Compartmentalized HIV-1 reservoir in intestinal monocytes/macrophages on antiretroviral therapy.

Background: The persistence of latently infected cells prevents a cure of HIV. The intestinal mucosa contains numerous target cells, and high levels of HIV-1 DNA persist in this compartment under ART. While CD4+ T cells are the best characterized reservoir of HIV-1, the role of long-lived intestinal macrophages in HIV-1 persistence on ART remains controversial.

Spermine and spermidine bind CXCR4 and inhibit CXCR4- but not CCR5-tropic HIV-1 infection.

Semen is an important vector for sexual HIV-1 transmission. Although CXCR4-tropic (X4) HIV-1 may be present in semen, almost exclusively CCR5-tropic (R5) HIV-1 causes systemic infection after sexual intercourse. To identify factors that may limit sexual X4-HIV-1 transmission, we generated a seminal fluid-derived compound library and screened it for antiviral agents.

Discovery of Bis-Imidazoline Derivatives as New CXCR4 Ligands.

The chemokine receptor CXCR4 and its ligand CXCL12 regulate leukocyte trafficking, homeostasis and functions and are potential therapeutic targets in many diseases such as HIV-1 infection and cancers. Here, we identified new CXCR4 ligands in the CERMN chemical library using a FRET-based high-throughput screening assay. These are bis-imidazoline compounds comprising two imidazole rings linked by an alkyl chain.

Protein overexpression can induce the elongation of cell membrane nanodomains.

In cell membranes, proteins and lipids are organized into submicrometric nanodomains of varying sizes, shapes, and compositions, performing specific functions. Despite their biological importance, the detailed morphology of these nanodomains remains unknown. Not only can they hardly be observed by conventional microscopy due to their small size, but there is no full consensus on the theoretical models to describe their structuring and their shapes.

Tracking receptor motions at the plasma membrane reveals distinct effects of ligands on CCR5 dynamics depending on its dimerization status.

G-protein-coupled receptors (GPCR) are present at the cell surface in different conformational and oligomeric states. However, how these states impact GPCRs biological function and therapeutic targeting remains incompletely known. Here, we investigated this issue in living cells for the CC chemokine receptor 5 (CCR5), a major receptor in inflammation and the principal entry co-receptor for Human Immunodeficiency Viruses type 1 (HIV-1).

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.

Modeling of CCR5 Recognition by HIV-1 gp120: How the Viral Protein Exploits the Conformational Plasticity of the Coreceptor.

The chemokine receptor CCR5 is a key player in HIV-1 infection. The cryo-EM 3D structure of HIV-1 envelope glycoprotein (Env) subunit gp120 in complex with CD4 and CCR5 has provided important structural insights into HIV-1/host cell interaction, yet it has not explained the signaling properties of Env nor the fact that CCR5 exists in distinct forms that show distinct Env binding properties. We used classical molecular dynamics and site-directed mutagenesis to characterize the CCR5 conformations stabilized by four gp120s, from laboratory-adapted and primary HIV-1 strains, and which were previously shown to bind differentially to distinct CCR5 forms and to exhibit distinct cellular tropisms.

Mechanisms of HIV-1 evasion to the antiviral activity of chemokine CXCL12 indicate potential links with pathogenesis.

HIV-1 infects CD4 T lymphocytes (CD4TL) through binding the chemokine receptors CCR5 or CXCR4. CXCR4-using viruses are considered more pathogenic, linked to accelerated depletion of CD4TL and progression to AIDS. However, counterexamples to this paradigm are common, suggesting heterogeneity in the virulence of CXCR4-using viruses.

Tuberculosis Exacerbates HIV-1 Infection through IL-10/STAT3-Dependent Tunneling Nanotube Formation in Macrophages.

The tuberculosis (TB) bacillus, Mycobacterium tuberculosis (Mtb), and HIV-1 act synergistically; however, the mechanisms by which Mtb exacerbates HIV-1 pathogenesis are not well known. Using in vitro and ex vivo cell culture systems, we show that human M(IL-10) anti-inflammatory macrophages, present in TB-associated microenvironment, produce high levels of HIV-1. In vivo, M(IL-10) macrophages are expanded in lungs of co-infected non-human primates, which correlates with disease severity.

