Single-Particle Tracking of Human Immunodeficiency Virus Type 1 Productive Entry into Human Primary Macrophages

Abstract

Macrophages are one of the major targets of human immunodeficiency virus (HIV-1), but the viral entry pathway remains poorly understood in these cells. Noninvasive virus labeling and single-virus tracking are effective tools for studying virus entry. Here, we constructed a quantum dot (QD)-encapsulated infectious HIV-1 particle to track viral entry at a single-particle level in live human primary macrophages. QDs were encapsulated in HIV-1 virions by incorporating viral accessory protein Vpr-conjugated QDs during virus assembly. With the HIV-1 particles encapsulating QDs, we monitored the early phase of viral infection in real time and observed that, during infection, HIV-1 was endocytosed in a clathrin-mediated manner; the particles were translocated into Rab5A-positive endosomes, and the core was released into the cytoplasm by viral envelope-mediated endosomal fusion. Drug inhibition assays verified that endosome fusion contributes to HIV-1 productive infection in primary macrophages. Additionally, we observed that a dynamic actin cytoskeleton is critical for HIV-1 entry and intracellular migration in primary macrophages. HIV-1 dynamics and infection could be blocked by multiple different actin inhibitors. Our study revealed a productive entry pathway in macrophages that requires both endosomal function and actin dynamics, which may assist in the development of inhibitors to block the HIV entry in macrophages.

Sensing of HIV-1 Entry Triggers a Type I Interferon Response in Human Primary Macrophages

“…HIV-1 entry is known to take place at the plasma membrane of T cells, but in other cell types, such as macrophages, it occurs most probably in the endosome following endocytosis (47) or macropinocytosis (48) depending on the cell type considered and the fusion protein carried by the virus. A recent study performed with quantum dots encapsulated in infectious HIV-1 particles further sup-ports the idea that viral entry occurs in macrophages at the endosomal level (49). These locations represent the first barriers of defense for a cell.…”

Single Virion Tracking Microscopy for the Study of Virus Entry Processes in Live Cells and Biomimetic Platforms

“…Simian virus 40 has the ability to induce actin rearrangement to further promote its own internalization whereas echovirus 1 does not [71,98]. Clathrin-mediated endocytosis is utilized for the uptake of many viruses including Australian bat lyssavirus [99], HIV [100], infectious hematopoietic necrosis virus [101], and rabiesvirus [74]. Other viruses are caveolinindependent or use macropinocytosis.…”

“…Adenovirus utilizes microtubules for movement within the cell, but detaches from them when the virions reach close proximity to the nucleus [119]. HIV cores move toward the nucleus with a microtuble-and actin-dependent motion; within the nucleus, the motion is slow and diffuse [100,116,120]. HIV RNA alone moves through the cytoplasm by diffusion [121].…”

“…This approach has been employed in the study of influenza virus, which fuses in early endosomes, revealing that the virus is able to enter a clathrin-dependent manner as well as a clathrin and caveolin independent fashion, and viral fusion can occur in both pathways [78]. In macrophages, HIV undergoes fusion in Rab5A positive endosomes [100]. Fusion of Dengue virus [70] and Ebola virus [11] colocalizes with Rab7, a marker of late stage endosomes.…”

Seeing the unseen: Imaging rotation in cells with designer anisotropic particles

“…Therefore, understanding the phagocytosis of particles has significant implications for both fundamental understanding of cell biology and biomedical innovation. Single-particle imaging and tracking methods have been used to dissect the complex interactions involved in the phagocytosis process (Chen et al, 2013b; Li et al, 2017). But, studies prior to our own measured only the translational motion of particles.…”