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.

Marek病（MD）是一种广泛的淋巴肿瘤疾病，其肿瘤是由Marek病毒（MDV）感染引起的。 MD是全球主要的家禽健康和福利问题之一，尽管每年使用超过200亿剂MD疫苗，但估计损失量高达2000万美元。事实证明，MDV是理解某些肿瘤发生原理的宝贵模型。它提供了良好定义的一般肿瘤发生的小动物模型，尤其是病毒诱导的淋巴作用。在天然病毒宿主系统中，可再现的疾病诱导和进展动力学使MDV独特，从而可以在其他非天然疱疹病毒诱导的肿瘤发生的非天然模型中进行研究。病毒感染细胞的研究一直是转化控制机制的重要信息来源。作为强制性细胞内寄生虫，病毒取决于细胞的复制。由于病毒缺乏翻译设备，因此没有比翻译系统中更清楚地看到这种依赖性的。病毒已经发展出为其mRNA获得翻译优势的方法。这在病毒感染的早期阶段尤其至关重要，在病毒感染的早期阶段，即立即产生了早期的转录本，需要翻译。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期间使用的策略之一，以及何时可以抑制帽依赖性翻译。我们已经发现，从MDV的立即获得的转录本具有两个IRES。第一个IRE包含在5'领导者序列中，并控制着编码PP14裂解蛋白的下游开放阅读框的翻译。 5'领导者的IRE似乎是由18S rRNA内的序列和40S核糖体亚基内的核糖体蛋白质的序列调节的。第二个IRES包含在间间区域内（ICR）中，并控制第二种蛋白质的翻译，称为Rlorf9。 ICR IRES活性似乎由一组称为微RNA的病毒小型非编码RNA控制。已知该转录物与致癌性和MDV转化细胞潜伏期的维持有关。该转录本在病毒衰减期间经历特定序列扩增的基因座。该序列放大是唯一与病毒衰减相关的唯一经过验证的标记。这项研究建议将带来什么好处，尤其是在MDV肿瘤发生的领域，通常会带来转化控制？初步工作使我们能够在翻译控制中提出新颖的概念，并直接影响对MDV肿瘤发生的理解。高度致癌的RB-1B菌株中传染性BAC克隆的可用性将使我们能够使用重组介导的基因工程来研究细胞培养和鸡肉中1.8-KB转录本的生物学功能。这种高度可重复的疾病模型将用于自然宿主中，以获取有关疱疹病毒诱导的淋巴疗法的机理，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在翻译中剖析基本机制，例如miRNA-IRES介导的翻译控制，核糖体滤波和IRES变构。在设计新型的多发性载体时，还将利用该结果，以借助报告基因，药物选择标记或自杀基因的范围内表达目标细胞内的感兴趣基因，这些基因均根据同一转录本表达。病毒在癌症，基因治疗和分娩中的新兴应用使人对病毒与宿主机制之间的相互作用的了解。翻译控制的研究影响了包括生物技术，细胞生物学，医学和农业在内的几个领域。

项目成果

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

• 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