Kinetic Analysis of Mouse Brain Proteome Alterations Following Chikungunya Virus Infection before and after Appearance of Clinical Symptoms

Recent outbreaks of Chikungunya virus (CHIKV) infection have been characterized by an increasing number of severe cases with atypical manifestations including neurological complications. In parallel, the risk map of CHIKV outbreaks has expanded because of improved vector competence. These features make CHIKV infection a major public health concern that requires a better understanding of the underlying physiopathological processes for the development of antiviral strategies to protect individuals from severe disease. To decipher the mechanisms of CHIKV infection in the nervous system, a kinetic analysis on the host proteome modifications in the brain of CHIKV-infected mice sampled before and after the onset of clinical symptoms was performed. The combination of 2D-DIGE and iTRAQ proteomic approaches, followed by mass spectrometry protein identification revealed 177 significantly differentially expressed proteins. This kinetic analysis revealed a dramatic down-regulation of proteins before the appearance of the clinical symptoms followed by the increased expression of most of these proteins in the acute symptomatic phase. Bioinformatic analyses of the protein datasets enabled the identification of the major biological processes that were altered during the time course of CHIKV infection, such as integrin signaling and cytoskeleton dynamics, endosome machinery and receptor recycling related to virus transport and synapse function, regulation of gene expression, and the ubiquitin-proteasome pathway. These results reveal the putative mechanisms associated with severe CHIKV infection-mediated neurological disease and highlight the potential markers or targets that can be used to develop diagnostic and/or antiviral tools.近期基孔肯雅病毒（Chikungunya virus, CHIKV）感染暴发的临床特征表现为重症病例数量持续攀升，且伴随包括神经系统并发症在内的非典型临床表现。与此同时，受媒介效能提升影响，基孔肯雅病毒暴发的风险范围不断扩大。上述特征使得基孔肯雅病毒感染成为重大公共卫生议题，亟需深入解析其潜在病理生理机制，以研发抗病毒策略，保护个体免受重症疾病侵害。 为解析基孔肯雅病毒神经系统感染的致病机制，本研究针对临床症状出现前后采样的基孔肯雅病毒感染小鼠脑组织内的宿主蛋白质组修饰情况开展了动态分析。本研究结合双向荧光差异凝胶电泳（two-dimensional difference gel electrophoresis, 2D-DIGE）与同位素相对标记与绝对定量技术（isobaric tags for relative and absolute quantitation, iTRAQ）两种蛋白质组学方法，并通过质谱进行蛋白质鉴定，最终筛选得到177个显著差异表达的蛋白质。 该动态分析结果显示，在临床症状出现前，宿主蛋白质组呈现显著的整体下调趋势；而在急性症状期，其中绝大多数蛋白质的表达水平均出现上调。对该蛋白质组数据集的生物信息学分析，鉴定出基孔肯雅病毒感染进程中发生显著改变的核心生物学过程，包括整合素信号通路与细胞骨架动力学、与病毒转运及突触功能相关的内体系统与受体循环、基因表达调控，以及泛素-蛋白酶体通路。 本研究结果揭示了与重症基孔肯雅病毒感染介导的神经系统疾病相关的潜在致病机制，并明确了可用于开发诊断工具或抗病毒疗法的潜在标志物与靶点。