HIV-AIDS Laboratory, Molecular Biology and Genetics Unit, Jawaharlal Nehru Centre for Advanced Scientific Research, Bangalore, India.
HIV-AIDS Laboratory, Molecular Biology and Genetics Unit, Jawaharlal Nehru Centre for Advanced Scientific Research, Bangalore, India
J Virol. 2020 Sep 15;94(19). doi: 10.1128/JVI.00503-20.
The magnitude of transcription factor binding site variation emerging in HIV-1 subtype C (HIV-1C), especially the addition of NF-κB motifs by sequence duplication, makes the examination of transcriptional silence challenging. How can HIV-1 establish and maintain latency despite having a strong long terminal repeat (LTR)? We constructed panels of subgenomic reporter viral vectors with varying copy numbers of NF-κB motifs (0 to 4 copies) and examined the profile of latency establishment in Jurkat cells. Surprisingly, we found that the stronger the viral promoter, the faster the latency establishment. Importantly, at the time of commitment to latency and subsequent points, Tat levels in the cell were not limiting. Using highly sensitive strategies, we demonstrate the presence of Tat in the latent cell, recruited to the latent LTR. Our data allude, for the first time, to Tat establishing a negative feedback loop during the late phases of viral infection, leading to the rapid silencing of the viral promoter. Over the past 10 to 15 years, HIV-1 subtype C (HIV-1C) has been evolving rapidly toward gaining stronger transcriptional activity by sequence duplication of major transcription factor binding sites. The duplication of NF-κB motifs is unique and exclusive to HIV-1C, a property not shared with any of the other eight HIV-1 genetic families. What mechanism(s) does HIV-1C employ to establish and maintain transcriptional silence despite the presence of a strong promoter and concomitant strong, positive transcriptional feedback is the primary question that we attempted to address in the present manuscript. The role that Tat plays in latency reversal is well established. Our work with the most common HIV-1 subtype, HIV-1C, offers crucial leads toward Tat possessing a dual role in serving as both a transcriptional activator and repressor at different phases of viral infection of the cell. The leads that we offer through the present work have significant implications for HIV-1 cure research.
HIV-1 亚型 C（HIV-1C）中出现的转录因子结合位点变异幅度很大，尤其是通过序列重复增加 NF-κB 基序，这使得检查转录沉默变得具有挑战性。尽管 HIV-1 具有强大的长末端重复序列（LTR），它是如何建立和维持潜伏期的？我们构建了具有不同 NF-κB 基序拷贝数（0 至 4 个拷贝）的亚基因组报告病毒载体面板，并在 Jurkat 细胞中检查了潜伏期建立的情况。令人惊讶的是，我们发现病毒启动子越强，潜伏期建立得越快。重要的是，在潜伏期建立和随后的时间点，细胞中的 Tat 水平并不是限制因素。我们使用高度敏感的策略证明了潜伏细胞中存在 Tat，它被募集到潜伏的 LTR 中。我们的数据首次暗示，Tat 在病毒感染的晚期建立负反馈回路，导致病毒启动子迅速沉默。在过去的 10 到 15 年中，HIV-1 亚型 C（HIV-1C）通过主要转录因子结合位点的序列重复，迅速进化为获得更强的转录活性。NF-κB 基序的重复是 HIV-1C 所独有的，与其他八种 HIV-1 遗传家族都没有共享。尽管存在强大的启动子和伴随的强大正向转录反馈，HIV-1C 采用什么机制来建立和维持转录沉默是我们在本研究中试图解决的主要问题。Tat 在潜伏期逆转中的作用已得到充分证实。我们对最常见的 HIV-1 亚型 HIV-1C 的研究为 Tat 在细胞病毒感染的不同阶段既作为转录激活剂又作为转录抑制剂发挥双重作用提供了重要线索。我们通过本研究提供的线索对 HIV-1 治愈研究具有重要意义。
