Naïve and memory cell turnover as drivers of CCR5-to-CXCR4 tropism switch in human immunodeficiency virus type 1: implications for therapy

Early human immunodeficiency virus infection is characterized by the predominance of CCR5-tropic (R5) virus. However, in many individuals CXCR4-tropic (X4) virus appears in late infection. The reasons for this phenotypic switch are unclear. The patterns of chemokine receptor expression suggest that X4 and R5 viruses have a preferential tropism for na?ve and memory T cells, respectively. Since memory cells divide approximately 10 times as often as na?ve cells in uninfected individuals, a tropism for memory cells in early infection may provide an advantage. However, with disease progression both na?ve and memory cell division frequencies increase, and at low CD4 counts, the na?ve cell division frequency approaches that of memory cells. This may provide a basis for the phenotypic switch from R5 to X4 virus observed in late infection. We show that a model of infection using observed values for cell turnover supports this mechanism. The phenotypic switch from R5 to X4 virus occurs at low CD4 counts and is accompanied by a rapid rise in viral load and drop in CD4 count. Thus, low CD4 counts are both a cause and an effect of X4 virus dominance. We also investigate the effects of different antiviral strategies. Surprisingly, these results suggest that both conventional antiretroviral regimens and CCR5 receptor-blocking drugs will promote R5 virus over X4 virus.     

CCR5- and CXCR4-tropic HIV-1 are equally cytopathic for their T-cell targets in human lymphoid tissue

A rapid decline in T-cell counts and the progression to AIDS is often associated with a switch from CCR5-tropic (R5) HIV-1 to CXCR4-tropic (X4) HIV-1 or R5/X4 HIV-1 variants. Experimental infection with R5 HIV-1 causes less T-cell depletion than infection with X4 or R5/X4 variants in T-cell cultures, in ex vivo infected human lymphoid tissue and in SCID/hu mice, despite similar replication levels. Experimental genetic changes in those sequences in gp120 that transform R5 HIV-1 variants into otherwise isogenic X4 viruses make them highly cytopathic. Thus, it is now believed that R5 variants are less cytopathic for T cells than are X4 variants. However, it is not known why CCR5-mediated HIV-1 infection does not lead to a massive CD4+ T-cell depletion, as occurs in CXCR4-mediated HIV-1 infection. Here we demonstrate that R5 HIV-1 isolates are indeed highly cytopathic, but only for CCR5+/CD4+ T cells. Because these cells constitute only a small fraction of CD4+ T cells, their depletion does not substantially change the total CD4+ T-cell count. These results may explain why the clinical stage of HIV disease correlates with viral tropism.     

Accelerated In Vivo Proliferation of Memory Phenotype CD4+ T-cells in Human HIV-1 Infection Irrespective of Viral Chemokine Co-receptor Tropism

CD4⁺ T-cell loss is the hallmark of HIV-1 infection. CD4 counts fall more rapidly in advanced disease when CCR5-tropic viral strains tend to be replaced by X4-tropic viruses. We hypothesized: (i) that the early dominance of CCR5-tropic viruses results from faster turnover rates of CCR5⁺ cells, and (ii) that X4-tropic strains exert greater pathogenicity by preferentially increasing turnover rates within the CXCR4⁺ compartment. To test these hypotheses we measured in vivo turnover rates of CD4⁺ T-cell subpopulations sorted by chemokine receptor expression, using in vivo deuterium-glucose labeling. Deuterium enrichment was modeled to derive in vivo proliferation (p) and disappearance (d*) rates which were related to viral tropism data. 13 healthy controls and 13 treatment-naive HIV-1-infected subjects (CD4 143–569 cells/ul) participated. CCR5-expression defined a CD4⁺ subpopulation of predominantly CD45R0⁺ memory cells with accelerated in vivo proliferation (p = 2.50 vs 1.60%/d, CCR5⁺ vs CCR5⁻; healthy controls; P<0.01). Conversely, CXCR4 expression defined CD4⁺ T-cells (predominantly CD45RA⁺ naive cells) with low turnover rates. The dominant effect of HIV infection was accelerated turnover of CCR5⁺CD45R0⁺CD4⁺ memory T-cells (p = 5.16 vs 2.50%/d, HIV vs controls; P<0.05), naïve cells being relatively unaffected. Similar patterns were observed whether the dominant circulating HIV-1 strain was R5-tropic (n = 9) or X4-tropic (n = 4). Although numbers were small, X4-tropic viruses did not appear to specifically drive turnover of CXCR4-expressing cells (p = 0.54 vs 0.72 vs 0.44%/d in control, R5-tropic, and X4-tropic groups respectively). Our data are most consistent with models in which CD4⁺ T-cell loss is primarily driven by non-specific immune activation.     

