人巨细胞病毒（HCMV）是围产期感染的常见病原，近期研究表明其致病性和免疫逃避可能与自噬相关，但确切机制尚未阐明。Beclin 1是自噬过程中的关键蛋白，其作用机制和相关信号通路有待深入研究。本项目在前期工作基础上，拟建立HCMV潜伏感染CD14+细胞模型，首先检测模型中的自噬标志物，探明HCMV潜伏感染的自噬水平变化特点；进而通过改变潜伏感染模型中的自噬水平，监测HCMV细胞易感性或感染状态的变化，明确自噬水平对HCMV潜伏感染的影响；最后调控IE2水平探讨其对Akt和Beclin 1表达及磷酸化的作用，并采用免疫沉淀和质谱分析鉴定IE2-Akt-Beclin 1通路上、下游结合Beclin 1的关键蛋白，揭示IE2-Akt-Beclin 1通路在HCMV潜伏感染自噬调控中的分子机制。本项目将为阐明HCMV潜伏感染的发病机制提供新思路，并为HCMV防治药物和疫苗的研发提供新靶标。

Human cytomegalovirus (HCMV) is the most frequent cause of congenital infection. Transplacental transmission of HCMV during pregnancy or neonatal infection of premature newborns can lead to neurological damage. Both primary and latency-activation infection of HCMV during gestation pose a risk of intrauterine transmission. Given the extremely high seropositive rate in pregnant women, latent infection seems to play an important role in congenital HCMV infection. Autophagy regulation and the molecular mechanisms in HCMV latency remain poorly understood. Most of the respective viral proteins affect autophagy via binding to Beclin 1, an autophagy-related protein that constitutes part of the phosphatidylinositol-3 kinase complexes and plays a crucial role in autophagy initiation. HCMV immediate-early protein IE2 is essential for HCMV replication due to its ability to transactivate critical viral early promoters. The expression level of IE2 is accordingly regarded as the most important hallmark to discriminate productive and latent infection of HCMV. Here, we propose a new IE2-Akt-Beclin 1 signaling pathway in the autophagy regulation of HCMV latency based on the latest research progress. In this study, a model of HCMV latency by infecting CD14+ monocyte will be constructed for exploring the regulatory mechanisms of autophagy and IE2-Akt-Beclin 1 pathway. The mRNA and protein levels of several autophagy-related markers such as Beclin 1, LC3, and p62 in this model will be firstly detected to evaluate the characteristics of autophagy regulation in HCMV latency. Furthermore, the autophagy level in this model will be modulated by chemical agents or small interfering RNA (siRNA) and microRNAs targeting to Beclin 1. The susceptibility of CD14+ monocyte to HCMV or the viral infection status will be compared between before and after the modulations to determine the effect of autophagy on HCMV latent infection. For the purpose of understanding the role of IE2-Akt-Beclin 1 pathway in autophagy regulation, the expressions and phosphorylations of Akt and Beclin 1 will be detected before and after changing the level of IE2 in HCMV productive and latent infection cell models. In addition, immunoprecipitation and mass spectrometric analysis will be carried out to identify the key molecules interacting with Beclin 1 in the upstream and downstream of the IE2-Akt-Beclin 1 pathway. The autophagy regulation in HCMV latency and the role of relevant IE2-Akt-Beclin 1 pathway revealed in this study will be helpful to further explore the pathogenesis of HCMV latency and to facilitate the development of novel vaccines and drugs against this virus.