烟草的TMV侵染耐受性与谷胱甘肽代谢途径的关系

Relationship of Tolerance to TMV and Glutathione Metabolic Pathway in Tobacco

为了深入了解烟草对烟草花叶病毒(TMV)的抗性机制,以耐受TMV感染品种豫烟8号和具有相同遗传背景对花叶病敏感的品种NC89为材料,通过对高通量测序获得的转录组数据进行生物信息学分析发现,二者接种TMV后的差异表达基因富集通路中共同存在谷胱甘肽代谢途径,且耐受性品种豫烟8号中该途径有增强的趋势。采用荧光定量PCR和分光光度法检测了两品种接种TMV前后谷胱甘肽代谢途径中关键酶谷胱甘肽S转移酶(GST)基因表达水平、酶活性以及谷胱甘肽(GSH)含量的变化。结果表明,接种TMV后两品种中GST基因表达水平升高,GST酶活性增强,GSH含量升高,耐受性品种豫烟8号增幅更明显。通过硫元素的丰缺试验研究了接种TMV对GST基因及与TMV侵染耐受性相关基因表达水平的影响,结果显示,+S和-S处理中两品种接种病毒后这些基因的表达趋势一致,-S处理的表达量低于+S处理。GST、PR1-a、HSP90和Catalase-3这些抗病相关基因都呈上调表达,其中豫烟8号接种后的表达量均高于NC89;而Psb A和PhotosystemⅡ10 k Da polypeptide 2个涉及光合作用的基因均呈现下调表达,但是接种后豫烟8号的下调幅度小于NC89。以上表明,在耐受TMV感染的品种中,谷胱甘肽代谢增强对细胞环境的氧化还原平衡有一定的促进作用,使光系统Ⅱ中的关键成分D1蛋白基因Psb A的表达相对稳定,从而减轻光合系统的破坏程度。

In order to further explore the resistance mechanism of tobacco to tobacco mosaic virus(TMV),two tobacco varieties,NC89 and Yuyan 8 which is tolerant to TMV and has the same genetic background as NC89,were used as materials,and bioinformatics analysis using transcriptome data obtained by high-throughput sequencing found that there was common response to glutathione metabolism after inoculation with TMV in the two varieties,and the response of Yuyan 8 was enhanced.We detected the change of transcription of glutathione S-transferase(GST) gene in glutathione metabolic pathway withquantitative PCR,the enzyme activity of GST and glutathione(GSH) content with spectrophotometry in the two varieties after inoculated by TMV.The results showed that the transcription level of GST gene,the enzyme activity of GST and the GSH content were increased,and the increase rate in Yuyan 8 was higher.The influence of TMV infection on the expression level of GST and other genes related to TMV tolerance was studied by the sulfur concentration experiment.The result showed that whether+S or-S,the expression trend of these genes in the two varieties was same,but the expression level in-S treatment was less than in the+S treatment.The expression of the genes related with disease resistance(GST,PR1-a,HSP90,Catalase-3) was increased after infection by TMV,and the expression level of Yuyan 8 was higher than NC89.The two genes involved in photosynthesis,Psb A and Photosystem Ⅱ 10 k Da polypeptide showed reduced expression,but the reduced rate in Yuyan 8 was less than NC89.The results above indicated that in the varieties resistant to TMV infection,enhancement of glutathione metabolic pathway was beneficial to cellular redox balance,which stabilized the expression of Psb A that encodes the key D1 protein in photosystem Ⅱ,and decreased the damage degree of TMV to photosynthetic system.