New Opportunities - Mechanisms of Early Vaccinia Viral Morphogensis

摘要 人们已经对痘病毒家族成员作为疫苗和表达载体的应用进行了研究，并且最近由于它们作为潜在的生物武器的用途而进行了深入研究。牛痘病毒作为原型成员，通过多种形式进化获得感染性，其中早期膜形态发生起着关键作用。尽管病毒通常会侵占宿主因子，但目前尚不清楚众所周知在细胞区室膜形态发生中起作用的关键宿主转运因子是否参与早期牛痘膜形态发生。我们现在发现了外壳蛋白的作用，外壳蛋白是一种宿主蛋白​​复合物，已知可形成外壳蛋白 I (COP!) 运输囊泡。然而，对衣壳异构体如何参与早期病毒形态发生的深入研究表明，该病毒绕过了形成宿主 COP1 囊泡的关键调节机制，而是与两个 蛋白质、病毒 K7 和宿主 KDEL 受体 (KDELR) 通过关键的二赖氨酸残基可能很重要。因此，为了进一步了解衣壳蛋白及其两种相互作用的蛋白如何促进病毒复制，我们提出了两个主要目标。在由 Victor Hsu 领导的一项由 NERCE 提议资助的目标中，将检查外衣异构体的扰动对于潜在积累特定病毒形式的影响。此外，还将检查 KDELR 在病毒形式上的分布以及是否 其配体在病毒复制中发挥作用。作为由 Stuart lsaacs 领导并获得 MARCE 资助的另一个目标，将检查病毒 K7 蛋白对其删除的影响，以及当表达不能与外衣异构体相互作用的突变形式时的影响。 这项合作以多种方式响应两个区域卓越中心在“新机会”倡议下的申请请求。首先，我们将深入了解 A 类病原体，为此我们不仅将阐明宿主蛋白在病毒复制过程中发挥作用的新机制，还将确定未来设计的潜在关键目标 针对最丰富的传染性病毒形式的干预措施。其次，该提案的资助将使主要研究人员之一（Victor Hsu）能够与痘病毒专家（Stuart Isaacs）合作，从而将囊泡运输机制方面的外部专业知识应用于痘病毒生物学的关键问题。由于病毒通常会征用宿主机制来与宿主相互作用，而不是发明 完全新颖的合作，互补的专业知识可能会加强对牛痘病毒如何与其宿主相互作用的阐明。第三，该提案的潜在资助可能会为未来最终更全面的应用奠定基础，例如通过 NIH R01 机制。

Abstract Members of the poxvirus family have been investigated for their applications as vaccines and expression vectors, and more recently intently studied because of their use as potential biological weapons. Vaccinia virus, the prototypic member, evolves through multiple forms in acquiring infectivity, for which early membrane morphogenesis plays a key role. Even though viruses generally usurp host factors for its use, it remains unclear whether key host transport factors that are well known to act in membrane morphogenesis of cellular compartments participate in early vaccinia membrane morphogenesis. We have now discovered a role for coatomer, a host protein complex known to shape Coat Protein I (COP!) transport vesicles. However, insights into how coatomer participates in early viral morphogenesis reveals that the virus bypasses key regulatory mechanisms that form host COP1 vesicles, but instead its interaction with two proteins, the viral K7 and the host KDEL receptor (KDELR) through critical di-lysine residues is likely important. Thus, to gain further insight into how coatomer and its two interacting proteins contribute to viral replication, we propose two major aims. In one aim headed by Victor Hsu with proposed funding from NERCE, perturbation of coatomer will be examined for its effect in potentially accumulating particular viral forms. Moreover, the KDELR will be examined with respect to its distribution on viral forms and also whether its ligands have a role in viral replication. As another aim headed by Stuart lsaacs with proposed funding from MARCE, the viral K7 protein will be examine for effects upon its deletion and also when a mutant form that cannot interact with coatomer is expressed. This collaborative effort is responsive in multiple ways to the request for application under the "New Opportunities" initiative by the two respective Regional Centers of Excellence. First, we will be gaining insights into a Category A pathogen, for which we will not only elucidate novel mechanisms by which a host protein functions during viral replication, but also identify potential key target(s) for the future design of intervention against the most abundant infectious form of the virus. Second, funding of this proposal will allow one of the main investigators (Victor Hsu) to collaborate with a poxvirus expert (Stuart Isaacs), and thus, applying outside expertise in mechanisms of vesicular transport to critical issues in poxvirus biology. As viruses generally commandeer host mechanisms for their interaction with the host rather than inventing completely novel ones, the complementary expertise of the collaboration will likely enhance the elucidation of how vaccinia virus interacts with its host. Third, potential funding of this proposal will likely set the stage for an eventual more comprehensive application in the future, such as through the NIH R01 mechanism.