RNA沉默在植物生物逆境反应中的作用

RNA沉默是真核生物共有的基因表达调节机制和防御机制。在植物RNA沉默中,一些小RNAs,如微小RNAs和小干扰RNAs,在植物防御病毒、细菌或食草动物的反应中具有重要作用。为了抑制宿主的RNA沉默系统,植物病毒或细菌进化出了在RNA沉默不同阶段起作用的病毒沉默抑制子或细菌沉默抑制子,来克服寄主的RNA沉默反应。文章就植物RNA沉默、病毒沉默抑制子、细菌沉默抑制子及其相关防御反应的一些新进展做一概述。     

植物抗病毒分子机制

在与植物病毒的长期斗争中,植物进化出多种抗病毒机制,其中RNA沉默和R基因介导的病毒抗性是最受人们关注的两种机制.一方面,RNA沉默是植物抵抗病毒侵染的重要手段.植物在病毒侵染过程中可形成病毒来源的双链RNA,经过DCL蛋白的切割、加工形成sRNA,与AGO蛋白结合形成RISC指导病毒RNA的沉默,用于清除病毒.相应地,病毒在与植物的竞争中进化出RNA沉默抑制子,抑制宿主RNA沉默系统以逃避宿主RNA沉默抗病毒反应,增强致病能力.另一方面,植物也进化出R基因介导植物对包括病毒在内的多类病原的抗性.R蛋白直接或间接识别病毒因子,通过一系列的信号转导途径激活植物防御反应,限制病毒的进一步侵染.对植物抗病毒的研究有助于人们对植物抗病分子基础的理解,有重要的科学意义和潜在应用价值.本文综述了植物抗病毒分子机制的重要进展.     

病毒编码的转录后沉默抑制蛋白

转录后水平沉默是植物体内天然存在的抗病毒防御机制,该机制基于双链RNA诱导的RNA干扰过程特异性降解植物病毒在体内复制时产生的双链RNA中间体,从而终止病毒的复制及扩散。而病毒在长期与植物体的互作进化过程中通过表达产生沉默抑制蛋白,也建立了针对寄主沉默机制的抗“防御”系统。     

RNA干扰与抗病毒感染

RNA干扰(RNAi)是由小干扰RNA(siRNA)引发的生物细胞内同源基因的转录后基因沉默(PTGS)现象,是一种古老的生物抵抗外在感染的防御机制。RNAi因其在维持基因组稳定、调控基因表达和保护基因组免受外源核酸侵入等方面发挥的重要作用,已被广泛用于探索基因功能、基因治疗和新药的研发。外源导入siRNA引发的RNAi可以特异性抑制病毒的复制与感染,为抗病毒感染治疗开辟了一条新的途径。     

病毒基因沉默抑制子及其作用机制

基因沉默或RNA 沉默是植物、真菌以及无脊椎动物抵御病毒侵染的重要机制, 为了应对这种机制, 病毒编码RNA 沉默抑制子抑制宿主的基因沉默, 抵抗由基因沉默介导的宿主对病毒的抗性. 病毒编码的RNA 沉默抑制子, 又被称为病毒基因沉默抑制子, 广泛存在于各种植物RNA 病毒和DNA 病毒以及部分动物病毒中. 近年来, 针对病毒基因沉默抑制子作用机制的研究表明, 病毒基因沉默抑制子通过与宿主基因沉默通路中的RNA 或者关键蛋白分子相互作用, 发挥抑制宿主对病毒的抗性以及干扰宿主正常的基因表达调控的功能. 由于植物基因沉默通路的复杂性, 病毒基因沉默抑制子的作用机制也是复杂而多样的.     

RNA干扰与植物抗病毒

RNA干扰是多种生物体内由双链RNA介导的同源mRNA降解现象,是植物体内天然的抗病毒机制。然而病毒在长期进化过程中也获得了通过编码沉默抑制蛋白来对抗植物体RNAi系统的能力。本文对RNA干扰过程、病毒编码的沉默抑制蛋白及利用干扰技术进行抗病毒基因工程研究进行简要综述。     

RNA沉默技术及其在烟草抗病毒研究中的应用

RNA沉默技术是一项基因沉默新技术。在抗病毒研究中，人为地将与病毒或宿主基因（宿主基因编码的蛋白质对病毒很重要而对宿主本身作用很小或不起作用）同源的双链RNA（double strand RNA，dsRNA）导入生物体内，引起与其同源的基因发生沉默，从而抑制病毒复制，达到抗病毒的目的。因此，RNA沉默技术技术在抗病毒研究中倍受关注，并取得了显著成绩。主要对RNA沉默技术的相关知识及其在烟草抗病毒中的应用进展作一综述。     

