Host proteins in Vaccinia viral membrane biogenesis

描述（由申请人提供）：人们对牛痘病毒的复制非常感兴趣，因为它代表了理解痘病毒的模型系统，包括天花的病原体天花病毒。病毒复制的一个关键阶段涉及获得膜，因为这一事件将非感染性组装病毒核心转化为感染性病毒粒子。积累的证据表明，病毒从宿主的细胞器室获得两套膜。内层膜来自内质网高尔基中间隔室（ERGIC），形成细胞内成熟病毒（IMV），而外层膜随后从反式高尔基网络获得，形成细胞内包膜病毒。参与膜包裹的病毒蛋白已经开始被确定，但是宿主蛋白也被预测在这一过程中发挥关键作用，因为膜来自宿主细胞器，仍然未知。在初步研究中，我们发现牛痘病毒特异性地将宿主蛋白（COPI外壳复合体的一个亚基）浓缩在其内层膜上。因此，我们建议阐明COPI外壳复合体如何在病毒生物发生中发挥作用。首先，我们将确定COPI首次在病毒膜上被检测到时病毒生物发生的阶段。其次，我们将确定当COPI功能被废除时，哪个病毒阶段可能会被阻止。第三，我们将确定病毒是否篡夺任何已知的宿主调节蛋白，将COPI招募到病毒膜上。第四，我们将测试当前已知的调节病毒形态发生的病毒蛋白是否会影响COPI在病毒膜上的募集。这些顺序方法还提出了一种系统的方法，可以识别可能影响IMV形成的潜在其他宿主蛋白质，因为这是病毒具有传染性的第一个阶段。因此，我们将筛选目前已知的在ERGIC区起作用的运输调节因子，以测试是否有类似的在病毒膜上富集。随后，候选宿主蛋白的功能将被取消，以确定病毒的生物发生是否受到影响。这些努力不仅可以更好地了解病毒在复制过程中如何与宿主相互作用，而且还可以为未来合理设计针对痘病毒的治疗干预提供新的生化靶点。

DESCRIPTION (provided by applicant): Much interest exists in understanding the replication of vaccinia virus, as it represents the model system to understand poxviruses that include variola virus, the causative agent of smallpox. A key phase of viral replication involves the acquisition of membranes, as this event transforms non-infectious assembling viral cores into infectious virions. Accumulated evidence suggests that the virus acquires two set of membranes from organellar compartments of the host. The inner set of membranes is derived from the Endoplasmic Reticulum Golgi Intermediate Compartment (ERGIC) to form an intracellular mature virus (IMV), while the outer set of membranes is subsequently acquired from the trans-Golgi network to form an intracellular envelope virus. Viral proteins that participate in membrane wrapping are beginning to be identified, but host proteins that would also be predicted to play a key role during this process, as the membranes are derived from host organelles, have remained unknown. In preliminary studies, we have found that vaccinia virus specifically concentrates a host protein, a subunit of the COPI coat complex, on its inner set of membranes. Thus, we propose to elucidate how the COPI coat complex might play a role in viral biogenesis. First, we will determine the stage of viral biogenesis when COPI is first detected on viral membranes. Second, we will determine which viral stage might be arrested when COPI function is abrogated. Third, we will determine whether the virus usurps any of the known host regulatory proteins to recruit COPI onto viral membranes. Fourth, we will test whether any of the current known viral proteins that regulate viral morphogenesis affects COPI recruitment onto viral membranes. These sequential approaches also suggest a systematic way of identifying potential other host proteins that would be predicted to affect IMV formation, as it is the first stage that the virus becomes infectious. Thus, we will screen the currently known transport regulators that function at the ERGIC compartment to test whether any is similarly enriched on viral membranes. Subsequently, the function of candidate host proteins will be abrogated to determine whether viral biogenesis is affected. These efforts will likely not only provide a better understanding of how the virus interacts with its host during replication, but also suggest novel biochemical targets in the future rational design of therapeutic intervention against poxviruses.