Engineering HIV-resistant human CD4+ T cells with CXCR4-specific zinc-finger nucleases

HIV-1 entry requires the cell surface expression of CD4 and either the CCR5 or CXCR4 coreceptors on host cells. Individuals homozygous for the ccr5Δ32 polymorphism do not express CCR5 and are protected from infection by CCR5-tropic (R5) virus strains. As an approach to inactivating CCR5, we introduced CCR5-specific zinc-finger nucleases into human CD4+ T cells prior to adoptive transfer, but the need to protect cells from virus strains that use CXCR4 (X4) in place of or in addition to CCR5 (R5X4) remains. Here we describe engineering a pair of zinc finger nucleases that, when introduced into human T cells, efficiently disrupt cxcr4 by cleavage and error-prone non-homologous DNA end-joining. The resulting cells proliferated normally and were resistant to infection by X4-tropic HIV-1 strains. CXCR4 could also be inactivated in ccr5Δ32 CD4+ T cells, and we show that such cells were resistant to all strains of HIV-1 tested. Loss of CXCR4 also provided protection from X4 HIV-1 in a humanized mouse model, though this protection was lost over time due to the emergence of R5-tropic viral mutants. These data suggest that CXCR4-specific ZFNs may prove useful in establishing resistance to CXCR4-tropic HIV for autologous transplant in HIV-infected individuals.     

Mycobacterium tuberculosis Complex Enhances Susceptibility of CD4 T Cells to HIV through a TLR2-Mediated Pathway

Among HIV-infected individuals, co-infection with Mycobacterium tuberculosis is associated with faster progression to AIDS. We investigated the hypothesis that M. bovis BCG and M. tuberculosis (Mtb complex) could enhance susceptibility of CD4+ cells to HIV infection. Peripheral blood mononuclear cells (PBMCs) collected from healthy donors were stimulated with M. bovis BCG, M. tuberculosis CDC1551 and M. smegmatis MC(2)155, and stimulated CD4+ cells were infected with R5-and X4-tropic single replication-competent pseudovirus. CD4+ cells stimulated with Mtb complex showed enhanced infection with R5- and X4-tropic HIV, compared to unstimulated cells or cells stimulated with M. smegmatis (p<0.01). Treatment with TLR2 siRNA reversed the increased susceptibility of CD4+ cells with R5- and X4-tropic virus induced by Mtb complex. These findings suggest that TB infection and/or BCG vaccination may be a risk factor for HIV acquisition.     

Siglecs facilitate HIV-1 infection of macrophages through adhesion with viral sialic acids

BACKGROUND: Human immunodeficiency virus type 1 (HIV-1) infects macrophages effectively, despite relatively low levels of cell surface-expressed CD4. Although HIV-1 infections are defined by viral tropisms according to chemokine receptor usage (R5 and X4), variations in infection are common within both R5- and X4-tropic viruses, indicating additional factors may contribute to viral tropism. METHODOLOGY AND PRINCIPAL FINDINGS: Using both solution and cell surface binding experiments, we showed that R5- and X4-tropic HIV-1 gp120 proteins recognized a family of I-type lectin receptors, the Sialic acid-binding immunoglobulin-like lectins (Siglec). The recognition was through envelope-associated sialic acids that promoted viral adhesion to macrophages. The sialic acid-mediated viral-host interaction facilitated both R5-tropic pseudovirus and HIV-1(BaL) infection of macrophages. The high affinity Siglec-1 contributed the most to HIV-1 infection and the variation in Siglec-1 expression on primary macrophages from different donors was associated statistically with sialic acid-facilitated viral infection. Furthermore, envelope-associated sialoglycan variations on various strains of R5-tropic viruses also affected infection. CONCLUSIONS AND SIGNIFICANCE OF THE FINDINGS: Our study showed that sialic acids on the viral envelope facilitated HIV-1 infection of macrophages through interacting with Siglec receptors, and the expression of Siglec-1 correlated with viral sialic acid-mediated host attachment. This glycan-mediated viral adhesion underscores the importance of viral sialic acids in HIV infection and pathogenesis, and suggests a novel class of antiviral compounds targeting Siglec receptors.     

