Determining the Essential Role of the Major Phosphoprotein of Vaccinia Virus: Vp11

病毒DNA复制和形态发生所需的许多病毒蛋白质的活性受磷酸化调节。 这是牛痘病毒（VV）的情况，这是痘病毒科最了解的成员，越来越清楚。 痘病毒编码其病毒基因组转录和复制所需的绝大多数自身酶组分。 这种来自宿主的显著自主性使得VV成为最重要的真核生物化学和遗传模型系统之一。 VV的研究继续揭示了平行系统，如DNA复制，染色质凝聚，蛋白质的翻译后修饰，以及激酶和磷酸酶的动态方面。 特别是，VV编码两种蛋白激酶（F10和B1）和一种双特异性磷酸酶（H1）。 这些酶都是病毒生命周期中必不可少的。 然而，动态磷酸化在病毒生命周期中所起的确切分子作用在很大程度上仍然不确定。 这项研究的核心是牛痘病毒蛋白Vp 11，它是一种11 kDa的磷酸丝氨酸蛋白。 尽管Vp 11（由基因F17编码）是病毒粒子中最丰富的蛋白质（就分子数量而言），但其精确的体内功能仍然难以捉摸。 因此，本研究的主要目标是确定Vp 11的体内功能。 Vp 11在体外与DNA结合，并且长期以来被假定参与病毒基因组DNA的缩合。 通过调节核酸结合活性，蛋白质磷酸化可以是基因表达的正或负调节剂。 磷酸化也可能是蛋白质寡聚化所必需的。此外，蛋白质磷酸化也可能是蛋白质的适当亚细胞定位所需的，或者可能是蛋白质体内稳定性的关键。 该项目旨在探索这些场景。 该项目的最新结果指出MAP/ERK激酶信号通路在VV生命周期中的重要作用。 牛痘作为模型系统的已被证明的易处理性允许对宿主细胞的这种可能贡献进行解剖。 这些过程的各个方面都在这个项目中得到了解决，通过集中的Vp 11的分子表征以及在痘病毒的生命周期中的MAP/ERK信号通路的作用。 牛痘是用来消灭天花的活疫苗。 生物化学家对牛痘病毒的研究长期以来为人类细胞更复杂的运作提供了见解。 为了人类细胞的正确生长，需要许多形式的控制（也称为“调节”）。 如果没有这种调节，细胞生长就会变得异常。 最常见的调节类型是一个简单的化学过程，通过这个过程，磷酸根离子被放置在某些类型的蛋白质上。 这种调节被称为“磷酸化”。 这项由NSF资助的研究将通过使用更简单，因此更易于处理的病毒模型系统，为这一关键事件提供一个窗口。

The activities of many viral proteins required for viral DNA replication and morphogenesis are regulated by phosphorylation. That this is the case in vaccinia virus (VV), which is the best-understood member of the Poxviridae, is increasingly clear. Poxviruses encode the vast majority of their own enzymatic components required for the transcription and replication of their viral genome. This remarkable autonomy from the host makes VV one of the most premier eukaryotic biochemical and genetic model systems available. The study of VV continues to shed light on parallel systems such as DNA replication, chromatin condensation, post-translational modification of proteins, and dynamic aspects of kinases and phosphatases. In particular, VV encodes two protein kinases (F10 and B1) and a dual-specificity phosphatase (H1). These enzymes are all essential in the viral life cycle. However, the precise molecular role dynamic phosphorylation plays during the viral life cycle remains largely undefined. The vaccinia viral protein that is at the heart of this research, Vp11, is an 11-kDa-phosphoserine protein. Although Vp11 (encoded by the gene F17) is the most abundant protein (in terms of number of molecules) in the virion, its precise in vivo function(s) remains elusive. As such, the primary goal of this research is to determine the in vivo function(s) of Vp11. Vp11 binds to DNA in vitro, and has long been postulated to be involved in condensation of the viral genomic DNA. By modulating nucleic acid-binding activity, protein phosphorylation can be a positive or negative regulator of gene expression. Phosphorylation may also be necessary for the oligomerization of a protein. Furthermore, protein phosphorylation can also be required for the proper subcellular localization of a protein, or may be critical for the protein's stability in vivo. This project is designed to explore these scenarios. Recent results from this project point to an important role of the MAP/ERK kinase-signaling pathway during the VV life cycle. The proven tractability of vaccinia as a model system allows a dissection of this possible contribution of the host cell. Aspects of these processes are addressed within this project by a focused molecular characterization of Vp11 as well as the role of the MAP/ERK signaling pathway in the life cycle of poxviruses.The research described in this project involves an animal virus called vaccinia. Vaccinia was the live vaccine used to eradicate smallpox. The study of vaccinia virus by biochemists has long provided insights into the more complicated workings of human cells. For the correct growth of a human cell, many forms of control (also called "regulation") are required. Without this regulation, cell growth becomes abnormal. The most common type of regulation is a simple chemical process by which a phosphate ion is placed on certain types of protein. Regulation of this sort is termed "phosphorylation". This NSF-funded research will provide a window into this crucial event by using a much simpler, and therefore more tractable, viral model system.