Identification of determinants of interaction between CXCR4 and gp120 of a dual-tropic HIV-1DH12 isolate.

Using a panel of chimeric viruses and their chimeric envelope glycoproteins, we have previously reported that the V1/V2 or the V3 regions of a dual-tropic primary human immunodeficiency virus type 1 (HIV-1) isolate (HIV-1DH12) could individually confer CXCR4 usage when introduced into the backbone of a macrophage-tropic (M-tropic) virus isolate (HIV-1AD8). In this study, chimeric CXCR4-CXCR2 chemokine receptors were employed to identify the determinants involved in the interaction between CXCR4 and the dual-tropic HIV-1DH12 gp120. Our results indicate that (i) HIV-1DH12 gp120 interacts primarily with the extracellular domains 1 (E1) and 2 (E2) of CXCR4, (ii) the V1/V2 and the V3 regions interact with different domains of CXCR4, and (iii) the V1/V2 region plays a more critical role in the interaction between CXCR4 and HIV-1DH12 gp120. Combining our data and those of others suggests that the pattern of CXCR4 usage is highly dependent on HIV-1 isolates. In addition, an M-tropic virus may evolve to become dual-tropic by first acquiring the ability to interact with CXCR4 through the V1/V2 region of gp120.     

Inhibition of R5X4 dualtropic HIV-1 primary isolates by single chemokine co-receptor ligands

The susceptibility of HIV-1 to chemokine-mediated inhibition may be lost as a consequence of the expanded usage of chemokine co-receptors frequently occurring in clade B isolates obtained from individuals with advanced disease. Since chemokine-based immune intervention is under intense investigation, it is crucial to determine its potential effect on primary dualtropic HIV isolates characterized by simultaneous utilization of CCR5 and CXCR4 chemokine co-receptors (R5X4 viruses). In the present study, the CCR5 binding chemokine regulated upon activation normal T cell expressed and secreted (RANTES) strongly inhibited the replication of two of eight primary R5X4 viruses in mitogen-activated primary peripheral blood mononuclear cells (PBMC). The CXCR4 antagonist AMD3100 efficiently suppressed the replication of other two HIV isolates, whereas the remaining four viruses were partially inhibited by treatment with either RANTES or AMD3100. The potency of chemokine-mediated inhibition was influenced by PBMC donor variability, but it was usually independent from the levels of expression of CCR5 or CXCR4. Dual co-receptor usage was maintained by the viruses after two serial passages on U87.CD4 astrocytic cell lines expressing exclusively either CCR5 or CXCR4. The gp120 env variable domains were sequenced before and after passages on U87.CD4 cells. Virus replication into U87.CD4-CXCR4 cells did not result in changes in the V3 region but perturbed the dominant env V4 sequence. Interestingly, double passage onto U87.CD4-CXCR4 cells determined the loss of susceptibility to RANTES inhibition. In conclusion, interference with CCR5 may efficiently inhibit the replication of at least some dualtropic HIV-1 strains, whereas forced CXCR4 usage may result in viral escape from CCR5-dependent inhibitory effects.     

Functional complementation of the envelope hypervariable V3 loop of human immunodeficiency virus type 1 subtype B by the subtype E V3 loop.

The hypervariable V3 loop within gp120 of human immunodeficiency virus type 1 (HIV-1) is the major determinant of cell tropism and the entry coreceptor usage of the virus. However, the information obtained thus far has been from only subtype B from North America and Europe, and little is known about other subtypes whose V3 amino acids differ by as much as 50% from subtype B V3. In this study, we examined the functional potential of the V3 element of the HIV-1 subtype E, the most crucial variant causing the AIDS epidemic throughout southeast Asia. A panel of HIV-1LAI recombinants was constructed by the overlap extension method, by which the LAI V3 loop was precisely replaced by that of the subtype E nonsyncytium-inducing (NSI) or syncytium-inducing (SI) variant. All of the recombinant viruses infected peripheral blood mononuclear cells, whereas only those with SI V3 infected MT2 cells, a CD4(+) T cell line. Consistently, the SI V3 recombinants used CXCR4, while the NSI V3 recombinants used CCR5 for infection of HOS-CD4(+) cells. Finally, only the NSI V3 sequence conferred CC-chemokine sensitivity on the parental virus. The data support the notion that the HIV-1 V3 loop consists of a relatively independent domain in gp120 and suggest that the subtype E V3 loop indeed contains the functional element to dictate the cell tropism, coreceptor preference, and chemokine sensitivity of the virus. These findings are of immediate importance in understanding V3 structure-function relationship and for examining phenotypic evolution of HIV-1 subtype E.     

