Payne Emily H, Ramalingam Dhivya, Fox Donald T, Klotman Mary E
Department of Pathology, Duke University, Durham, North Carolina, USA.
Department of Medicine, Duke University, Durham, North Carolina, USA.
J Virol. 2018 Jan 2;92(2). doi: 10.1128/JVI.01718-17. Print 2018 Jan 15.
Prior studies have found that HIV, through the Vpr protein, promotes genome reduplication (polyploidy) in infection-surviving epithelial cells within renal tissue. However, the temporal progression and molecular regulation through which Vpr promotes polyploidy have remained unclear. Here we define a sequential progression to Vpr-mediated polyploidy in human renal tubule epithelial cells (RTECs). We found that as in many cell types, Vpr first initiates G cell cycle arrest in RTECs. We then identified a previously unreported cascade of Vpr-dependent events that lead to renal cell survival and polyploidy. Specifically, we found that a fraction of G-arrested RTECs reenter the cell cycle. Following this cell cycle reentry, two distinct outcomes occur. Cells that enter complete mitosis undergo mitotic cell death due to extra centrosomes and aberrant division. Conversely, cells that abort mitosis undergo endoreplication to become polyploid. We further show that multiple small-molecule inhibitors of the phosphatidylinositol 3-kinase-related kinase (PIKK) family, including those that target ATR, ATM, and mTOR, indirectly prevent Vpr-mediated polyploidy by preventing G arrest. In contrast, an inhibitor that targets DNA-dependent protein kinase (DNA-PK) specifically blocks the Vpr-mediated transition from G arrest to polyploidy. These findings outline a temporal, molecularly regulated path to polyploidy in HIV-positive renal cells. Current cure-focused efforts in HIV research aim to elucidate the mechanisms of long-term persistence of HIV in compartments. The kidney is recognized as one such compartment, since viral DNA and mRNA persist in the renal tissues of HIV-positive patients. Further, renal disease is a long-term comorbidity in the setting of HIV. Thus, understanding the regulation and impact of HIV infection on renal cell biology will provide important insights into this unique HIV compartment. Our work identifies mechanisms that distinguish between HIV-positive cell survival and death in a known HIV compartment, as well as pharmacological agents that alter these outcomes.
先前的研究发现，人类免疫缺陷病毒（HIV）通过病毒蛋白R（Vpr）促进肾组织中存活的上皮细胞发生基因组复制（多倍体化）。然而，Vpr促进多倍体化的时间进程和分子调控机制仍不清楚。在此，我们确定了人肾小管上皮细胞（RTECs）中Vpr介导的多倍体化的连续进程。我们发现，与许多细胞类型一样，Vpr首先在RTECs中引发G期细胞周期停滞。然后，我们确定了一系列以前未报道的Vpr依赖性事件，这些事件导致肾细胞存活和多倍体化。具体而言，我们发现一部分G期停滞的RTECs重新进入细胞周期。在这种细胞周期重新进入之后，会出现两种不同的结果。进入完全有丝分裂的细胞由于额外的中心体和异常分裂而发生有丝分裂细胞死亡。相反，中止有丝分裂的细胞进行核内复制而成为多倍体。我们进一步表明，磷脂酰肌醇3激酶相关激酶（PIKK）家族的多种小分子抑制剂，包括那些靶向共济失调毛细血管扩张症突变基因（ATR）、共济失调毛细血管扩张症突变基因（ATM）和雷帕霉素靶蛋白（mTOR）的抑制剂，通过阻止G期停滞间接预防Vpr介导的多倍体化。相比之下，一种靶向DNA依赖性蛋白激酶（DNA-PK）的抑制剂特异性地阻断了Vpr介导的从G期停滞到多倍体化的转变。这些发现概述了HIV阳性肾细胞中多倍体化的时间性、分子调控途径。目前HIV研究中以治愈为重点的努力旨在阐明HIV在不同区室中长期持续存在的机制。肾脏被认为是这样一个区室，因为病毒DNA和mRNA在HIV阳性患者的肾组织中持续存在。此外，肾脏疾病是HIV感染背景下的一种长期合并症。因此，了解HIV感染对肾细胞生物学的调控和影响将为这个独特的HIV区室提供重要的见解。我们的工作确定了在已知的HIV区室中区分HIV阳性细胞存活和死亡的机制，以及改变这些结果的药物制剂。
