Targeting the host NDP kinase to abrogate viral dissemination

项目摘要 目前迫切需要开发安全有效的治疗方法，以对抗严重的 人类病原体，如卡波西氏肉瘤疱疹病毒(KSHV)，它会导致显著的发病率和 免疫受损个体的死亡率仍然是一个临床挑战。目前，没有FDA批准的 有治疗方法或疫苗可用于KSHV感染。鉴于KSHV和其他人类疱疹病毒 劫持宿主蛋白和途径以完成其复制周期并在细胞之间传播的策略 针对这些途径和机制将提供广谱的基因复盖和很高的障碍 与抗药性有关。包括KSHV在内的疱疹病毒早就知道利用COPII介导的 它们成熟的分泌途径。然而，管理这一进程的机制仍然较少。 明白了。这个项目将填补这一空白，利用我们最近的发现，宿主核苷 二磷酸激酶NM23-H2是COPII囊泡运输的重要调节因子，被KSHV开发用于 它们的病毒粒子的形态发生和出口。我们发现KSHV的病毒Bcl2(Ks-Bcl2)直接与KSHV的Bcl2相互作用， 在KSHV裂解期稳定并激活NM23-H2。NM23-H2丢失或Ks-Bcl2突变 取消NM23-H2相互作用严重损害了病毒粒子的产生。因此，我们假设KSHV激活 宿主蛋白NM23-H2利用COPII途径进行有效的病毒粒子组装和释放，解开 病毒生命周期中的一个关键检查点，可以作为新的抗病毒疗法的靶点。我们现在带来的是 在这项提案中，KSHV生物学和小分子抑制剂设计方面的专家合作 确定KSHV病毒粒子产生的细胞途径并开发抑制病毒的新策略 变速箱。为了实现这一目标，我们提出了两个具体的目标，包括(1)定义分子 KSHV Ks-Bcl2靶向NM23-H2激活Sar1介导的COPII高效转运机制 病毒粒子组装；(2)靶向KS-Bcl2-NM23-H2介导的COPII机制以阻断KSHV的传播。 这些目标将使用多学科方法来解决，这些方法结合了最先进的遗传学、 生物化学、活细胞成像和生理分析。我们预计，我们的研究将确定 宿主基因/途径在病毒组装和出口中起作用，并提供令人信服的体内验证 靶向NM23-H2依赖宿主机制可以消除病毒在内部和之间的传播 个人。

Project Summary There is an urgent and unmet need for the development of safe and effective therapeutics against serious human pathogens such as Kaposi's sarcoma herpesvirus (KSHV), which causes significant morbidity and mortality in immune-compromised individuals and remains a clinical challenge. Currently, no FDA approved therapeutics or vaccines are available for KSHV infection. Given that KSHV and other human herpesviruses hijack host proteins and pathways to complete their replication cycles and spread from cell to cell, strategies targeting these pathways and mechanisms will provide broad-spectrum genotype coverage and a high barrier to drug resistance. Herpesviruses including KSHV have long been known to exploit the COPII-mediated secretory pathway for their maturation. However, the mechanisms governing this process remain less understood. This project will fill this gap, capitalizing on our recent discovery that the host nucleoside diphosphates kinase NM23-H2, an important regulator of COPII vesicle transport, is exploited by KSHV for their virion morphogenesis and egress. We found that viral Bcl-2 of KSHV (ks-Bcl-2) directly interacted with, stabilized, and activated NM23-H2 during lytic phase of KSHV. Loss of NM23-H2 or mutations in ks-Bcl-2 that abolished NM23-H2 interaction severely impaired virion production. We thus hypothesize that KSHV activates host protein NM23-H2 to exploit the COPII pathway for efficient virion assembly and release, unraveling a key checkpoint in virus lifecycle that can be targeted for new antiviral therapeutics. We now bring within this proposal a collaboration of experts in KSHV biology and in design of small-molecule inhibitors to identify cellular pathway responsible for virion production of KSHV and develop a new strategy to dampen virus transmission. To achieve this goal, we propose two specific aims, including (1) defining the molecular mechanism by which KSHV ks-Bcl-2 targets NM23-H2 to activate Sar1-mediated COPII transport for efficient virion assembly; and (2) targeting ks-Bcl-2-NM23-H2-mediated COPII mechanism to block KSHV propagation. These aims will be addressed using multidisciplinary approaches that integrate state-of-the-art genetic, biochemistry, live-cell imaging, and physiological assays. Together, we anticipate that our studies will identify host genes/pathways that function in virus assembly and egress, and provide compelling in vivo validation that targeting NM23-H2-dependent host mechanism can abrogate virus transmission within and between individuals.