研究所,南京210014 [2]南京农业大学园艺学院,南京210095 [3]长江大学农学院,湖北荆州434025

出　　处：《园艺学报》2021年第8期1531-1540,共10页Acta Horticulturae Sinica

基　　金：国家重点研发计划项目(2018YFD0201208);国家自然科学基金项目(32072506,31770168);国家现代农业产业技术体系建设专项资金项目(CARS-24-C-01);江苏省农业自主创新基金项目[CX(19)3108];江苏省农业科学院基金项目(6111614)。

摘　　要：以瓜类褪绿黄化病毒(cucurbit chlorotic yellows virus,CCYV)山东寿光分离物作为研究对象,对其RNA1编码的P6蛋白进行克隆,发现其基因全长为159 bp,编码1个大约6 kD的蛋白,并含1个跨膜区。进一步构建了带有YFP标签的荧光表达载体P6-YFP,浸润本氏烟叶片后利用激光共聚焦显微镜观察,发现P6-YFP在细胞质和细胞核均有分布。将P6构建到马铃薯X病毒(potato virus X,PVX)异源表达载体获得的PVX-P6接种本氏烟,发现接种PVX-P6的植株新叶伴有明显黄化、皱缩等症状,并沿叶脉出现坏死,而接种PVX空载体的植株仅有轻微的花叶,表明在PVX异源表达系统中,P6诱导寄主植物出现更严重的症状,暗示P6能增强PVX的致病性,可能是参与症状形成的致病相关因子。Cucurbit chlorotic yellows virus(CCYV)is an important whitefly-transmitted Crinivirus,which has become an emerging infectious agent of cucurbits leading to severe disease and significant economic losses.P6 is a small protein encoded by ORF2 of RNA1,one segment of CCYV,and its functions still remains unknown.In this study,P6 was cloned from an isolate of CCYV from Shouguang,Shandong Provience using specific primers by RT-PCR.The results of sequence analysis revealed that P6 gene was 159 bp in length,encoding a protein of approximately 6 kD with a transmembrane domain.To clarify the subcellular localization of P6,a YFP-tagged P6(P6-YFP)was constructed using a 35 S:YFP vector and transiently expressed in the Nicotiana benthamiana by agrobacterium infiltration.Fluorescence signal was observed in the cytoplasm and nucleus of the epidermal cells of infiltrated N.benthamiana leaves,which indicated that P6-YFP was distributed in the cytoplasm and nucleus.To test the pathogenicity of P6,a potato virus X(PVX)-based vector was introduced and PVX-P6 was constructed.Chlorotic,crumple and necrosis symptoms were observed in the emerging leaves of N.benthamiana plants inoculated by PVX-P6 and only slight mosaic symptoms was observed in the plant inoculated by PVX,which showed that heterologous expression of P6 in N.benthamiana enhanced the pathogenicity of PVX,indicating that P6 might be an important protein involved in the pathology of CCYV.

关 键 词：瓜类褪绿黄化病毒 P6蛋白 亚细胞定位 致病性 

分 类 号：S642[农业科学—蔬菜学]