Potato virus X amplicons in arabidopsis mediate genetic and epigenetic gene silencing
Potato virus X amplicons in arabidopsis mediate genetic and epigenetic gene silencing
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DOI：
10.1105/tpc.12.3.369
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发表时间：
2000-03-01
期刊：
影响因子：
11.6
通讯作者：
Baulcombe, DC
中科院分区：
文献类型：
--
作者：
Dalmay, T;Hamilton, A;Baulcombe, DC
来自马铃薯X病毒（PVX）的扩增子转基因基于病毒基因组的修饰形式，并且是转录后基因沉默（PTGS）的有效激活剂。为了确定PVX扩增子是否激活拟南芥中的PTGS，我们使用了基于携带绿色荧光蛋白（GFP）报告基因的PVX基因组的构建体。我们对转基因表型的分析利用了以前的观察结果，表明PTGS与短的25个核苷酸的RNA种类、转基因甲基化和同源依赖的病毒抗性相关。我们还利用芜菁花叶病毒抑制基因沉默的能力，作为一种手段，解剖阶段的机制。结果表明，PVX：GFP扩增子诱导弱PTGS，并且该PTGS在GFP报告基因存在下增强。我们对这些数据的解释是，扩增子诱导的PTGS是遗传决定的，相当于PTGS机制的起始阶段。由组合的扩增子和报告基因诱导的PTGS相当于维持阶段，并且与转基因的表观遗传转化相关。遗传和表观遗传PTGS之间的区别解释了转基因剂量对PTGS的良好表征的影响，其先前已在RNA表达阈值方面进行了解释。
Amplicon transgenes from potato virus X (PVX) are based on a modified version of the viral genome and are efficient activators of post-transcriptional gene silencing (PTGS). To determine whether PVX amplicons activate PTGS in Arabidopsis, we used constructs based on the genome of PVX carrying a green fluorescent protein (GFP) reporter gene. Our analysis of the transgene phenotype exploited previous observations indicating that PTGS is associated with short 25-nucleotide RNA species, transgene methylation, and homology-dependent virus resistance. We also used the ability of turnip mosaic virus to suppress gene silencing as a means of dissecting stages of the mechanism. The results showed that a PVX:GFP amplicon induces weak PTGS and that this PTGS was enhanced in the presence of a GFP reporter gene. Our interpretation of these data is that the PTGS induced by the amplicon was genetically determined and equivalent to the initiation stage of the PTGS mechanism. The PTGS induced by the combined amplicon and reporter gene was equivalent to the maintenance stage and was associated with an epigenetic conversion of the transgene, The distinction between genetic and epigenetic PTGS explains the well-characterized effects of transgene dosage on PTGS that have been previously interpreted in terms of RNA expression thresholds.