Enhanced CD4 Down-modulation by Late Stage HIV-1 nef Alleles Is Associated with Increased Env Incorporation and Viral Replication

Abstract

Three viral proteins participate in the down-modulation of CD4 in human immunodeficiency virus type 1 (HIV-1)-infected cells. The underlying mechanisms have been extensively investigated. However, the physiological relevance of this phenomenon remains poorly understood. To address the role of CD4 down-modulation in HIV-1 pathogenesis in vivo, we have characterized the functional properties of nef alleles isolated from seven HIV-1-infected patients at either the stage of AIDS (late alleles) or during the asymptomatic phase of infection (early alleles). HIV-1 variants carrying these nef alleles showed striking differences in CD4 down-modulation, virus infectivity, and replication properties. Infection of T cells with late strains resulted in production of viral particles with enhanced infectivity, as compared with variants carrying early nef alleles. These differences in infectivity were observed only when viruses were produced in cells with high levels of the viral receptor, suggesting a functional link between CD4 levels and the ability of Nef to down-modulate CD4 and to enhance viral infectivity. Similarly, late nef alleles were substantially more active than early nef genes in stimulating HIV-1 replication in high CD4-positive cells, including primary lymphocytes, but not in cells expressing low levels of the CD4 receptor. Single-round assays showed that differences in infectivity between late and early strains are largely reduced when evaluated in target cells with high levels of CD4, suggesting that the inhibitory effect occurs at the entry step. Supporting this, enhanced CD4 down-modulation by late nef alleles was associated with higher levels of envelope incorporation into viral particles, a phenomenon that likely accounted for the augmented infectivity. Our data suggest a mechanistic link between the Nef-mediated CD4 down-modulation and the enhancement of replication in CD4-positive lymphocytes. As progression to disease occurs, HIV-1 Nef variants with enhanced ability to down-modulate CD4 are selected. These strains efficiently overcome the deleterious effects of CD4 and replicate more aggressively in CD4-positive primary lymphocytes. These results highlight the importance of the virus-induced CD4 down-modulation in HIV-1 pathogenesis.

Nef Stimulates Human Immunodeficiency Virus Type 1 Replication in Primary T Cells by Enhancing Virion-Associated gp120 Levels: Coreceptor-Dependent Requirement for Nef in Viral Replication

“…We propose that, through removal of CD4 from the cell surface, Nef enhances viral infectivity by facilitating the incorporation of gp120 into nascent virions and by preventing virion incorporation of CD4. Similar conclusions were reached in studies by Lama and coworkers (4,39), who showed that expression of CD4 in 293T and Jurkat cells inhibited the incorporation of Env into HIV-1 particles and that Nef overcame the inhibition by downregulating CD4 expression. Our findings extend these conclusions to a physiologically relevant cell type, primary CD4 ϩ T cells.…”

“…Analysis of the Nef genes isolated from infected individuals and monkeys have shown that major histocompatibility complex (MHC) class I downregulation is a common feature of Nef isolates present at early stages of infection (10,51). In contrast, CD4 downregulation is more highly conserved in Nef genes from late stages of HIV and SIV infection (4,55) and in AIDS patients (10,66). Together, these studies suggest that while MHC I downregulation by Nef is important for HIV-1 persistence during the early asymptomatic phase of HIV-1 infection, downregulation of CD4 is more important for HIV-1 replication during late symptomatic stages of HIV-1 disease, when X4-tropic HIV-1 strains emerge.…”

Vpu Augments the Initial Burst Phase of HIV-1 Propagation and Downregulates BST2 and CD4 in Humanized Mice

“…It has been reported that CD4 expressed on the surface of infected cells has the potential to impair HIV-1 replication by (i) inhibiting the release of progeny virions (48), (ii) impairing the incorporation of Env into nascent virions (6,29), and (iii) sequestering Env from the virion budding site (3,6,29,31,32). We confirmed that CD4 suppresses the production of infectious virions (Fig.…”

Vpr and Vpu Are Important for Efficient Human Immunodeficiency Virus Type 1 Replication and CD4 + T-Cell Depletion in Human Lymphoid Tissue Ex Vivo

“…We have demonstrated that the ability of Nef to enhance HIV-1 replication in ex vivo-infected human lymphoid tissue correlates with its functional activity in CD4 downmodulation (25). This Nef function might be critical for the production of fully infectious viral particles from CD4 ϩ T cells (6). Vpu might enhance viral spread by both the same and different mechanisms because it downmodulates CD4 (34, 53) but it also increases the release of virus particles (49,54).…”