Identification of an N-linked glycan in the V1-loop of HIV-1 gp120 influencing neutralization by anti-V3 antibodies and soluble CD4

Summary Glycosylation is necessary for HIV-1 gp120 to attain a functional conformation, and individual N-linked glycans of gp120 are important, but not essential, for replication of HIV-1 in cell culture. We have constructed a mutant HIV-1 infectious clone lacking a signal for N-linked glycosylation in the V1-loop of HIV-1 gp120. Lack of an N-linked glycan was verified by a mobility enhancement of mutant gp120 in SDS-gel electrophoresis. The mutated virus showed no differences in either gp120 content per infectious unit or infectivity, indicating that the N-linked glycan was neither essential nor affecting viral infectivity in cell culture. We found that the mutated virus lacking an N-linked glycan in the V1-loop of gp120 was more resistant to neutralization by monoclonal antibodies to the V3-loop and neutralization by soluble recombinant CD4 (sCD4). Both viruses were equally well neutralized by ConA and a conformation dependent human antibody IAM-2G12. This suggests that the N-linked glycan in the V1-loop modulates the three-dimensional conformation of gp120, without changing the overall functional integrity of the molecule.     

HIV-1 envelope, integrins and co-receptor use in mucosal transmission of HIV

It is well established that HIV-1 infection typically involves an interaction between the viral envelope protein gp120/41 and the CD4 molecule followed by a second interaction with a chemokine receptor, usually CCR5 or CXCR4. In the early stages of an HIV-1 infection CCR5 using viruses (R5 viruses) predominate. In some viral subtypes there is a propensity to switch to CXCR4 usage (X4 viruses). The receptor switch occurs in ~ 40% of the infected individuals and is associated with faster disease progression. This holds for subtypes B and D, but occurs less frequently in subtypes A and C. There are several hypotheses to explain the preferential transmission of R5 viruses and the mechanisms that lead to switching of co-receptor usage; however, there is no definitive explanation for either. One important consideration regarding transmission is that signaling by R5 gp120 may facilitate transmission of R5 viruses by inducing a permissive environment for HIV replication. In the case of sexual transmission, infection by HIV requires the virus to breach the mucosal barrier to gain access to the immune cell targets that it infects; however, the immediate events that follow HIV exposure at genital mucosal sites are not well understood. Upon transmission, the HIV quasispecies that is replicating in an infected donor contracts through a "genetic bottleneck", and often infection results from a single infectious event. Many details surrounding this initial infection remain unresolved. In mucosal tissues, CD4(+) T cells express high levels of CCR5, and a subset of these CD4(+)/CCR5(high) cells express the integrin α?β?, the gut homing receptor. CD4(+)/CCR5(high)/ α4β7(high) T cells are highly susceptible to infection by HIV-1 and are ideal targets for an efficient productive infection at the point of transmission. In this context we have demonstrated that the HIV-1 envelope protein gp120 binds to α?β? on CD4(+) T cells. On CD4(+)/CCR5(high)/ α4β7(high) T cells, α?β? is closely associated with CD4 and CCR5. Furthermore, α?β? is ~3 times the size of CD4 on the cell surface, that makes it a prominent receptor for an efficient virus capture. gp120-α?β? interactions mediate the activation of the adhesion-associated integrin LFA-1. LFA-1 facilitates the formation of virological synapses and cell-to-cell spread of HIV-1. gp120 binding to α?β? is mediated by a tripeptide located in the V1/V2 domain of gp120. Of note, the V1/V2 domain of gp120 has been linked to variations in transmission fitness among viral isolates raising the intriguing possibility that gp120-α?β? interactions may be linked to transmission fitness. Although many details remain unresolved, we hypothesize that gp120-α?β? interactions play an important role in the very early events following sexual transmission of HIV and may have important implication in the design of vaccine strategies for the prevention of acquisition of HIV infection.     

Clinical resistance to vicriviroc through adaptive V3 loop mutations in HIV-1 subtype D gp120 that alter interactions with the N-terminus and ECL2 of CCR5

The HIV-1 CCR5 co-receptor is a member of the chemokine receptor family of G-protein coupled receptors; for which a number of small molecule antagonists, such as vicriviroc (VCV), have been developed to inhibit HIV-1 R5-tropic replication. In this study, we analyzed an HIV-1 subtype D envelope gene from a clinical trial subject who developed complete resistance to VCV. The HIV-1 resistant envelope has six predominant amino acid changes in the V3 loop, together with one change in the C4 domain of gp120, which are fully responsible for the resistance phenotype. V3 loop mutations Q315E and R321G are essential for resistance to VCV, whereas E328K and G429R in C4 contribute significantly to the infectivity of the resistant variant. Collectively, these amino acid changes influenced the interaction of gp120 with both the N-terminus and ECL2 region of CCR5.     

