登革病毒NS3蛋白与丙酮酸羧化酶相互作用调控糖酵解在病毒复制中的作用及分子机制

Dengue virus (DENV) is one of the major arbovirus that seriously threat global human health. However, there is no specific antiviral drug available for the treatment of DENV infection. DENV NS3 is a multifunctional protein which is a main target for antiviral drugs. Our research group discovered that pyruvate carboxylase (PC) interacted with NS3 protein through affinity purifications and Co-IP analyses. Our research also showed that NS3 interacted with the carboxyltransferase domain of PC, and over-expression of PC promoted the DENV-2 replication. PC is the rate-limiting enzyme in gluconeogenesis and catalyses the ATP-dependent carboxylation of pyruvate to oxaloacetate, which plays a crucial role in cell metabolism. In recent years, studies revealed that some viruses activate glycolysis to supplie the intermediates necessary for their rapid viral genome replication and virion assembly by different mechanisms. So, we proposed that the interaction between NS3 and PC influences the function of the CT domain and then induces the glycolytic activity to promote the DENV replication. Thus, this project will determine the effects of NS3 on PC activity, and then identify the key amino acids involved in the interaction between NS3 protein and CT domain of PC. Finally, this project will investigate the mechanism of the interaction between NS3 and PC in glycolysis during DENV-2 replication and may provide novel therapeutic target against DENV infection.

登革病毒（DENV）是世界范围内广泛流行、严重危害人类健康的虫媒病毒，尚无有效治疗药物。DENV NS3蛋白具有多种功能，是抗病毒药物的主要靶点之一。本课题组通过亲和层析筛选和Co-IP等发现丙酮酸羧化酶（PC）与NS3相互作用，主要作用区域为PC羧基转移酶（CT）区，且过表达PC促进DENV复制。PC是糖异生的限速酶，可催化丙酮酸转变成草酰乙酸，在细胞代谢中发挥重要作用。新近研究表明，多种病毒均可通过激活糖酵解途径为其复制或子代病毒产生提供所需物质和能量，其机制各不相同。据此，我们推测NS3与PC的相互作用影响了PC CT区功能的发挥，诱导糖酵解途径代谢升高，促进DENV复制。为此，本项目拟明确NS3蛋白对PC活性的影响、NS3蛋白和PC CT区相互作用关键氨基酸及NS3与PC相互作用调控糖酵解的分子机制，旨在深入探索DENV感染细胞后调控糖酵解的具体机制，为抗DENV药物发现提供新的靶点。

登革病毒（DENV）是世界范围内广泛流行、严重危害人类健康的虫媒病毒，尚无有效治疗药物。DENV NS3蛋白具有多种功能，是抗病毒药物的主要靶点。本课题组通过亲和层析筛选和质谱鉴定发现丙酮酸羧化酶（PC）与NS3可能相互作用。PC是糖异生的限速酶，在细胞代谢中发挥重要作用。新近研究表明，多种病毒均可通过激活糖酵解途径为其复制或子代病毒产生提供所需物质和能量，但其机制各不相同。为此，本项目通过检测PC 过表达和低表达对DENV-2复制的影响，DENV-2感染或NS3过表达对PC以及糖酵解通路的影响以及确定NS3和PC相互作用的结构区，深入研究NS3与PC相互作用调控糖酵解的分子机制。结果表明，在DENV-2感染早期，PC可促进病毒复制，并且NS3的helicase区可以和PC的CT区相互作用调控糖酵解通路活性，从而有可能进一步为DENV-2复制提供ATP和原料。通过深入探索DENV感染细胞后调控糖酵解的具体机制，可为抗DENV药物发现提供新的靶点。