黄瓜花叶病毒基因时序表达差异与致病相关性研究

项目名称： 黄瓜花叶病毒基因时序表达差异与致病相关性研究

项目编号： No.30800716

项目类型： 青年科学基金项目

立项/批准年度： 2009

项目学科： 轻工业、手工业

项目作者： 冯俊丽

作者单位： 浙江理工大学

项目金额： 21万元

中文摘要： 黄瓜花叶病毒（CMV）具有(+)ssRNA三分体基因组，卫星RNA通常会改变CMV引起的寄主症状；本项目以CMV-Fny，CMV-Fny?2b，CMV-Fny-satYn12和CMV-Fny-satT1为研究对象，利用荧光定量RT-PCR实验体系，分析了2b基因缺失、致弱/非致弱卫星共侵染对CMV基因组各区段复制、移动和积累的影响。此外，首创了植物miRNAs的Stem-loop Real-time RT-PCR检测方法，比较了不同病毒侵染后寄主根、茎、叶、花、果实中miRNAs及其靶标mRNAs表达谱的变化。根据得到的实验结果，并结合寄主植物不同组织的症状表现，我们认为寄主症状的严重程度与CMV的积累量之间无直接相关性，卫星RNA并非通过影响辅助病毒基因组的表达量来改变寄主症状。病毒感染会干扰寄主miRNA调控途径，引起特定mRNA表达量发生变化，产生相应寄主症状。在此过程中CMV的致病决定因子（2b基因）起着非常重要的作用，推测卫星RNA通过影响寄主miRNA调控途径来改变寄主症状。该结果为探讨病毒的致病机理提供新的着眼点，为研究卫星RNA影响辅助病毒的分子机制提供了新的理论依据。

中文关键词： 黄瓜花叶病毒；基因组RNA；卫星RNA；基因表达；致病性

英文摘要： Cucumber mosaic virus (CMV) is one of the most important plant virus of worldwide distribution. It is well known that CMV is a positive-sense RNA virus with a tripartite genome, designed RNAs 1, 2, 3 in decreasing order of their size. The nature isolates of CMV also contained a satellite RNA (satRNA), which can usually attenuate the symptoms induced by help virus. However, it has been reported some satRNAs could aggravate or had no effect on the pathogenicity of CMV. In this research, CMV-Fny, CMV-Fny?2b, CMV-Fny-satYn12, CMV-Fny-satT1 were used to analysis the effects of 2b gene and satRNAs on the accumulation of CMV genomic RNAs in different tissues of susceptible/resistant host varieties, using the established Real-time RT-PCR experiment system. In addition, we pioneered the Stem-loop Real-time RT-PCR for quantification of plant miRNAs, and compared the expression profiles of miRNAs and target mRNAs in tomato root, stem, leave, flower, green and ripe fruit upon different virus infections. Based on the obtained experimental data and combined with the symptoms of different viruses infected plants, we regarded that the pathogenicity of CMV was not determined by the abundance of viral RNAs in host tissues, and the symptoms alteration in presence of satRNA was not due to its effects on the accumulation of CMV RNAs. Nevertheless, tomato miRNAs pathways were perturbed by virus infection, indicated by the alteration of miRNAs and target mRNAs expression levels. CMV 2b has important roles in this process. We speculate satRNAs would interference tomato miRNAs pathway through unknown mechanism, and subsequently modified the symptoms of host plants. These results point new sights to recognize pathogenicity of CMV, and provided theoretical basis for understanding the effects of satRNAs on help virus.

英文关键词： Cucumber mosaic virus; genome RNA; satellite RNA; gene expression; pathogenicity