The CLIC/GEEC pathway regulates particle uptake and formation of the virus-containing compartment (VCC) in HIV-1-infected macrophages

Abstract

HIV-1 particles are captured by the immunoglobulin superfamily member Siglec-1 on the surface of macrophages and dendritic cells, leading to particle internalization and facilitating trans-infection of CD4+ T cells. HIV-1-infected macrophages develop a unique intracellular compartment termed the virus-containing compartment (VCC) that exhibits characteristic markers of the late endosome and is enriched in components of the plasma membrane (PM). The VCC has been proposed as the major site of particle assembly in macrophages. Depleting Siglec-1 from macrophages significantly reduces VCC formation, implying a link between the capture and uptake of external HIV-1 particles and the development of VCCs within HIV-infected cells. We found that internalization of particles to the VCC was independent of clathrin, but required dynamin-2. CD98 and CD44, classical markers of the CLIC/ GEEC pathway, colocalized with Siglec-1 and HIV-1 particles within the VCC. Virus-like particles (VLPs) were taken up within CD98 and Siglec-1-enriched tubular membranes that migrated centripetally over time to form VCC-like structures. Inhibition of CLIC/GEEC-mediated endocytosis resulted in the arrest of captured HIV-1 particles on the macrophage cell surface, prevented VCC formation, and significantly reduced the efficiency of trans-infection of T cells. These findings indicate that following capture of virus by Siglec-1, particles follow an endocytic route to the VCC that requires both the CLIC/GEEC pathway and dynamin-2. We propose a model in which internalization of HIV-1 particles together with CLIC/GEEC membranes leads to the formation of the VCC in HIV-infected macrophages, creating an intracellular platform that facilitates further particle assembly and budding. of virus within tissues, and because infected macrophages contribute to HIV-related complications that include neurologic disorders, endocrine disorders, and cardiovascular disease. HIV infection of macrophages may also create a latent reservoir that persists in infected individuals despite administration of antiretroviral therapy. Here we focused on a compartment that forms in HIV-infected macrophages termed the virus-containing compartment or VCC. The VCC is an intracellular compartment that has been described as a site of assembly and as a holding compartment for viruses, and contains components of both intracellular organelles and the plasma membrane. We identified an endocytosis pathway that helps to explain the origins of the VCC, the CLIC/GEEC pathway. Inhibition of the CLIC/GEEC pathway prevented virus-like particle delivery to the VCC and inhibited transmission of infection from macrophages to T cells. The GTPase dynamin-2 was also required for delivery of HIV particles to the VCC. This study identifies a new facet of how HIV interacts with macrophages, suggesting that disruption of this pathway could be a therapeutic strategy with implications for HIV cure efforts.