CCR5 structural plasticity shapes HIV-1 phenotypic properties.

CCR5 plays immune functions and is the coreceptor for R5 HIV-1 strains. It exists in diverse conformations and oligomerization states. We interrogated the significance of the CCR5 structural diversity on HIV-1 infection.

CCR5 adopts three homodimeric conformations that control cell surface delivery.

Biophysical methods and x-ray crystallography have revealed that class A G protein-coupled receptors (GPCRs) can form homodimers. We combined computational approaches with receptor cross-linking, energy transfer, and a newly developed functional export assay to characterize the residues involved in the dimerization interfaces of the chemokine receptor CCR5, the major co-receptor for HIV-1 entry into cells. We provide evidence of three distinct CCR5 dimeric organizations, involving residues of transmembrane helix 5.

Buffering deleterious polymorphisms in highly constrained parts of HIV-1 envelope by flexible regions.

Background: Covariation is an essential process that leads to coevolution of parts of proteins and genomes. In organisms subject to strong selective pressure, coevolution is central to keep the balance between the opposite requirements of antigenic variation and retention of functionality. Being the viral component most exposed to the external environment, the HIV-1 glycoprotein gp120 constitutes the main target of the immune response.

Guidelines for cloning, expression, purification and functional characterization of primary HIV-1 envelope glycoproteins.

The trimeric HIV-1 envelope (Env) glycoproteins gp120 and gp41 mediate virus entry into target cells by engaging CD4 and the coreceptors CCR5 or CXCR4 at the cell surface and driving membrane fusion. Receptor/gp120 interactions regulate the virus life cycle, HIV infection transmission and pathogenesis. Env is also the target of neutralizing antibodies.

A single-residue change in the HIV-1 V3 loop associated with maraviroc resistance impairs CCR5 binding affinity while increasing replicative capacity.

Background: Maraviroc (MVC) is an allosteric CCR5 inhibitor used against HIV-1 infection. While MVC-resistant viruses have been identified in patients, it still remains incompletely known how they adjust their CD4 and CCR5 binding properties to resist MVC inhibition while preserving their replicative capacity. It is thought that they maintain high efficiency of receptor binding.

Targeting spare CC chemokine receptor 5 (CCR5) as a principle to inhibit HIV-1 entry.

CCR5 binds the chemokines CCL3, CCL4, and CCL5 and is the major coreceptor for HIV-1 entry into target cells. Chemokines are supposed to form a natural barrier against human immunodeficiency virus, type 1 (HIV-1) infection. However, we showed that their antiviral activity is limited by CCR5 adopting low-chemokine affinity conformations at the cell surface.

Modeling the allosteric modulation of CCR5 function by Maraviroc.

Maraviroc is a non-peptidic, low molecular weight CC chemokine receptor 5 (CCR5) ligand that has recently been marketed for the treatment of HIV infected individuals. This review discusses recent molecular modeling studies of CCR5 by homology to CXC chemokine receptor 4, their contribution to the understanding of the allosteric mode of action of the inhibitor and their potential for the development of future drugs with improved efficiency and preservation of CCR5 biological functions.

HIV-1 exploits CCR5 conformational heterogeneity to escape inhibition by chemokines.

CC chemokine receptor 5 (CCR5) is a receptor for chemokines and the coreceptor for R5 HIV-1 entry into CD4(+) T lymphocytes. Chemokines exert anti-HIV-1 activity in vitro, both by displacing the viral envelope glycoprotein gp120 from binding to CCR5 and by promoting CCR5 endocytosis, suggesting that they play a protective role in HIV infection. However, we showed here that different CCR5 conformations at the cell surface are differentially engaged by chemokines and gp120, making chemokines weaker inhibitors of HIV infection than would be expected from their binding affinity constants for CCR5.

β-arrestin-dependent activation of the cofilin pathway is required for the scavenging activity of the atypical chemokine receptor D6.