V3 loop-determined coreceptor preference dictates the dynamics of CD4+-T-cell loss in simian-human immunodeficiency virus-infected macaques

We used experimental infection of rhesus macaques with envelope gp120 V3 loop isogenic simian-human immunodeficiency virus (SHIV) molecular clones to more clearly define the impact of human immunodeficiency virus type 1 coreceptor usage in target cell selectivity and the rates of CD4+-T-cell depletion. Functional assays demonstrate that substitution of the V3 loop of the pathogenic CXCR4-tropic (X4) SHIV(SF33A2) molecular clone with the corresponding sequences from the CCR5-tropic (R5) SHIV(SF162P3) isolate resulted in a switch of coreceptor usage from CXCR4 to CCR5. The resultant R5 clone, designated SHIV(SF33A2(V3)), is replication competent in vivo, infecting two of two macaques by intravenous inoculation with peak viremia that is comparable to that seen in monkeys infected with X4-SHIV(SF33A2). But while primary infection with the X4 clone was accompanied by rapid and significant loss of peripheral and secondary lymphoid CD4+ T lymphocytes, infection with R5-SHIV(SF33A2(V3)) led to only a modest and transient loss. However, substantial depletion of intestinal CD4+ T cells was observed in R5-SHIV(SF33A2(V3))-infected macaques. Moreover, na?ve T cells that expressed high levels of CXCR4 were rapidly depleted in X4-SHIV(SF33A2)-infected macaques, whereas R5-SHIV(SF33A2(V3)) infection mainly affected memory T cells that expressed CCR5. These findings in a unique isogenic system illustrate that coreceptor usage is the principal determinant of tissue and target cell specificity of the virus in vivo and dictates the dynamics of CD4+-T-cell depletion during SHIV infection.     

TLR2 signaling renders quiescent naive and memory CD4+ T cells more susceptible to productive infection with X4 and R5 HIV-type 1

It has been recently demonstrated that circulating microbial products are responsible for a systemic immune activation in individuals infected with HIV-type 1. Bacterial products carry structural conserved motifs recognized by TLRs. Some TLR members are expressed in primary human CD4+ T cells but the precise functional role played by these pattern recognition receptors is still imprecise. In this study, we report that engagement of TLR2 in quiescent naive and memory CD4+ T cells leads to the acquisition of an effector-like phenotype. Interestingly, engagement of TLR2 renders both cell subsets more susceptible to productive infection with X4 virions and a higher virus production was seen with R5 viruses. It can be proposed that exposure of resting CD4+ T cells to pathogen-derived products that can engage TLR2 induces the acquisition of an effector-like phenotype in naive and memory CD4+ T lymphocytes, a phenomenon that might result in an acceleration of virus replication, immune dysregulation, and HIV-type 1-mediated disease progression.     

R5 human immunodeficiency virus type 1 (HIV-1) replicates more efficiently in primary CD4+ T-cell cultures than X4 HIV-1

In this report, we present evidence that R5 human immunodeficiency virus type 1 (HIV-1) replicates more efficiently in primary CD4+ T cells than X4 HIV-1. By comparing CD3/CD28-costimulated CD4+ T-cell cultures infected by several X4 and R5 HIV-1 strains, we determined that R5-infected CD4+ T cells produce more virus over time than X4-infected CD4+ T cells. In the first comparison, we found that more cells were infected by the X4-tropic strain LAI than by the R5-tropic strain JR-CSF and yet that higher levels of viral production were detected in the R5-infected cultures. The differential viral production was partially due to the severe cytopathic effects of the X4 virus. We also compared cultures infected with the isogenic HIV-1 strains NL4-3 (X4) and 49.5 (R5). We found that fewer cells were infected by the R5 strain, and yet similar levels of viral production were detected in both infected cultures. Cell death played less of a role in the differential viral production of these strains, as the cell viability remained comparable in both X4- and R5-infected cultures over time. The final comparison involved the primary R5-tropic isolate KP1 and the primary dual-tropic isolate KP2. Although both strains infected similar numbers of cells and induced comparable levels of cytopathicity, viral production was considerably higher in the R5-infected culture. In summary, these data demonstrate that R5 HIV-1 has an increased capacity to replicate in costimulated CD4+ T cells compared to X4 HIV-1.     

