Understand the role of CARD8 inflammasome in HIV-1 infection

摘要 尽管进行了抗逆转录病毒治疗（ART），但由于病毒感染的存在，HIV-1感染是不可治愈的。 主要存在于长寿命的静息记忆CD4 + T细胞中的潜在储库。病毒潜伏库 是根除HIV-1的主要障碍。消灭HIV-1的"休克和杀死"战略 涉及使用潜伏期逆转剂（LRA）诱导病毒基因表达， 使感染的细胞易受病毒致细胞病变效应或免疫清除的影响。然而，在这方面， 在接受ART治疗的患者中，潜伏在细胞中的HIV-1对病毒或免疫介导的 凋亡性细胞死亡，并且经常携带突变以逃避T细胞或抗体的识别。 因此，靶向病毒的不可变组分如必需的病毒抗原的新方法， 蛋白质功能是必需的。炎性小体是介导炎症反应的重要分子复合物， 炎症和细胞因微生物或危险信号而死亡。人类的 CARD 8炎性体的生理配体仍然未知。我们的研究表明 HIV-1蛋白酶降解CARD 8 N-末端结构域并释放C-末端， 炎性小体激活。在HIV-1感染的细胞中，病毒蛋白酶作为亚单位保持不活动 病毒性Gag-Pol多聚蛋白在病毒出芽后，它通过Gag-Pol二聚化被激活。我们 显示了通过非核苷逆转录过早激活细胞内HIV-1蛋白酶 转录酶抑制剂（NNRTI）触发CARD8感应和杀死HIV感染的巨噬细胞 CD4 + T细胞。CARD8可以检测到HIV-1的所有亚型， 多样性我们的发现表明，CARD 8炎性小体的靶向激活是一种有希望的治疗方法。 消除潜伏HIV-1宿主的战略。在这项拟议的研究中，我们将：1）进行离体 评估和优化基于CARD8的"震慑战略"， HIV-1在患者CD4 + T细胞中的表达; 2）了解HIV-1蛋白酶的分子机制- 介导的CARD8炎性小体激活; 3）测试CARD8为基础的"休克和杀伤"策略， HIV-1潜伏期的人源化小鼠模型。我们提出的研究将促进理解 CARD8炎性小体激活的生理机制及其在HIV-1中的作用 感染，这将提供关键的影响，艾滋病毒治疗的研究。

Abstract Despite antiretroviral therapy (ART), HIV-1 infection is not curable due to the presence of viral latent reservoirs primarily in long-lived resting memory CD4+ T cells. The viral latent reservoirs are the major barrier to HIV-1 eradication. The “shock and kill” strategy for HIV-1 eradication involves the use of latency-reversing agents (LRAs) to induce viral gene expression, which renders the infected cells susceptible to viral cytopathic effects or immune clearance. However, in ART-treated patients, the latent HIV-1 resides in cells is resistant to viral- or immune-mediated apoptotic cell death and often carries mutations to escape recognition by T cells or antibodies. Therefore, novel approaches to target immutable components of the virus such as essential viral protein functions are needed. Inflammasome is a critical molecular complex that mediates inflammation and pyroptotic cell death in response to microbial or danger signals. In humans, the physiologic ligand(s) for the CARD8 inflammasome remains unknown. Our studies demonstrate that HIV-1 protease degrades the CARD8 N-terminal domain and releases the C-terminus for inflammasome activation. In HIV-1-infected cells, the viral protease remains inactive as a subunit of viral Gag-Pol polyprotein. After virus budding, it is activated through Gag-Pol dimerization. We show that premature activation of intracellular HIV-1 protease by non-nucleoside reverse transcriptase inhibitors (NNRTI) triggers CARD8 sensing and killing of HIV-infected macrophages and CD4+ T cells. All subtypes of HIV-1 can be sensed by CARD8 despite substantial viral diversity. Our finding suggests that targeted activation of CARD8 inflammasome is a promising strategy to eliminate latent HIV-1 reservoirs. In this proposed study, we will: 1) perform ex vivo assessment and optimization of the CARD8-based “shock and kill strategy” for clearing latent HIV-1 in patient CD4+ T cells; 2) understand the molecular mechanisms of HIV-1 protease- mediated CARD8 inflammasome activation; 3) test CARD8-based “shock and kill” strategy in humanized mouse model of HIV-1 latency. Our proposed study will advance the understanding of the physiological mechanisms of CARD8 inflammasome activation and its role in HIV-1 infection, which will provide critical implications to HIV cure research.