RNA干扰分子的制作

小干扰RNA是一种能够在各种生物体和细胞(包括蠕虫、果蝇、植物、哺乳动物)中减弱基因表达的有效工具。在哺乳动物中转染的siRNA能够抑制特殊基因的表达,这已经证明是探索基因功能、基因敲除、抗病毒研究、基因治疗的有效方法。简单、有效、特异性地抑制基因的表达具有巨大的科学、商业和医学治疗价值。如何设计和制作siRNA是影响RNA干扰效率的一个很重要的方面。本文就siRNA的设计和制作等方面作扼要的介绍。     

RNA 沉默的病毒抑制子

RNA 沉默是一种在真核生物体内普遍保守的、通过核酸序列特异性的相互作用来抑制基因表达的调控机制 . RNA 沉默的一种重要生物学效应是防御病毒的侵染,而针对寄主的这种防御机制,许多植物病毒已演化通过编码 RNA 沉默的抑制子来克服这种防御反应 . 目前,已从植物、动物和人类病毒中鉴定了 20 多种 RNA 沉默的抑制子,围绕抑制子的鉴定和作用机理研究已成为病毒学研究的一个热点 . 对 RNA 沉默抑制子的发现、鉴定方法、作用机理及与病毒病症状形成的关系、动物病毒的沉默抑制子等方面的最新进展做了综述,并对沉默抑制子的应用和存在的问题进行了讨论 .     

水稻瘤矮病毒S12编码第2个RNA沉默抑制子

RNA沉默是一种保守而普遍存在于真核生物体内用于调控基因表达和发育的一种机制,也是植物抵抗病毒侵染和转录的一种防御机制.作为一种反防御机制,许多植物病毒编码沉默抑制子.本研究利用农杆菌共浸润的方法发现,RGDV基因组S12片段编码的Pns12蛋白是该病毒的第2个RNA沉默抑制因子.Pns12能够抑制正义链RNA诱发的局...     

Suppressors of RNA silencing encoded by plant viruses and their role in viral infections

RNA silencing as a robust host defense mechanism against plant viruses is generally countered by virus-encoded silencing suppressors. This strategy is now increasingly recognized to be used by animal viruses as well. We present here an overview of the common features shared by some of the better studied plant viral silencing suppressors. We then briefly describe the characteristics of the few reported animal viral suppressors, notably their extraordinary ability of cross-kingdom suppression. We next discuss the basis for biased protection of viral RNA and subviral parasites by silencing suppressors, the link between movement and silencing suppression, the influence of temperature on the outcome of viral infection and the effect of viral silencing suppressors on the microRNA pathway.     

假马鞭曲叶病毒和中国胜红蓟黄脉病毒C4蛋白是RNA沉默的抑制子

为了研究中国胜红蓟黄脉病毒(Ageratum yellow vein Chin virus,AYVCNV)和假马鞭曲叶病毒(Stachytarpheta leafcurl virus,StaLCV)C4蛋白的功能,利用烟草脆裂病毒(Tobacco rattle virus,TRV)载体在本氏烟(Nicotianabenthamiana)中分别表达了这两种病毒的C4蛋白,结果发现它们均能在本氏烟中引起类似于病毒侵染的症状,推测AYVCNV和StaLCV的C4蛋白是病毒的致病因子;在RNA沉默的抑制试验中,AYVCNV和StaLCV的C4蛋白均能够在表达gfp基因的转基因本氏烟(16c)上抑制由gfp基因正义链引起的基因沉默的建立,证明它们都是RNA沉默的抑制子。     

Small RNA binding is a common strategy to suppress RNA silencing by several viral suppressors

RNA silencing is an evolutionarily conserved system that functions as an antiviral mechanism in higher plants and insects. To counteract RNA silencing, viruses express silencing suppressors that interfere with both siRNA- and microRNA-guided silencing pathways. We used comparative in vitro and in vivo approaches to analyse the molecular mechanism of suppression by three well-studied silencing suppressors. We found that silencing suppressors p19, p21 and HC-Pro each inhibit the intermediate step of RNA silencing via binding to siRNAs, although the molecular features required for duplex siRNA binding differ among the three proteins. None of the suppressors affected the activity of preassembled RISC complexes. In contrast, each suppressor uniformly inhibited the siRNA-initiated RISC assembly pathway by preventing RNA silencing initiator complex formation.     

