Defining viral-host interactions between arthritogenic alphaviruses and MARCO

项目总结 虫媒病毒维持在人-蚊子-人传播周期中，负责为周期性的 在全球范围内暴发，是日益严重的公共卫生威胁。虫媒病毒传播周期的一个关键特征， 病毒血症的大小和持续时间是它们地理传播和致病的主要决定因素。 在脊椎动物的宿主中。然而，很少有研究研究病毒血症的分子决定因素。近期 莫里森实验室发表的研究表明，小鼠肝脏上的清道夫受体Marco 巨噬细胞清除基孔肯雅(CHIKV)颗粒和包括Ross在内的其他致关节炎甲型病毒 由于对赖氨酸(K)的识别，RRV和ONNV病毒从小鼠循环中分离出来 CHIKV和ONNV E2糖蛋白第200位残基，RRV E2糖蛋白第251位残基。我的初选 研究进一步表明，当谷氨酸发生突变时，CHIKV的清除也会被取消 (E)E_2的208和E_1糖蛋白的K_(61)，并对重要的生化特征进行质谱分析 病毒清除提示E1K61是甲基化的。CHIKV E2基因208位的进一步分析 糖蛋白揭示了这个位置的负电荷对CHIKV从循环中清除的重要性。 作为一种模式识别受体，Marco识别修饰的自我和非自我分子以及多态 在人类中，马可可使携带者易患结核病等传染病。因为清道夫 MARCO的受体富含半胱氨酸(SRCR)结构域是内源性配体的结合部位，如修饰的 低密度脂蛋白，我假设Marco的SRCR结构域稳定和非共价相互作用 与CHIKV的E1和E2糖蛋白之间暴露的界面，允许去除 减少循环中的病毒颗粒，减少病毒血症的程度和持续时间。在目标1中， 我将定义CHIKV的残留物和生化特征，这些特征对于从 通过操纵病毒颗粒的表面特征，评估特定突变如何影响病毒的循环 传播，并确定在E1和E2糖蛋白的特定位置的翻译后修饰。在……里面 目的2，我将阐明MARCO上负责与致关节炎甲型病毒结合的细胞和 生化方法。此外，我将确定病毒颗粒与人类相互作用的程度 Marco，以及已知的Marco基因多态性是否会影响病毒-Marco相互作用、病毒血症或临床 结果。综上所述，通过定义Marco和CHIKV之间相互作用的分子机制， 这一建议可以提供对影响甲型病毒致病因素的洞察，阐明两者之间的关系 Marco基因多态和病毒血症之间的关系，并识别具有增加的 对严重的甲型病毒感染和暴发的易感性。

PROJECT SUMMARY Arboviruses maintained in a human-mosquito-human transmission cycle are responsible for fueling periodic outbreaks worldwide and are an increasing public health threat. A critical feature of arbovirus transmission cycles, and a major determinant of their geographic spread and pathogenesis, is the magnitude and duration of viremia in vertebrate hosts. However, few studies have investigated the molecular determinants of viremia. Recent studies published by the Morrison laboratory demonstrated that the murine scavenger receptor MARCO on liver macrophages removes chikungunya (CHIKV) particles and other arthritogenic alphaviruses, including Ross River (RRV) and o’nyong ‘nyong (ONNV) viruses, from murine circulation due to recognition of the lysine (K) residue at position 200 of CHIKV and ONNV E2 glycoprotein and 251 of RRV E2 glycoprotein. My preliminary studies further revealed that CHIKV clearance is also abrogated when mutations were introduced at glutamate (E)208 of E2 and K61 of E1 glycoproteins, and mass spectrometry analysis of the biochemical features important for viral clearance suggested that E1 K61 is methylated. Further analysis of position 208 of CHIKV E2 glycoprotein revealed the importance of a negative charge at this position for CHIKV removal from circulation. As a pattern recognition receptor, MARCO recognizes modified self and non-self molecules, and polymorphisms in human MARCO can predispose carriers to infectious diseases such as tuberculosis. Because the scavenger receptor cysteine-rich (SRCR) domain of MARCO is a binding site for endogenous ligands, such as modified low-density lipoprotein, I hypothesize that the SRCR domain of MARCO stably and noncovalently interacts with an exposed interface between the E1 and E2 glycoproteins of CHIKV, allowing for the removal of viral particles from circulation and a reduction in both the magnitude and duration of viremia. In Aim 1, I will define the residues and biochemical features of CHIKV important for MARCO-dependent clearance from circulation by manipulating surface features of virus particles, assessing how specific mutations impact viral dissemination, and identifying post-translational modifications at specific sites in the E1 and E2 glycoproteins. In Aim 2, I will elucidate the sites on MARCO responsible for binding arthritogenic alphaviruses with cell-based and biochemical approaches. In addition, I will determine the extent to which virus particles interact with human MARCO, and whether known polymorphisms in MARCO affect virus-MARCO interactions, viremia, or clinical outcomes. Taken together, by defining the molecular mechanism of interaction between MARCO and CHIKV, this proposal could provide insights into factors that influence alphaviral pathogenesis, elucidate the relationship between MARCO polymorphisms and viremia, and identify individuals or populations with an increased susceptibility to severe alphaviral infections and outbreaks, respectively.