Anandalakshmi R, Pruss G J, Ge X, Marathe R, Mallory A C, Smith T H, Vance V B
Department of Biological Sciences, University of South Carolina, Columbia, SC 29208, USA.
Proc Natl Acad Sci U S A. 1998 Oct 27;95(22):13079-84. doi: 10.1073/pnas.95.22.13079.
Gene silencing is an important but little understood regulatory mechanism in plants. Here we report that a viral sequence, initially identified as a mediator of synergistic viral disease, acts to suppress the establishment of both transgene-induced and virus-induced posttranscriptional gene silencing. The viral suppressor of silencing comprises the 5'-proximal region of the tobacco etch potyviral genomic RNA encoding P1, helper component-proteinase (HC-Pro) and a small part of P3, and is termed the P1/HC-Pro sequence. A reversal of silencing assay was used to assess the effect of the P1/HC-Pro sequence on transgenic tobacco plants (line T4) that are posttranscriptionally silenced for the uidA reporter gene. Silencing was lifted in offspring of T4 crosses with four independent transgenic lines expressing P1/HC-Pro, but not in offspring of control crosses. Viral vectors were used to assess the effect of P1/HC-Pro expression on virus-induced gene silencing (VIGS). The ability of a potato virus X vector expressing green fluorescent protein to induce silencing of a green fluorescent protein transgene was eliminated or greatly reduced when P1/HC-Pro was expressed from the same vector or from coinfecting potato virus X vectors. Expression of the HC-Pro coding sequence alone was sufficient to suppress virus-induced gene silencing, and the HC-Pro protein product was required for the suppression. This discovery points to the role of gene silencing as a natural antiviral defense system in plants and offers different approaches to elucidate the molecular basis of gene silencing.
基因沉默是植物中一种重要但了解甚少的调控机制。在此我们报告，一个最初被鉴定为协同病毒病介导因子的病毒序列，可抑制转基因诱导的和病毒诱导的转录后基因沉默的建立。沉默抑制因子包含烟草蚀纹马铃薯Y病毒基因组RNA的5'-近端区域，该区域编码P1、辅助成分蛋白酶（HC-Pro）和P3的一小部分，被称为P1/HC-Pro序列。采用沉默逆转试验来评估P1/HC-Pro序列对uidA报告基因转录后沉默的转基因烟草植株（T4系）的影响。在T4与四个独立表达P1/HC-Pro的转基因系杂交的后代中，沉默被解除，但在对照杂交的后代中则没有。使用病毒载体来评估P1/HC-Pro表达对病毒诱导的基因沉默（VIGS）的影响。当从同一载体或共感染的马铃薯X病毒载体中表达P1/HC-Pro时，表达绿色荧光蛋白的马铃薯X病毒载体诱导绿色荧光蛋白转基因沉默的能力被消除或大大降低。单独表达HC-Pro编码序列就足以抑制病毒诱导的基因沉默，并且抑制作用需要HC-Pro蛋白产物。这一发现表明基因沉默在植物中作为一种天然抗病毒防御系统的作用，并为阐明基因沉默的分子基础提供了不同的方法。
