Molecular virology of insecr baculoviruses

不安全杆状病毒的分子病毒学

• 批准号: 8395-2009
• 负责人: Krell, Peter
• 金额: $ 6.05万
• 依托单位: University of Guelph
• 依托单位国家: 加拿大
• 项目类别: Discovery Grants Program - Individual
• 财政年份: 2010
• 资助国家: 加拿大
• 起止时间: 2010-01-01 至 2011-12-31
• 项目状态: 已结题
• 来源: https://www.nserc-crsng.gc.ca/ase-oro/Details-Detailles_eng.asp?id=450286
• 关键词: Molecular; virology; insecr; baculoviruses

While much is known (and still unknown) about the interactions between mammalian viruses and their host cells that between insect viruses and insect cells is poorly understood. Baculoviruses are unique to insects and as such their study should reveal novel aspects of their replication strategies. This proposal involves two baculoviruses. CfMNPV infects the spruce budworm (a major defoliator in Canadian forests) and AcMNPV infects many lepidopteran insects, particularly pests of agricultural crops. With about 150 baculovirus genes, there is a need to regulate when and at what levels those genes are expressed. It would be wasteful if all genes were expressed simultaneously and at the same level. Some virus products are needed at only low amounts and early in infection (e.g. enzymes for DNA replication) whereas others are needed in higher amounts and later in infection (e.g. virion proteins). Similarly viral DNA replication, which occurs before viral assembly, requires prior production of viral DNA polymerase. This study focuses on genome transcriptional regulation and viral DNA replication. In particular we will characterize two baculovirus proteins, ME53 and Cf103 both of which may be involved in regulation of gene expression. We will determine when these proteins and their transcripts are made, where in the cells these proteins are localized, what viral and cellular proteins or DNA sequences they interact with (and how) and evaluate how they influence the expression of other baculovirus genes (we already know that Cf103 delays expression of late genes). We noted that CfMNPV generates several antisense, non-coding RNAs. While this may be a natural consequence of transcription of upstream sense mRNAs we feel some of the antisense transcripts have a regulatory role in the virus replication cycle, perhaps using RNA silencing, and we wish to address that hypothesis. In addition, since viral DNA replication is such a central feature of the virus replication cycle we wish to determine the mechanism of DNA replication. To that end we will try to identify where on the DNA genome replication actually starts (an ori of DNA replication) and what viral and cellular components exist at the origin of replication complex (ORC).

虽然我们对哺乳动物病毒和它们的宿主细胞之间的相互作用了解很多（仍然是未知的），但对昆虫病毒和昆虫细胞之间的相互作用却知之甚少。杆状病毒是昆虫所特有的，因此对它们的研究应该揭示它们复制策略的新方面。该提议涉及两种杆状病毒。CfMNPV感染云杉芽虫（加拿大森林中的主要食叶虫），AcMNPV感染许多鳞翅目昆虫，特别是农作物的害虫。大约有150个杆状病毒基因，需要调节这些基因何时表达以及表达水平。如果所有的基因都同时在同一水平上表达，那将是一种浪费。一些病毒产物在感染早期仅需要少量（例如用于DNA复制的酶），而另一些病毒产物在感染后期需要大量（例如病毒体蛋白）。类似地，在病毒组装之前发生的病毒DNA复制需要预先产生病毒DNA聚合酶。本研究的重点是基因组转录调控和病毒DNA复制。特别是，我们将表征两种杆状病毒蛋白，ME53和Cf103，这两种蛋白都可能参与基因表达的调控。我们将确定这些蛋白质及其转录物是何时产生的，这些蛋白质在细胞中的位置，它们与哪些病毒和细胞蛋白质或DNA序列相互作用（以及如何相互作用），并评估它们如何影响其他杆状病毒基因的表达（我们已经知道Cf103延迟晚期基因的表达）。我们注意到CfMNPV产生几种反义非编码RNA。虽然这可能是上游正义mRNA转录的自然结果，但我们认为一些反义转录物在病毒复制周期中具有调节作用，可能使用RNA沉默，我们希望解决这一假设。此外，由于病毒DNA复制是病毒复制周期的核心特征，我们希望确定DNA复制的机制。为此，我们将尝试确定DNA基因组复制实际上从哪里开始（DNA复制的起点），以及复制复合物起点（ORC）存在哪些病毒和细胞成分。