Structure-based,targeted deglycosylation of HIV-1 gp120 and effects on neutralization sensitivity and antibody recognition

The human immunodeficiency virus (HIV-1) exterior envelope glycoprotein, gp120, mediates receptor binding and is the major target for neutralizing antibodies. Primary HIV-1 isolates are characteristically more resistant to broadly neutralizing antibodies, although the structural basis for this resistance remains obscure. Most broadly neutralizing antibodies are directed against functionally conserved gp120 regions involved in binding to either the primary virus receptor, CD4, or the viral coreceptor molecules that normally function as chemokine receptors. These antibodies are known as CD4 binding site (CD4BS) and CD4-induced (CD4i) antibodies, respectively. Inspection of the gp120 crystal structure reveals that although the receptor-binding regions lack glycosylation, sugar moieties lie proximal to both receptor-binding sites on gp120 and thus in proximity to both the CD4BS and the CD4i epitopes. In this study, guided by the X-ray crystal structure of gp120, we deleted four N-linked glycosylation sites that flank the receptor-binding regions. We examined the effects of selected changes on the sensitivity of two prototypic HIV-1 primary isolates to neutralization by antibodies. Surprisingly, removal of a single N-linked glycosylation site at the base of the gp120 third variable region (V3 loop) increased the sensitivity of the primary viruses to neutralization by CD4BS antibodies. Envelope glycoprotein oligomers on the cell surface derived from the V3 glycan-deficient virus were better recognized by a CD4BS antibody and a V3 loop antibody than were the wild-type glycoproteins. Absence of all four glycosylation sites rendered a primary isolate sensitive to CD4i antibody-mediated neutralization. Thus, carbohydrates that flank receptor-binding regions on gp120 protect primary HIV-1 isolates from antibody-mediated neutralization.     

Functional roles of the V3 hypervariable region of HIV-1 gp160 in the processing of gp160 and in the formation of syncytia in CD4+ cells.

To study the roles of the V3 hypervariable region (amino acid residues 301-336) of the HIV-1 envelope glycoprotein (gp160) during infection, we constructed recombinant vaccinia viruses that expressed either wild-type gp160 (v-env10) or mutant gp160 in which the V3 region was deleted (v-dl29.1 and v-dl29.2). In v-dl29.1 the V3 loop, formed by disulfide bonding between cysteine residues 301 and 336, was deleted from cys301 to cys336 (inclusive) and replaced by one serine residue. In v-dl29.2 the V3 loop was deleted from arg303 to ala334 and replaced by three residues: gly-ala-gly. Cells infected with all three recombinant vaccinia viruses expressed gp160 on the cell surface, but v-dl29.1-derived gp160 was not cleaved into gp120 and gp41 and did not bind the CD4 glycoprotein. In contrast, gp160 produced by recombinant v-dl29.2 was cleaved normally, and the mutant gp120 produced was secreted and retained binding activity to CD4+ cells. However, both mutants failed to induce syncytia in HeLa CD4+ cells. Thus a disulfide loop at the V3 portion of gp160 is required for cleavage into gp120 and gp41, presumably because the loop is required for proper tertiary structure. The sequence within the V3 loop, however, is not required for cleavage and secretion of gp160, or for binding to CD4+, but this region is essential for gp120-mediated syncytia formation.     

Small amino acid changes in the V3 loop of human immunodeficiency virus type 2 determines the coreceptor usage for CXCR4 and CCR5.

HIV-2 GH-1 is a molecular clone derived from an AIDS patient from Ghana. In contrast to the prototypic molecular clone ROD, GH-1 exhibits a narrow range of target cell specificity. By an infectious assay using HeLa-CD4 cells stably transfected with an HIV-1 LTR-beta-galactosidase reporter gene and transiently expressing various cloned chemokine receptors, we have examined the coreceptor usage of GH-1. In contrast to ROD, which uses principally CXCR4, GH-1 was found to use mainly if not exclusively CCR5 but not CXCR4. The distinct coreceptor usage of these two molecular clones allowed us to further map the region of gp120 that is important for the coreceptor specificity. By constructing a series of chimeric viruses between GH-1 and ROD, we have demonstrated that the C-terminal half of the V3 loop region of gp120 determines the differential coreceptor usage between GH-1 and ROD, and only a few amino acid differences in this region appear to be able to shift the specificity between CCR5 and CXCR4. Notably, the shift in the coreceptor usage from CCR5 to CXCR4 is associated with an increase in the net positive charge in the V3 region.     