Effects and side-effects of viral RNA silencing suppressors on short RNAs

In eukaryotes, short RNAs play a crucial regulatory role in many processes including development, maintenance of genome stability and antiviral responses. These different but overlapping RNA-guided pathways are collectively termed 'RNA silencing'. To counteract an antiviral RNA silencing response, plant viruses express silencing suppressor proteins. Recent results have shown that silencing suppressors operate by modifying the accumulation and/or activity of short RNAs involved in the antiviral response. Because RNA silencing pathways intersect, silencing suppressors can also inhibit other short-RNA-regulated pathways. Thus, suppressors contribute to viral symptoms. These findings fuel further research to test whether certain symptoms caused by animal viruses are also manifestations of altered RNA regulatory pathways.     

Structural insights into the arms race between host and virus along RNA silencing pathways in Arabidopsis thaliana

RNA silencing refers to a conserved sequence‐specific gene‐regulation mechanism mediated by small RNA molecules. In plants, microRNA (miRNA) and small interfering RNA (siRNA) represent two major types of small RNA molecules which play pivotal roles in plant developmental control and antiviral defences. To escape these plant defences, plant viruses have encoded a vast array of viral suppressors of RNA silencing (VSRs) to attack the host antiviral silencing pathway by interfering with small RNA processing, RNA‐induced silencing complex (RISC) assembly, viral mRNA cleavage etc. Transgenic plants expressing distinct VSRs often show developmental aberrations that resemble the phenotype of miRNA‐deficient mutants, implying a potential intrinsic link between VSRs and the miRNA pathway (at least in Arabidopsis thaliana) even though their pathogenic mechanisms remain largely unknown. In this review, we summarise our current structural understandings of the arms race between the host and virus along the RNA silencing pathway in A. thaliana by focusing on several important ribonucleoprotein (RNP) structures involved in RNA silencing and unique structural features adopted by VSRs.     

Viral induction and suppression of RNA silencing in plants

RNA silencing in plants and insects can function as a defence mechanism against invading viruses. RNA silencing-based antiviral defence entails the production of virus-derived small interfering RNAs which guide specific antiviral effector complexes to inactivate viral genomes. As a response to this defence system, viruses have evolved viral suppressors of RNA silencing (VSRs) to overcome the host defence. VSRs can act on various steps of the different silencing pathways. Viral infection can have a profound impact on the host endogenous RNA silencing regulatory pathways; alterations of endogenous short RNA expression profile and gene expression are often associated with viral infections and their symptoms. Here we discuss our current understanding of the main steps of RNA-silencing responses to viral invasion in plants and the effects of VSRs on endogenous pathways. This article is part of a Special Issue entitled: MicroRNAs in viral gene regulation.     

Crystal structure of p19--a universal suppressor of RNA silencing

RNA silencing in plants has an antiviral role and, consequently, plant viruses encode counter-defensive suppressor proteins that block this process. The recently reported crystal structure of two Tombusvirus suppressor proteins reveals a novel RNA-binding structure and illustrates precisely how the silencing mechanism is blocked. These suppressor protein structures, combined with molecular analyses of their effects in animal and plant cells, are informative about RNA silencing mechanisms. They also suggest various ways that Tombusvirus suppressors can be used to investigate RNA silencing in plants and animals.     

Polerovirus protein P0 prevents the assembly of small RNA‐containing RISC complexes and leads to degradation of ARGONAUTE1

RNA silencing plays an important role in plants in defence against viruses. To overcome this defence, plant viruses encode suppressors of RNA silencing. The most common mode of silencing suppression is sequestration of double‐stranded RNAs involved in the antiviral silencing pathways. Viral suppressors can also overcome silencing responses through protein–protein interaction. The poleroviral P0 silencing suppressor protein targets ARGONAUTE (AGO) proteins for degradation. AGO proteins are the core component of the RNA‐induced silencing complex (RISC). We found that P0 does not interfere with the slicer activity of pre‐programmed siRNA/miRNA containing AGO1, but prevents de novo formation of siRNA/miRNA containing AGO1. We show that the AGO1 protein is part of a high‐molecular‐weight complex, suggesting the existence of a multi‐protein RISC in plants. We propose that P0 prevents RISC assembly by interacting with one of its protein components, thus inhibiting formation of siRNA/miRNA–RISC, and ultimately leading to AGO1 degradation. Our findings also suggest that siRNAs enhance the stability of co‐expressed AGO1 in both the presence and absence of P0.     

RNAi的作用机制及其在抗病毒领域的应用

RNA干扰是外源性或内源性双链RNA诱发的mRNA水平上的基因沉默机制。RNA技术具有高效性、特异性。最近将RNA干扰应用于许多病毒性疾病的治疗研究均取得了显著的基因沉默效果,为病毒的预防和治疗开辟了一条新途径。就RNA干扰作用机制及抗病毒效应作一综述。