Effects of CCR5 and CD4 Cell Surface Concentrations on Infections by Macrophagetropic Isolates of Human Immunodeficiency Virus Type 1

Abstract

It has been proposed that changes in cell surface concentrations of coreceptors may control infections by human immunodeficiency virus type 1 (HIV-1), but the mechanisms of coreceptor function and the concentration dependencies of their activities are unknown. To study these issues and to generate stable clones of adherent cells able to efficiently titer diverse isolates of HIV-1, we generated two panels of HeLa-CD4/CCR5 cells in which individual clones express either large or small quantities of CD4 and distinct amounts of CCR5. The panels were made by transducing parental HeLa-CD4 cells with the retroviral vector SFF-CCR5. Derivative clones expressed a wide range of CCR5 quantities which were between 7.0 × 102 and 1.3 × 105 molecules/cell as measured by binding antibodies specific for CCR5 and the chemokine [125I]MIP1β. CCR5 was mobile in the membranes, as indicated by antibody-induced patching. In cells with a large amount of CD4, an unexpectedly low trace of CCR5 (between 7 × 102 and 2.0 × 103molecules/cell) was sufficient for maximal susceptibility to all tested HIV-1, including primary patient macrophagetropic and T-cell-tropic isolates. Indeed, the titers as indicated by immunoperoxidase staining of infected foci were as high as the tissue culture infectious doses measured in human peripheral blood mononuclear cells. In contrast, cells with a small amount of CD4 required a much larger quantity of CCR5 for maximal infection by macrophagetropic HIV-1 (ca. 1.0 × 104 to 2.0 × 104 molecules/cell). Cells that expressed low and high amounts of CD4 were infected with equal efficiencies when CCR5 concentrations were above threshold levels for maximal infection. Our results suggest that the concentrations of CD4 and CCR5 required for efficient infections by macrophagetropic HIV-1 are interdependent and that the requirements for each are increased when the other component is present in a limiting amount. We conclude that CD4 and CCR5 directly or indirectly interact in a concentration-dependent manner within a pathway that is essential for infection by macrophagetropic HIV-1. In addition, our results suggest that multivalent virus-receptor bonds and diffusion in the membrane contribute to HIV-1 infections.

Cell Entry of Enveloped Viruses

“…Similar slight shifts are involved in the change from the CCR5 to CXCR4 coreceptor usage during AIDS progression (Scarlatti et al, 1997). Small shifts in usages of highly similar receptors have also been reported during cell culture selections of subgroup B, D, and E avian leukosis viruses that all use polymorphic variants of the same TVB receptor (Taplitz and Coffin, 1997) and during cell culture selections of HIV-1 variants (Platt et al, 1998). Therefore, despite the rarity of receptor repertoire expansions throughout millions of years of retrovirus evolution, limited switches can occur within single infected animals.…”

“…The assembly of a complex containing several receptors may be a prerequisite for the membrane fusion steps that require multiple EnvGP molecules to cooperatively participate in this process. For example, in the case of HIV-1, the presence of more than one CD4 in contact with the virus enhances the infectivity dramatically and reduces the concentration of coreceptors needed for infection (Platt et al, 1998). Further investigation of this system has implied that a critical complex containing approximately four to six coreceptors is a requirement for infection, although it is not known whether this complex performs a transient role and then disperses or is maintained throughout the membrane fusion process (Kuhmann et al, 2000).…”

The broad anti-viral agent glycyrrhizin directly modulates the fluidity of plasma membrane and HIV-1 envelope

“…Because minimal changes in membrane fluidity are logarithmically responsible for infectivity [15], it is plausible that a reduction of fluidity by GL is the primary mechanism causing inhibition of viral infectivity. It has been proposed that membrane and envelope fluidities play a pivotal role in the formation of the wide fusion pore of enveloped viruses, by clustering a substantial number of activated viral fusion proteins derived from multiple ligand-receptor complexes [39][40][41][42][43]. In this respect, GL mainly acts against enveloped viruses.…”

Reduced Susceptibility to VIRIP-Based HIV-1 Entry Inhibitors Has a High Genetic Barrier and Severe Fitness Costs

“…The reasons for these target cell type and/or VIRIP derivativedependent differences remain to be determined and might involve differences in CD4 and coreceptor expression levels as well as altered affinity and accessibility of the target region in Env to the inhibitors. For example, TZM-bl cells express very high levels of CD4 and CCR5 (24) and are more susceptible to R5 than to X4 HIV-1 infection (25), while a substantially higher number of CD4 ϩ T cells in PBMC cultures express CXCR4 than CCR5. While VIR-353 is monomeric, VIR-576 is a dimer (11).…”