Primary HIV-1 R5 isolates from end-stage disease display enhanced viral fitness in parallel with increased gp120 net charge

To better understand the evolution of the viral envelope glycoproteins (Env) in HIV-1 infected individuals who progress to AIDS maintaining an exclusive CCR5-using (R5) virus population, we cloned and sequenced the env gene of longitudinally obtained primary isolates. A shift in the electrostatic potential towards an increased net positive charge was revealed in gp120 of end-stage viruses. Residues with increased positive charge were primarily localized in the gp120 variable regions, with the exception of the V3 loop. Molecular modeling indicated that the modifications clustered on the gp120 surface. Furthermore, correlations between increased Env net charge and lowered CD4⁺ T cell counts, enhanced viral fitness, reduced sensitivity to entry inhibitors and augmented cell attachment were disclosed. In summary, this study suggests that R5 HIV-1 variants with increased gp120 net charge emerge in an opportunistic manner during severe immunodeficiency. Thus, we here propose a new mechanism by which HIV-1 may gain fitness.     

Maturation of the viral core enhances the fusion of HIV-1 particles with primary human T cells and monocyte-derived macrophages

HIV-1 infection requires fusion of viral and cellular membranes in a reaction catalyzed by the viral envelope proteins gp120 and gp41. We recently reported that efficient HIV-1 particle fusion with target cells is linked to maturation of the viral core by an activity of the gp41 cytoplasmic domain. Here, we show that maturation enhances the fusion of a variety of recombinant viruses bearing primary and laboratory-adapted Env proteins with primary human CD4+ T cells. Overall, HIV-1 fusion was more dependent on maturation for viruses bearing X4-tropic envelope proteins than for R5-tropic viruses. Fusion of HIV-1 with monocyte-derived macrophages was also dependent on particle maturation. We conclude that the ability to couple fusion to particle maturation is a common feature of HIV-1 Env proteins and may play an important role during HIV-1 replication in vivo.     

A novel denaturing heteroduplex tracking assay for genotypic prediction of HIV-1 tropism

Human immunodeficiency virus type 1 (HIV-1) is characterized by sequence variability. The third variable region (V3) of the HIV-1 envelope glycoprotein gp120 plays a key role in determination of viral coreceptor usage (tropism) and pathogenesis. This report describes a novel denaturing heteroduplex tracking assay (HTA) to analyze the genetic variation of HIV-1 V3 DNA. It improved upon previous non-denaturing HTA approaches to distinguish HIV-1 CCR5 and CXCR4 tropic viruses in mixed populations. The modifications included the use of a single-stranded fluorescent probe based on the consensus V3 sequence of HIV-1 CCR5 tropic viruses, Locked Nucleic Acid (LNA) "clamps" at both ends of heteroduplex DNA, and denaturing gel electrophoresis using Mutation Detection Enhancement (MDE(?)) as matrix. The analysis demonstrated that the LNA "clamps" increased its melting temperature (T(m)) and the thermal stability of heteroduplex DNA. The partially denaturing gel used a defined concentration of formamide, and significantly induced mobility shifts of heteroduplex DNA that was dependent on the number and patterns of DNA mismatches and insertions/deletions. This new technique successfully detected tropisms of 53 HIV-1 V3 clones of known tropism, and was able to separate and detect multiple V3 DNA variants encoding tropisms for CCR5 or CXCR4 in a mixture. The assay had the sensitivity to detect 0.5% minority species. This method may be useful as a research tool for analysis of viral quasispecies and for genotypic prediction of HIV-1 tropism in clinical specimens.     

