ABCF1识别HBV_cccDNA诱导IFN-Lambda应答抑制HBV复制的作用及分子机制

The covalently closed circular DNA (cccDNA) of hepatitis B virus (HBV) is viral persistence reservoir and key obstacle for a cure of chronic hepatitis B. Pattern recognition receptors recognize viral nucleic acids to induce the production of interferon (IFN), which is an important prerequisite to stimulate an effective innate immune response for HBV clearance. Up to now, how the host DNA sensors recognize HBV cccDNA to activate the innate immune response remains unclear. Using a minicircle DNA model with high similarity to cccDNA to screen host proteins that bind to cccDNA, we found a potential cccDNA binding protein ABCF1, a vital innate immune regulatory factor. Preliminary studies have shown that ABCF1 can bind to HBV cccDNA in vitro, its over-expression not only significantly inhibits HBV replication, but also significantly enhances the IFN-Lambda response induced by HBV infection, which strongly suggests that ABCF1 is a pattern recognition receptor that recognizes cccDNA. We will further demonstrate the inhibitory role of ABCF1 in HBV replication as DNA sensor，and illuminate the molecular mechanism of ABCF1 recognizing cccDNA to inhibit HBV replication through activating TBK1-IRF3 pathway to induce IFN-Lambda response depending STING or other critical molecules. This study will provide novel knowledge of understanding innate immune recognition of HBV infection and the development of new antiviral therapeutic targets.

HBV cccDNA是病毒持续复制的关键，模式识别受体识别病毒核酸是机体激发有效固有免疫应答清除HBV的重要前提，宿主DNA感受器如何识别cccDNA并激活固有免疫应答的分子机制尚不清楚。本项目前期利用高度模拟cccDNA的微环DNA模型，筛选cccDNA互作宿主蛋白时发现，固有免疫调节因子ABCF1可与cccDNA相互作用。初步研究显示，ABCF1可结合cccDNA，其过表达不仅显著抑制HBV复制，还显著增强HBV感染诱导的IFN-L应答，强烈提示ABCF1是识别cccDNA的模式识别受体。本课题将进一步明确ABCF1作为cccDNA感受器，抑制HBV复制的作用，并系统解析ABCF1识别cccDNA后通过STING依赖性或非依赖方式激活TBK1-IRF3信号通路，诱导IFN-L反应发挥抗病毒作用的分子机制,以期为揭示固有免疫识别HBV感染的新机制及抗病毒治疗新靶点提供理论依据。

乙肝病毒（HBV）共价闭合环状DNA（cccDNA）结合多种宿主蛋白形成微染色体驱动HBV复制，是慢乙肝难以清除的根本原因。模式识别受体识别病毒核酸诱导IFN应答是乙肝病毒清除的重要方式，然而，宿主DNA识别受体如何识别cccDNA并激活干扰素反应目前仍不清楚。基于创新性建立的cccDNA-宿主互作筛选模型，本项目发现重要的固有免疫调节因子ATP结合盒转运蛋白F1(ABCF1)是新的cccDNA结合蛋白和乙肝病毒复制的宿主限制因子。机制上，一方面，ABCF1识别cccDNA诱导IFN反应发挥部分抑制HBV复制的作用。更为重要的是，ABCF1通过其N末端无序低复杂结构域与cccDNA相互作用，发生液液相分离，进而抑制RNA转录酶Pol II的招募和cccDNA转录活性。总之，本项目揭示了ABCF1识别cccDNA后以相分离依赖的方式转录抑制HBV复制，为乙肝病毒cccDNA活性调控和慢乙肝治疗提供新策略。