A preponderance of CCR5(+) CXCR4(+) mononuclear cells enhances gastrointestinal mucosal susceptibility to human immunodeficiency virus type 1 infection

The gastrointestinal mucosa harbors the majority of the body's CD4(+) cells and appears to be uniquely susceptible to human immunodeficiency virus type 1 (HIV-1) infection. We undertook this study to examine the role of differences in chemokine receptor expression on infection of mucosal mononuclear cells (MMCs) and peripheral blood mononuclear cells (PBMCs) by R5- and X4-tropic HIV-1. We performed in vitro infections of MMCs and PBMCs with R5- and X4-tropic HIV-1, engineered to express murine CD24 on the infected cell's surface, allowing for quantification of HIV-infected cells and their phenotypic characterization. A greater percentage of MMCs than PBMCs are infected by both R5- and X4-tropic HIV-1. Significant differences exist in terms of chemokine receptor expression in the blood and gastrointestinal mucosa; mucosal cells are predominantly CCR5(+) CXCR4(+), while these cells make up less than 20% of the peripheral blood cells. It is this cell population that is most susceptible to infection with both R5- and X4-tropic HIV-1 in both compartments. Regardless of whether viral isolates were derived from the blood or mucosa of HIV-1-infected patients, HIV-1 p24 production was greater in MMCs than in PBMCs. Further, the chemokine receptor tropism of these patient-derived viral isolates did not differ between compartments. We conclude that, based on these findings, the gastrointestinal mucosa represents a favored target for HIV-1, in part due to its large population of CXCR4(+) CCR5(+) target cells and not to differences in the virus that it contains.     

Humanized mice dually challenged with R5 and X4 HIV-1 show preferential R5 viremia and restricted X4 infection of CCR5+CD4+ T cells

CCR5-tropic (R5) immunodeficiency virus type 1 (HIV-1) strains are highly transmissible during the early stage of infection in humans, whereas CXCR4-tropic (X4) strains are less transmissible. This study aimed to explore the basis for early phase R5 and X4 HIV-1 infection in vivo by using humanized mice dually challenged with R5 HIV-1_NLAD8-D harboring DsRed and X4 HIV-1_NL-E harboring EGFP. Whereas R5 HIV-1 replicated well, X4 HIV-1 caused only transient viremia with variable kinetics; however, this was distinct from the low level but persistent viremia observed in mice challenged with X4 HIV-1 alone. Flow cytometric analysis of HIV-1-infected cells revealed that X4 HIV-1 infection of CCR5⁺CD4⁺ T cells was significantly suppressed in the presence of R5 HIV-1. X4 HIV-1 was more cytopathic than R5 HIV-1; however, this was not the cause of restricted X4 HIV-1 infection because there were no significant differences in the mortality rates of CCR5⁺ and CCR5⁻ cells within the X4 HIV-1-infected cell populations. Taken together, these results suggest that restricted infection of CCR5⁺CD4⁺ T cells by X4 HIV-1 (occurring via a still-to-be-identified mechanism) might contribute to the preferential transmission of R5 HIV-1 during the early phase of infection.     

Cannabinoid receptor 2-mediated attenuation of CXCR4-tropic HIV infection in primary CD4+ T cells

Agents that activate cannabinoid receptor pathways have been tested as treatments for cachexia, nausea or neuropathic pain in HIV-1/AIDS patients. The cannabinoid receptors (CB(1)R and CB(2)R) and the HIV-1 co-receptors, CCR5 and CXCR4, all signal via Gαi-coupled pathways. We hypothesized that drugs targeting cannabinoid receptors modulate chemokine co-receptor function and regulate HIV-1 infectivity. We found that agonism of CB(2)R, but not CB(1)R, reduced infection in primary CD4+ T cells following cell-free and cell-to-cell transmission of CXCR4-tropic virus. As this change in viral permissiveness was most pronounced in unstimulated T cells, we investigated the effect of CB(2)R agonism on to CXCR4-induced signaling following binding of chemokine or virus to the co-receptor. We found that CB(2)R agonism decreased CXCR4-activation mediated G-protein activity and MAPK phosphorylation. Furthermore, CB(2)R agonism altered the cytoskeletal architecture of resting CD4+ T cells by decreasing F-actin levels. Our findings suggest that CB(2)R activation in CD4+ T cells can inhibit actin reorganization and impair productive infection following cell-free or cell-associated viral acquisition of CXCR4-tropic HIV-1 in resting cells. Therefore, the clinical use of CB(2)R agonists in the treatment of AIDS symptoms may also exert beneficial adjunctive antiviral effects against CXCR4-tropic viruses in late stages of HIV-1 infection.     