Generation and properties of a human immunodeficiency virus type 1 isolate resistant to the small molecule CCR5 inhibitor, SCH-417690 (SCH-D)

We describe the generation of two genetically related human immunodeficiency virus type 1 (HIV-1) isolates highly (>20,000-fold) resistant to the small molecule CCR5 inhibitor, SCH-417690 (formerly SCH-D). Both viruses were cross-resistant to other small molecules targeting entry via CCR5, but they were inhibited by some MAbs against the same coreceptor on primary CD4+ T-cells. The resistant isolates remained sensitive to inhibitors of other stages of virus entry, and to replication inhibitors acting post-entry. Neither escape mutant could replicate detectably in peripheral blood mononuclear cells (PBMC) from two donors homozygous for the CCR5-Delta32 allele and both were insensitive to the CXCR4-specific inhibitor, AMD3100. Hence, the SCH-D escape mutants retained the R5 phenotype. One of the resistant isolates was, however, capable of replication in U87.CD4.CXCR4 cells and, after expansion in those cells, was sensitive to AMD3100 in primary CD4+ T-cells. Hence, some X4 variants may be present in this escape mutant swarm. A notable observation was that the SCH-D escape mutants were also cross-resistant to PSC-RANTES and AOP-RANTES, chemokine derivatives that are reported to down-regulate cell surface CCR5 almost completely. However, the extent to which CCR5 is down-regulated was dependent upon the detection MAb. Hence, the escape mutants may be using a CCR5 configuration that is only detected by some anti-CCR5 MAbs. Finally, two SCH-D-resistant clonal viruses revealed no amino acid changes in the gp120 V3 region relative to the parental viruses, in marked contrast to clones resistant to the AD101 small molecule CCR5 inhibitor that possess 4 such sequence changes. Several sequence changes elsewhere in gp120 (V2, C3 and V4) were present in the SCH-D-resistant clones. Their influence on the resistant phenotype remains to be determined.     

A simian immunodeficiency virus V3 loop mutant that does not efficiently use CCR5 or common alternative coreceptors is moderately attenuated in vivo

Sexually transmitted HIV-1 strains utilize the chemokine receptor CCR5 for viral entry and inhibitors targeting this coreceptor offer great promise for antiretroviral therapy. They also raise the question, however, whether viral variants exhibiting altered coreceptor interactions and resistance against these antiviral agents might still be pathogenic. In the present study, we analyzed a SIVmac239 envelope (Env) mutant (239DL) containing two mutations in the V3 loop which reduced viral entry via CCR5 by 10- to 20-fold, disrupted utilization of common alternative SIV coreceptors and changed the way Env engaged CCR5. To evaluate its replicative capacity and pathogenic potential in vivo we infected six rhesus macaques with 239DL. We found that 239DL replication was only slightly attenuated early during infection. Thereafter, a D324V change, which restored efficient CCR5 usage and coincided with 239wt-like levels of viral replication, emerged in two animals. In contrast, the viral geno- and phenotype remained stable in the other four rhesus macaques. Although these animals had about 100-fold reduced viral RNA loads relative to 239wt-infected macaques, they showed pronounced CD4 T-cell depletion in the intestinal lamina propria, and one developed opportunistic infections and died with simian AIDS. Thus, changes in the V3 loop that diminished CCR5 usage and altered Env interactions with CCR5 reduced the pathogenic potential of SIVmac in rhesus macaques but did not abolish it entirely.     

Simple determination of human immunodeficiency virus type 1 syncytium-inducing V3 genotype by PCR.

Human immunodeficiency virus type 1 (HIV-1) phenotype variability plays an important role in the pathogenesis of AIDS. The presence of syncytium-inducing (SI) HIV-1 isolates in infected individuals is associated with a rapid decline of CD4+ T cells, rapid disease progression, and reduced survival time after AIDS diagnosis. The strong association between the SI capacity of HIV-1 and the presence of positively charged amino acid residues at positions 306 and/or 320 in the third variable domain (V3) of gp120 could here be confirmed in 97% of 402 primary HIV-1 isolates, indicating that the V3 genotype may be useful for prediction of the viral phenotype. The V3 DNA sequences revealed a remarkably limited codon usage for the amino acid residues that are responsible for virus phenotype. On the basis of this limited SI-specific DNA sequence variation, four SI-specific oligonucleotides were designed for selective amplification of V3 from SI but not non-SI HIV-1 isolates. This PCR analysis allowed the prediction of the biological phenotype of HIV-1 isolates on the basis of the V3 genotype and may prove to be useful for monitoring SI capacity of HIV-1 isolates in infected individuals.     

