Translational control by non-coding RNA in Marek's disease herpesvirus: implications in oncogenesis and exploitation in bioengineering

马立克氏病疱疹病毒中非编码 RNA 的翻译控制：对肿瘤发生和生物工程开发的影响

• 批准号: BB/H010696/1
• 负责人: Venugopal Nair
• 金额: $ 56.49万
• 依托单位: The Pirbright Institute
• 依托单位国家: 英国
• 项目类别: Research Grant
• 财政年份: 2010
• 资助国家: 英国
• 起止时间: 2010 至 无数据
• 项目状态: 已结题
• 来源: https://gtr.ukri.org/projects?ref=BB%2FH010696%2F1
• 关键词: Translational; control; non; coding; RNA

Marek's Disease (MD) is a widespread lymphoid neoplastic disease with tumours caused by infection with the Marek's Disease Virus (MDV). MD is one of the major poultry health and welfare problems throughout the world, with losses estimated to be up to US$ 2,000 million despite more than 20 billion doses of the MD vaccines used annually. MDV has proven to be a valuable model for understanding some of the principles of oncogenesis. It provides a well defined small-animal model of general tumorigenesis, and virus-induced lymphomagenesis in particular. The reproducible kinetics of disease induction and progression, in a natural virus-host system, make MDV unique allowing studies that are impossible to perform in other non-natural models of herpesvirus-induced oncogenesis. Studies of virus-infected cells have been a prominent source of information on the mechanism of translational control. As obligate intracellular parasites, viruses depend on cells for their replication. Nowhere is this dependency seen more clearly than in the translation system, as viruses lack a translational apparatus. Viruses have evolved ways to gain a translational advantage for their mRNAs. This is particularly crucial during the very early stages of viral infection where immediate-early transcripts are produced and need to be translated. Internal ribosome entry site (IRES)-mediated translation initiation is one of the strategies that viruses use during time of cellular stress and when cap-dependent translation might be inhibited. We have discovered that an immediate-early transcript from MDV has two IRESes. The first IRES is contained within the 5' leader sequence and controls the translation of the downstream open reading frame that encode for the pp14 lytic protein. The 5' leader IRES appears to be regulated by base-pairing with sequences within the 18S rRNA and by a ribosomal protein within the 40S ribosomal subunit. The second IRES is contained within the intercistronic region (ICR) and controls the translation of the second protein known as RLORF9. The ICR IRES activity appears to be controlled by a set of viral small non-coding RNA known as micro-RNAs. This transcript is known to be associated with oncogenicity and the maintenance of latency of MDV transformed cells. This transcript is within a locus that undergoes specific sequence amplification during viral attenuation. This sequence amplification is the only validated marker that always correlates with viral attenuation. What benefits this research proposal will bring to the field of MDV oncogenesis in particular and to translational control in general? Preliminary work has allowed us to put forward novel concepts in translational control with direct impact on the understanding of MDV oncogenesis. The availability of an infectious BAC clone of the highly oncogenic RB-1B strain will allow us to use recombination-mediated genetic engineering to investigate the biological function of the 1.8-kb transcript in cell culture and in chicken. This highly reproducible disease model in the natural host will be used to obtain insights into the mechanisms of herpesvirus-induced lymphomagenesis for designing better vaccines. Detailed analysis of the structure and the function of the two IRESes within the 1.8-kb transcript will allow us to dissect fundamental mechanisms in translation such as miRNA-IRES mediated translational control, ribosomal filtering and IRES allostery. The results will also be exploited in designing novel multicistronic vectors to allow the expression of a gene of interest inside the target cells with the aid of a reporter, a drug selection marker, or a suicidal gene, all expressed from the same transcript. The emerging application of viruses in cancer, gene therapy and delivery mandate an understanding of the interplay between viruses and the host mechanisms. The study of translational control impinges on several fields including biotechnology, cell biology, Medicine and Agriculture.

马立克氏病（MD）是由马立克氏病病毒（MDV）感染引起的一种广泛的淋巴样肿瘤疾病。口蹄疫是全世界家禽健康和福利的主要问题之一，尽管每年使用的口蹄疫疫苗超过200亿剂，但估计损失高达20亿美元。MDV已被证明是理解肿瘤发生的一些原理的一个有价值的模型。它提供了一个定义良好的一般肿瘤发生的小动物模型，特别是病毒诱导的淋巴瘤发生。在自然病毒-宿主系统中，疾病诱导和进展的可复制动力学使MDV独一无二，使研究不可能在其他非自然疱疹病毒诱导的肿瘤发生模型中进行。对病毒感染细胞的研究已成为翻译控制机制的重要信息来源。作为专性细胞内寄生虫，病毒依靠细胞进行复制。这种依赖性在转译系统中表现得最为明显，因为病毒缺乏转译装置。病毒已经进化出获得mrna翻译优势的方法。这在病毒感染的非常早期阶段尤其重要，在这个阶段产生了立即早期转录本，需要进行翻译。内部核糖体进入位点（IRES）介导的翻译起始是病毒在细胞应激和帽依赖性翻译可能被抑制时使用的策略之一。我们已经发现，MDV的立即早期转录本有两个IRESes。第一个IRES包含在5'先导序列中，并控制编码pp14裂解蛋白的下游开放阅读框的翻译。5'先导IRES似乎通过与18S rRNA内序列的碱基配对和40S核糖体亚基内的核糖体蛋白进行调节。第二个IRES包含在顺胞间区（ICR）中，并控制第二个蛋白质RLORF9的翻译。ICR IRES活性似乎是由一组被称为微RNA的病毒小非编码RNA控制的。已知该转录物与MDV转化细胞的致癌性和维持潜伏期有关。该转录本位于病毒衰减过程中经历特定序列扩增的位点内。该序列扩增是唯一经过验证的始终与病毒衰减相关的标记。这项研究计划将给MDV致癌领域带来什么好处，特别是在转译控制方面？初步的工作使我们能够提出翻译控制的新概念，直接影响对MDV肿瘤发生的理解。高致癌性RB-1B菌株的感染性BAC克隆的获得将使我们能够利用重组介导的基因工程来研究1.8 kb转录本在细胞培养和鸡体内的生物学功能。这种在自然宿主中高度可复制的疾病模型将用于深入了解疱疹病毒诱导的淋巴瘤形成机制，以设计更好的疫苗。对1.8 kb转录本中两个IRES的结构和功能的详细分析将使我们能够剖析翻译中的基本机制，如miRNA-IRES介导的翻译控制、核糖体过滤和IRES变构。这些结果也将被用于设计新的多顺反子载体，以允许在报告基因、药物选择标记或自杀基因的帮助下在靶细胞内表达感兴趣的基因，这些基因都来自同一转录本。病毒在癌症、基因治疗和药物传递等领域的新应用要求我们了解病毒与宿主机制之间的相互作用。转译控制的研究涉及生物技术、细胞生物学、医学和农业等多个领域。

项目成果

Poly(A) binding protein 1 enhances cap-independent translation initiation of neurovirulence factor from avian herpesvirus.

聚（a）结合蛋白1增强了来自禽疱疹病毒的神经电动性因子的帽非依赖性翻译起始。

• DOI: 10.1371/journal.pone.0114466
• 发表时间: 2014
• 期刊: PloS one
• 影响因子: 3.7
• 作者: Tahiri-Alaoui A;Zhao Y;Sadigh Y;Popplestone J;Kgosana L;Smith LP;Nair V
• 通讯作者: Nair V