Inhibition of Dual/Mixed Tropic HIV-1 Isolates by CCR5-Inhibitors in Primary Lymphocytes and Macrophages

Background

Dual/mixed-tropic HIV-1 strains are predominant in a significant proportion of patients, though little information is available regarding their replication-capacity and susceptibility against CCR5-antagonists in-vitro. The aim of the study was to analyze the replication-capacity and susceptibility to maraviroc of HIV-1 clinical isolates with different tropism characteristics in primary monocyte-derived-macrophages (MDM), peripheral-blood-mononuclear-cells (PBMC), and CD4⁺T-lymphocytes.

Methods

Twenty-three HIV-1 isolates were phenotipically and genotipically characterized as R5, X4 or dual (discriminated as R5⁺/X4, R5/X4, R5/X4⁺). Phenotypic-tropism was evaluated by multiple-cycles-assay on U87MG-CD4⁺-CCR5⁺−/CXCR4⁺-expressing cells. Genotypic-tropism prediction was obtained using Geno2Pheno-algorithm (false-positive-rate [FPR] = 10%). Replication-capacity and susceptibility to maraviroc were investigated in human-primary MDM, PBMC and CD4⁺T-cells. AMD3100 was used as CXCR4-inhibitor. Infectivity of R5/Dual/X4-viruses in presence/absence of maraviroc was assessed also by total HIV-DNA, quantified by real-time polymerase-chain-reaction.

Results

Among 23 HIV-1 clinical isolates, phenotypic-tropism-assay distinguished 4, 17 and 2 viruses with R5-tropic, dual/mixed-, and X4-tropic characteristics, respectively. Overall, viruses defined as R5⁺/X4-tropic were found with the highest prevalence (10/23, 43.5%). The majority of isolates efficiently replicated in both PBMC and CD4⁺T-cells, regardless of their tropism, while MDM mainly sustained replication of R5- or R5⁺/X4-tropic isolates; strong correlation between viral-replication and genotypic-FPR-values was observed in MDM (rho = 0.710;p-value = 1.4e-4). In all primary cells, maraviroc inhibited viral-replication of isolates not only with pure R5- but also with dual/mixed tropism (mainly R5⁺/X4 and, to a lesser extent R5/X4 and R5/X4⁺). Finally, no main differences by comparing the total HIV-DNA with the p24-production in presence/absence of maraviroc were found.

Conclusions

Maraviroc is effective in-vitro against viruses with dual-characteristics in both MDM and lymphocytes, despite the potential X4-mediated escape. This suggests that the concept of HIV-entry through one of the two coreceptors “separately” may require revision, and that the use of CCR5-antagonists in patients with dual/mixed-tropic viruses may be a therapeutic-option that deserves further investigations in different clinical settings.     

HIV-1 cell-to-cell spread overcomes the virus entry block of non-macrophage-tropic strains in macrophages

Macrophages (MΦ) are increasingly recognized as HIV-1 target cells involved in the pathogenesis and persistence of infection. Paradoxically, in vitro infection assays suggest that virus isolates are mostly T-cell-tropic and rarely MΦ-tropic. The latter are assumed to emerge under CD4+ T-cell paucity in tissues such as the brain or at late stage when the CD4 T-cell count declines. However, assays to qualify HIV-1 tropism use cell-free viral particles and may not fully reflect the conditions of in vivo MΦ infection through cell-to-cell viral transfer. Here, we investigated the capacity of viruses expressing primary envelope glycoproteins (Envs) with CCR5 and/or CXCR4 usage from different stages of infection, including transmitted/founder Envs, to infect MΦ by a cell-free mode and through cell-to-cell transfer from infected CD4+ T cells. The results show that most viruses were unable to enter MΦ as cell-free particles, in agreement with the current view that non-M-tropic viruses inefficiently use CD4 and/or CCR5 or CXCR4 entry receptors on MΦ. In contrast, all viruses could be effectively cell-to-cell transferred to MΦ from infected CD4+ T cells. We further showed that viral transfer proceeded through Env-dependent cell-cell fusion of infected T cells with MΦ targets, leading to the formation of productively infected multinucleated giant cells. Compared to cell-free infection, infected T-cell/MΦ contacts showed enhanced interactions of R5 M- and non-M-tropic Envs with CD4 and CCR5, resulting in a reduced dependence on receptor expression levels on MΦ for viral entry. Altogether, our results show that virus cell-to-cell transfer overcomes the entry block of isolates initially defined as non-macrophage-tropic, indicating that HIV-1 has a more prevalent tropism for MΦ than initially suggested. This sheds light into the role of this route of virus cell-to-cell transfer to MΦ in CD4+ T cell rich tissues for HIV-1 transmission, dissemination and formation of tissue viral reservoirs.     