Chemokines promote the recruitment of leukocytes to sites of infection and inflammation by activating conventional heterotrimeric guanine nucleotide-binding protein (G protein)-coupled receptors (GPCRs). Chemokines are also recognized by a set of atypical chemokine receptors (ACRs), which cannot induce directional cell migration but are required for the generation of chemokine gradients in tissues. ACRs are presently considered "silent receptors" because no G protein-dependent signaling activity is observed after their engagement by cognate ligands.

CCR5 mutations distinguish N-terminal modifications of RANTES (CCL5) with agonist versus antagonist activity.

CCR5 is the major HIV-1 entry coreceptor. RANTES/CCL5 analogs are more potent inhibitors of infection than native chemokines; one class activates and internalizes CCR5, one neither activates nor internalizes, and a third partially internalizes without activation. Here we show that mutations in CCR5 transmembrane domains differentially impact the activity of these three inhibitor classes, suggesting that the transmembrane region of CCR5, a key interaction site for inhibitors, is a sensitive molecular switch, modulating receptor activity.

Allosteric model of maraviroc binding to CC chemokine receptor 5 (CCR5).

Maraviroc is a nonpeptidic small molecule human immunodeficiency virus type 1 (HIV-1) entry inhibitor that has just entered the therapeutic arsenal for the treatment of patients. We recently demonstrated that maraviroc binding to the HIV-1 coreceptor, CC chemokine receptor 5 (CCR5), prevents it from binding the chemokine CCL3 and the viral envelope glycoprotein gp120 by an allosteric mechanism. However, incomplete knowledge of ligand-binding sites and the lack of CCR5 crystal structures have hampered an in-depth molecular understanding of how the inhibitor works.

New insights into the mechanisms whereby low molecular weight CCR5 ligands inhibit HIV-1 infection.

CC chemokine receptor 5 (CCR5) is a G-protein-coupled receptor for the chemokines CCL3, -4, and -5 and a coreceptor for entry of R5-tropic strains of human immunodeficiency virus type 1 (HIV-1) into CD4(+) T-cells. We investigated the mechanisms whereby nonpeptidic, low molecular weight CCR5 ligands block HIV-1 entry and infection. Displacement binding assays and dissociation kinetics demonstrated that two of these molecules, i.

CXCR7 heterodimerizes with CXCR4 and regulates CXCL12-mediated G protein signaling.

The stromal cell-derived factor-1/CXCL12 chemokine engages the CXCR4 and CXCR7 receptors and regulates homeostatic and pathologic processes, including organogenesis, leukocyte homeostasis, and tumorigenesis. Both receptors are widely expressed in mammalian cells, but how they cooperate to respond to CXCL12 is not well understood. Here, we show that CXCR7 per se does not trigger G(alphai) protein-dependent signaling, although energy transfer assays indicate that it constitutively interacts with G(alphai) proteins and undergoes CXCL12-mediated conformational changes.

The CXCL12gamma chemokine displays unprecedented structural and functional properties that make it a paradigm of chemoattractant proteins.

The CXCL12gamma chemokine arises by alternative splicing from Cxcl12, an essential gene during development. This protein binds CXCR4 and displays an exceptional degree of conservation (99%) in mammals. CXCL12gamma is formed by a protein core shared by all CXCL12 isoforms, extended by a highly cationic carboxy-terminal (C-ter) domain that encompass four overlapped BBXB heparan sulfate (HS)-binding motifs.

Small neutralizing molecules to inhibit actions of the chemokine CXCL12.

The chemokine CXCL12 and the receptor CXCR4 play pivotal roles in normal vascular and neuronal development, in inflammatory responses, and in infectious diseases and cancer. For instance, CXCL12 has been shown to mediate human immunodeficiency virus-induced neurotoxicity, proliferative retinopathy and chronic inflammation, whereas its receptor CXCR4 is involved in human immunodeficiency virus infection, cancer metastasis and in the rare disease known as the warts, hypogammaglobulinemia, immunodeficiency, and myelokathexis (WHIM) syndrome. As we screened chemical libraries to find inhibitors of the interaction between CXCL12 and the receptor CXCR4, we identified synthetic compounds from the family of chalcones that reduce binding of CXCL12 to CXCR4, inhibit calcium responses mediated by the receptor, and prevent CXCR4 internalization in response to CXCL12.