Molecular mechanisms linking viral replication and neuropathogenesis

项目摘要/摘要 感染发育中的大脑的病毒，包括寨卡病毒、巨细胞病毒和风疹病毒会引起 严重的出生缺陷。小头畸形就是这样一种出生缺陷，它的头部和大脑尺寸严重缩小， 而且常常伴随着智力残疾。这种由病毒引起的神经系统疾病，或称病毒 神经发病机制，可由多种机制引起。最近发现的一种ZIKV非结构方式 蛋白4A(NS4A)通过破坏人类ANKLE2蛋白而导致小头畸形。有趣的是，个人 由于编码ANKLE2的基因发生突变，导致小头畸形。ANKLE2是从蠕虫到 在大脑发育过程中，对协调细胞分裂是必不可少的。ANKLE2派生出此函数 在细胞分裂和发育中通过调节蛋白质的相互作用。NS4A与ANKLE2在物理上相互作用 在果蝇大脑发育模型中以依赖ANKLE2的方式干扰大脑发育。确认2 还会促进ZIKV复制。综上所述，这些研究表明，在寻找宿主蛋白质的过程中 对于复制，ZIKV失调了一条重要的发育途径。因此，NS4A-ANKLE2蛋白 相互作用是研究病毒神经致病机制及其如何与病毒联系的重要模型 分子水平上的复制和遗传性疾病。 这项工作的长期目标是破译病毒-宿主蛋白相互作用如何影响病毒复制和 发病机制，因为这些发现将推动治疗靶点的确定和药物开发。这个 本研究的目的是分析ZIKV NS4A蛋白相互作用的机制。 人类ANKLE2促进ZIKV复制，抑制大脑发育。为了实现这一目标，我们 将检验核心假设，即ANKLE2通过与NS4A和NS4A的相互作用促进病毒复制 通过招募参与ZIKV复制的其他宿主因素到复制站点，这扰乱了生理 ANKLE2蛋白相互作用是大脑发育所必需的。以下具体目标将检验这一假设： 目的1：分析NS4A-ANKLE2蛋白相互作用在ZIKV复制和致病中的作用。 目的2：揭示ANKLE2在ZIKV复制和致病中的分子功能。 当这项工作完成后，这项工作将描绘出单个病毒-宿主蛋白相互作用如何改变发育 促进病毒复制并在分子水平上造成神经疾病的途径。这将揭示 详细的生化洞察病毒与宿主蛋白的相互作用与氨基酸水平的解析，新的宿主因子 在ZIKV复制中发挥作用的蛋白质，以及以前未知的大脑发育关键蛋白 其他遗传性发育障碍。从长远来看，这里建立的方法可以用来 揭示其他具有病毒和遗传病因的疾病背后的分子机制。

PROJECT SUMMARY/ABSTRACT Viruses that infect the developing brain, including Zika virus (ZIKV), cytomegalovirus, and rubella virus cause major birth defects. Microcephaly is one such birth defect, in which head and brain size are severely reduced, and is often accompanied by intellectual disability. This virally-inflicted neurological disease, or viral neuropathogenesis, can be caused by multiple mechanisms. One recently identified way ZIKV non-structural protein 4A (NS4A) causes microcephaly is by disrupting the human ANKLE2 protein. Interestingly, individuals with mutations in the gene encoding ANKLE2 suffer from microcephaly. ANKLE2 is conserved from worms to humans, and is essential for coordinating cell division during brain development. ANKLE2 derives this function in cell division and development by mediating protein interactions. NS4A physically interacts with ANKLE2 and disrupts brain development in an ANKLE2-dependent manner in a fruit fly model of brain development. ANKLE2 also promotes ZIKV replication. Taken together, these studies show that in the process of coopting a host protein for replication, ZIKV dysregulates an important developmental pathway. Thus, the NS4A-ANKLE2 protein interaction represents an important model to study viral neuropathogenesis and how it is connected to viral replication and hereditary disorders at the molecular level. The long-term goal of this work is to decipher how virus-host protein interactions impact virus replication and pathogenesis, as these discoveries will fuel therapeutic target identification and drug development. The objective of this proposal is to dissect the mechanisms by which the protein interaction between ZIKV NS4A and human ANKLE2 promote ZIKV replication and inhibit brain development. To accomplish this objective, we will test the central hypothesis that ANKLE2 promotes viral replication through its interaction with NS4A and by recruiting other host factors involved in ZIKV replication to sites of replication, and this disrupts physiological ANKLE2 protein interactions required for brain development. The following specific aims will test this hypothesis: Aim 1: Dissect the impact of the NS4A-ANKLE2 protein interaction in ZIKV replication and pathogenesis. Aim 2: Unravel the molecular function of ANKLE2 in ZIKV replication and pathogenesis. When completed, this work will delineate how a single virus-host protein interaction rewires a developmental pathway to facilitate virus replication and inflict neurological disease at the molecular level. This will reveal detailed biochemical insight into a virus-host protein interaction with amino acid-level resolution, new host factors that play a role in ZIKV replication, and previously unknown proteins key to brain development and disrupted in other hereditary developmental disorders. In the long term, the methods established here could be employed to uncover the molecular mechanisms behind other diseases with viral and hereditary etiologies.