Primary intestinal epithelial cells selectively transfer R5 HIV-1 to CCR5+ cells

The upper gastrointestinal tract is a principal route of HIV-1 entry in vertical transmission and after oral-genital contact. The phenotype of the newly acquired virus is predominantly R5 (CCR5-tropic) and not X4 (CXCR4-tropic), although both R5 and X4 viruses are frequently inoculated onto the mucosa. Here we show that primary intestinal (jejunal) epithelial cells express galactosylceramide, an alternative primary receptor for HIV-1, and CCR5 but not CXCR4. Moreover, we show that intestinal epithelial cells transfer R5, but not X4, viruses to CCR5+ indicator cells, which can efficiently replicate and amplify virus expression. Transfer was remarkably efficient and was not inhibited by the fusion blocker T-20, but was substantially reduced by colchicine and low (4 degrees C) temperature, suggesting endocytotic uptake and microtubule-dependent transcytosis of HIV-1. Our finding that CCR5+ intestinal epithelial cells select and transfer exclusively R5 viruses indicates a mechanism for the selective transmission of R5 HIV-1 in primary infection acquired through the upper gastrointestinal tract.     

Coreceptor tropism in human immunodeficiency virus type 1 subtype D: high prevalence of CXCR4 tropism and heterogeneous composition of viral populations

In human immunodeficiency virus type 1 (HIV-1) subtype B, CXCR4 coreceptor use ranges from approximately 20% in early infection to approximately 50% in advanced disease. Coreceptor use by non-subtype B HIV is less well characterized. We studied coreceptor tropism of subtype A and D HIV-1 collected from 68 pregnant, antiretroviral drug-naive Ugandan women (HIVNET 012 trial). None of 33 subtype A or 10 A/D-recombinant viruses used the CXCR4 coreceptor. In contrast, nine (36%) of 25 subtype D viruses used both CXCR4 and CCR5 coreceptors. Clonal analyses of the nine subtype D samples with dual or mixed tropism revealed heterogeneous viral populations comprised of X4-, R5-, and dual-tropic HIV-1 variants. In five of the six samples with dual-tropic strains, V3 loop sequences of dual-tropic clones were identical to those of cocirculating R5-tropic clones, indicating the presence of CXCR4 tropism determinants outside of the V3 loop. These dual-tropic variants with R5-tropic-like V3 loops, which we designated "dual-R," use CCR5 much more efficiently than CXCR4, in contrast to dual-tropic clones with X4-tropic-like V3 loops ("dual-X"). These observations have implications for pathogenesis and treatment of subtype D-infected individuals, for the association between V3 sequence and coreceptor tropism phenotype, and for understanding potential mechanisms of evolution from exclusive CCR5 use to efficient CXCR4 use by subtype D HIV-1.     

Rhabdovirus-Based Vectors with Human Immunodeficiency Virus Type 1 (HIV-1) Envelopes Display HIV-1-Like Tropism and Target Human Dendritic Cells

We describe replication-competent, vaccine strain-based rabies viruses (RVs) that lack their own single glycoprotein and express, instead, a chimeric RV-human immunodeficiency virus type 1 (HIV-1) envelope protein composed of the ectodomain and transmembrane domains of HIV-1 gp160 and the cytoplasmic domain of RV G. The envelope proteins from both X4 (NL4-3)- and R5X4 (89.6)-tropic HIV-1 strains were utilized. These recombinant viruses very closely mimicked an HIV-1- like tropism, as indicated by blocking experiments. Infection was inhibited by SDF-1 on cells expressing CD4 and CXCR4 for both viruses, whereas RANTES abolished infection of cells expressing CCR5 in addition to CD4 in studies of the RV expressing HIV-1(89.6) Env. In addition, preincubation with soluble CD4 or monoclonal antibodies directed against HIV-1 gp160 blocked the infectivity of both G-deficient viruses but did not affect the G-containing RVs. Our results also indicated that the G-deficient viruses expressing HIV-1 envelope protein, in contrast to wild-type RV but similar to HIV-1, enter cells by a pH-independent pathway. As observed for HIV-1, the surrogate viruses were able to target human peripheral blood mononuclear cells, macrophages, and immature and mature human dendritic cells (DC). Moreover, G-containing RV-based vectors also infected mature human DC, indicating that infection of these cells is also supported by RV G. The ability of RV-based vectors to infect professional antigen-presenting cells efficiently further emphasizes the potential use of recombinant RVs as vaccines.