Immunodominance and antigenic variation of the principal neutralization domain of HIV-1

The principal neutralization domain (PND) of HIV-1 is located in the third variable region (V3) of the envelope glycoprotein gp 120. Cross-reactivity of experimental and natural sera with recombinant proteins containing the V3 region of four HIV-1 variants showed that a group of viruses (among which HIV-1 MN) had antigenically similar V3 regions. The V3 regions of HIV-1 IIIB and HIV-1 RF were antigenically distinct. Antibodies raised to V3 domains of two isolates from the "MN group" neutralized HIV-1 MN (and not HIV-1 IIIB), thus confirming the antigenic similarity of V3 of these isolates to that of HIV-1 MN. Human antibodies to the V3 region were shown to be mainly directed to the central area in V3, where the neutralization domain is. In addition, antibody reactivities in sera of 397 Dutch and 39 Tanzanian HIV-1-infected individuals show that the V3 neutralization domain is highly antigenic, and that viruses from the MN group predominate in The Netherlands and to a lesser extent in Tanzania. Thus, if the protective value of antibodies to the PND can be established, then PND (poly)peptides derived from the MN group may be important components of a subunit vaccine against HIV-1.     

Fifteen years of env C2V3C3 evolution in six individuals infected clonally with human immunodeficiency virus type 1

The study of the evolution of human immunodeficiency virus type 1 (HIV-1) requires blood samples collected longitudinally and data on the approximate time point of infection. Although these requirements were fulfilled in several previous studies, the infectious sources were either unknown or heterogeneous genetically. In the present study, HIV-1 env C2V3C3 (nt 7029-7315) evolution was examined retrospectively in a cohort of hemophiliacs. Compared to other cohorts, the area of interest here was the infection of six hemophiliacs by the same virus strain, that is, the infecting viruses shared an identical genome. As expected, divergence from the founder sequence as well as interpatient divergence of the predominant virus strains increased significantly over time. Based on the V3 nucleotide sequences, CCR5 usage was predicted exclusively throughout the whole period of infection in all patients. Interestingly, common patterns of viral evolution were detected in the patients of the cohort. Four amino acid substitutions within the V3 loop emerged and persisted subsequently in five (positions 305 and 308 of the HXB2 gp120 reference sequence) and six patients (positions 325 and 328 in HXB2 gp120), respectively. These common changes within the V3 loop are likely to be enforced by HIV-1 specific immune response.     

Genetic composition of replication competent clonal HIV-1 variants isolated from peripheral blood mononuclear cells (PBMC), HIV-1 proviral DNA from PBMC and HIV-1 RNA in serum in the course of HIV-1 infection

The HIV-1 quasispecies in peripheral blood mononuclear cells (PBMC) is considered to be a mix of actively replicating, latent, and archived viruses and may be genetically distinct from HIV-1 variants in plasma that are considered to be recently produced.Here we analyzed the genetic relationship between gp160 env sequences from replication competent clonal HIV-1 variants that were isolated from PBMC and from contemporaneous HIV-1 RNA in serum and HIV-1 proviral DNA in PBMC of four longitudinally studied therapy naïve HIV-1 infected individuals.Replication competent clonal HIV-1 variants, HIV-1 RNA from serum, and HIV-1 proviral DNA from PBMC formed a single virus population at most time points analyzed. However, an under-representation in serum of HIV-1 sequences with predicted CXCR4 usage was sometimes observed implying that the analysis of viral sequences from different sources may provide a more complete assessment of the viral quasispecies in peripheral blood in vivo.     

Asymmetric HIV-1 co-receptor use and replication in CD4(+) T lymphocytes

Susceptibility to infection by the human immunodeficiency virus type-1 (HIV-1), both in vitro and in vivo, requires the interaction between its envelope (Env) glycoprotein gp120 Env and the primary receptor (R), CD4, and Co-R, either CCR5 or CXCR4, members of the chemokine receptor family. CCR5-dependent (R5) viruses are responsible for both inter-individual transmission and for sustaining the viral pandemics, while CXCR4-using viruses, usually dualtropic R5X4, emerge in ca. 50% of individuals only in the late, immunologically suppressed stage of disease. The hypothesis that such a major biological asymmetry is explained exclusively by the availability of cells expressing CCR5 or CXCR4 is challenged by several evidences. In this regard, binding of the HIV-1 gp120 Env to the entry R complex, i.e. CD4 and a chemokine R, leads to two major events: virion-cell membrane fusion and a cascade of cell signaling. While the fusion/entry process has been well defined, the role of R/Co-R signaling in the HIV-1 life cycle has been less characterized. Indeed, depending on the cellular model studied, the capacity of HIV-1 to trigger a flow of events favoring either its own latency or replication remains a debated issue. In this article, we will review the major findings related to the role of HIV R/Co-R signaling in the steps following viral entry and leading to viral spreading in CD4(+) T lymphocytes.