Department of Biomedical Sciences, University of North Dakota School of Medicine and Health Sciences, Grand Forks, North Dakota, USA.
FASEB J. 2022 Mar;36(3):e22184. doi: 10.1096/fj.202101722R.
The presence of latent HIV-1 reservoirs in the periphery and brain represents a major obstacle to curing HIV-1 infection. As an essential protein for HIV-1 viral replication, HIV-1 Tat, mostly intracellular, has been implicated in latent HIV-1 infection. From HIV-1 infected cells, HIV-1 Tat is actively secreted and bystander cells uptake the released Tat whereupon it is endocytosed and internalized into endolysosomes. However, to activate the HIV-1 LTR promoter and increase HIV-1 replication, HIV-1 Tat must first escape from the endolysosomes and then enter the nucleus. Here, we tested the hypothesis that HIV-1 Tat can accumulate in endolysosomes and contribute to the activation of latent HIV-1 in astrocytes. Using U87MG astrocytoma cells expressing HIV-1 LTR-driven luciferase and primary human astrocytes we found that exogenous HIV-1 Tat enters endolysosomes, resides in endolysosomes for extended periods of time, and induces endolysosome de-acidification as well as enlargement. The weak base chloroquine promoted the release of HIV-1 Tat from endolysosomes and induced HIV-1 LTR transactivation. Similar results were observed by activating endolysosome Toll-like receptor 3 (TLR3) and TLR7/8. Conversely, pharmacological block of TLRs and knocking down expression levels of TLR3 and TLR7, but not TLR8, prevented endolysosome leakage and attenuated HIV-1 Tat-mediated HIV-1 LTR transactivation. Our findings suggest that HIV-1 Tat accumulation in endolysosomes may play an important role in controlling HIV-1 transactivation.
潜伏的 HIV-1 储库存在于外周血和大脑中，这是治愈 HIV-1 感染的主要障碍。HIV-1 Tat 作为 HIV-1 病毒复制的必需蛋白，与潜伏的 HIV-1 感染有关。HIV-1 Tat 主要存在于细胞内，从 HIV-1 感染的细胞中被主动分泌出来，旁观者细胞摄取释放的 Tat，随后 Tat 被内吞并内化到内溶酶体中。然而，为了激活 HIV-1 LTR 启动子并增加 HIV-1 的复制，HIV-1 Tat 必须首先从内溶酶体中逃逸，然后进入细胞核。在这里，我们检验了假设，即 HIV-1 Tat 可以在内溶酶体中积累，并有助于激活星形胶质细胞中的潜伏 HIV-1。使用表达 HIV-1 LTR 驱动的荧光素酶的 U87MG 星形胶质细胞瘤细胞和原代人星形胶质细胞，我们发现外源性 HIV-1 Tat 进入内溶酶体，在内溶酶体中长时间停留，并诱导内溶酶体去酸化和扩大。弱碱性氯喹促进 HIV-1 Tat 从内溶酶体中释放，并诱导 HIV-1 LTR 转录激活。通过激活内溶酶体 Toll 样受体 3（TLR3）和 TLR7/8 也观察到类似的结果。相反，TLRs 的药理学阻断以及 TLR3 和 TLR7 的敲低表达水平，但不是 TLR8，可防止内溶酶体渗漏，并减弱 HIV-1 Tat 介导的 HIV-1 LTR 转录激活。我们的研究结果表明，HIV-1 Tat 在内溶酶体中的积累可能在控制 HIV-1 转录激活中发挥重要作用。
