from Medscape General Medicine™
Posted 05/23/2003

Majid Masso

http://www.medscape.com/viewarticle/455538

Dendritic cell (DC)-specific intercellular adhesion molecule 3 (ICAM-3) grabbing nonintegrin (DC-SIGN), a recently discovered type II transmembrane protein on DCs with a C-type lectin extracellular domain, is capable of binding ICAM-3 on resting T cells in the secondary lymphoid organs, providing the initial contact between these cells during the establishment of cell-mediated immunity. DC-SIGN also binds the HIV-1 envelope glycoprotein gp120 but does not function as a receptor for viral entry into DCs. Instead, DC-SIGN allows DCs in the peripheral mucosa to carry HIV-1 through the lymphatics in a "Trojan horse" fashion, where it is eventually delivered to the T cells. Also, the period of infectivity of HIV-1 is increased by several days as a result of DC-SIGN-gp120 binding, allowing for efficient trans-infection of T cells on DC arrival. The discovery of a cluster of related genes colocalized with DC-SIGN on chromosome 19p13.2-3, all displaying complex alternative splicing patterns, has led to a reexamination of the mechanisms underlying both the interactions between antigen-presenting cells (APCs) and T cells and the pathogenesis of HIV-1 infection.

Peripheral mucosal tissues, such as those lining the cervix and rectum, contain DCs that capture invading infectious microorganisms and intracellularly digest them for presentation with proteins of the major histocompatibility complex (MHC) as MHC-peptide complexes on the DC plasma membrane.^[1-4] Following antigen uptake, these immature DCs migrate to the T-cell areas of secondary organs to present the MHC-peptide ligands to antigen-specific T-cell receptors (TCRs) on resting T cells, while also maturing by altering their cell-surface receptor profile to make this initiation of cell-mediated immunity more effective.^[2] Although ICAM-3 is expressed at high levels on resting T cells, no high-affinity receptors for ICAM-3 on DCs had been previously identified.^[2]

Recently, though, an abundantly expressed receptor unique to DCs, referred to as DC-SIGN, has been discovered and shown to bind ICAM-3 with high affinity.^[2,5] Additionally, DC-SIGN has been observed to bind the HIV-1 envelope glycoprotein gp120.^[1,5] However, rather than being used by DCs to internalize HIV-1 for antigen processing, DC-SIGN allows DCs to unwittingly carry HIV-1 from the mucosal tissue to the lymph nodes while also increasing the period of infectivity of HIV-1 by several days and promoting efficient trans-infection of CD4⁺ T-helper cells.^[1,5]

from Medscape General Medicine™

Of all APCs of the immune system, including DCs, macrophages, and B cells, mature DCs are the most effective at activating naïve T cells.^[6,7] Mucosal DCs, although still immature, provide an important first-line of defense by ingesting foreign invaders via both pinocytosis and receptor-mediated endocytosis.^[2] While the receptor is recycled to the cell surface, the endocytic compartment that contains the internalized antigen fuses with a lysosome that contains hydrolytic enzymes, which degrade the antigen into oligopeptides.^[3,4] Next, an endosome containing DC class II MHC (MHC II) molecules fuses with the compartment containing the peptides.^[3,4,8] Each MHC II receptor binds a single peptide while being transported to the DC membrane, where MHC II is constitutively expressed.^[3,4,9] The MHC II-peptide receptor complex serves to initiate a signaling pathway for activation in CD4⁺ T cells via interaction with the TCR.^[3,4]

Also known as T-helper cells, these activated CD4⁺ T cells mobilize key players in the humoral and cell-mediated arms of the immune system and coordinate their activities via secretion of cytokines.^[3,4] The formation of a MHC II-peptide-TCR complex is specific: a large repertoire of TCRs with highly variable extracellular regions exist, since a TCR must recognize and bind both the MHC II molecule and the attached peptide.^[3,4] Binding of the CD4 receptor to MHC II helps to strengthen the interaction.^[3,4] Following uptake of antigen and while traveling through the lymphatics to reach the T cells, DCs differentiate into mature cells by both losing their ability to ingest antigen and expressing cell-surface receptors complementary to those on T cells for a more effective interaction.^[1,2] Specifically, a costimulatory signal provided by interactions between the CD28 molecule on the T cell and B7 on the DC is necessary for T-cell activation.^[3,4,10]

The story in virally infected DCs involves an alternate endogenous processing pathway.^[4,8] Most viral proteins are produced in the cytosol, where a ubiquinating enzyme complex covalently links several small ubiquitin proteins to a lysine-amino group near the amino terminus of the viral protein.^[4] Ubiquitin-protein complexes are targeted for degradation by a cylindrical protease complex called a proteasome, and eukaryotic cells use this general mechanism to regulate all protein levels, especially those of abnormal or foreign proteins.^[4] The viral peptides produced are transported by a transporter associated with antigen processing (TAP) protein into the rough endoplasmic reticulum, where each peptide is bound by a DC MHC I molecule.^[4,8] MHC I is similar to MHC II in that it also is constitutively expressed on the DC membrane.^[4,8] However unlike MHC II, the MHC I-peptide receptor complex is recognized by CD8⁺ T cells via their TCRs.^[3,4,8] In addition to the CD8 molecule binding MHC I for delivery of a more effective signal to the CD8⁺ T cell, there are other accessory receptors on the T-cell membrane that bind complementary receptor molecules on the mature DC.^[3,4] Activated CD8⁺ T cells differentiate into cytotoxic T-lymphocytes primed to destroy other infected cells.^[3,4]

An exception to the above pathway may occur following synthesis of viral envelope proteins, which are translocated into the endoplasmic reticulum and destined for the cell surface in endocytic compartments after passing through the Golgi.^[8] As described earlier, first lysosomes and then MHC II-containing endosomes fuse with these compartments. MHC II molecules bind peptides from these proteins, and these complexes are presented on the DC surface for T-helper cell recognition. Alternately, the envelope proteins may reach the cell membrane, and fusion with MHC II-containing endosomes may occur following endocytosis of these envelope proteins from the cell surface.^[8]

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The TCR, with its short cytoplasmic domains, is unable to singularly mediate signal transduction following interaction with the MHC-peptide complex.^[4] In fact, the TCR associates closely with CD3, a protein complex composed of 5 invariant polypeptide chains in 3 dimers, to form a TCR-CD3 complex.^[4] Immunoreceptor tyrosine-based activation motifs, located in the cytoplasmic domains of each CD3 chain, associate with the src-related protein tyrosine kinase (PTK) Fyn and the Zap-70 PTK to mediate signal transduction.^[4,10] In addition, the cytoplasmic domains of CD4 and CD8 associate with the src-related PTK Lck.^[4,10] It is believed that cross-linking of the TCR with CD4/CD8 brings the respective kinases into close proximity to facilitate activation of the CD4+/CD8+ T cell.^[4,10]

Initiating these events is the CD45 membrane protein on T cells, whose cytoplasmic domain exhibits tyrosine phosphatase activity.^[4] Association of CD45 with CD4/CD8 molecules results in dephosphorylation of tyrosine residues on Fyn and Lck to activate them.^[4] Next, the phosphorylation of a CD3 chain by Lck provides a binding site for Zap-70, which then phosphorylates downstream substrates, including phospholipase C (PLC-gamma) and mitogen-activated kinases.^[4,10] This phosphorylated CD3 chain is also capable of binding the SH2 domain of an SHC adaptor protein, which in turn activates Ras via the Grb-2 intermediate along with mSos.^[10]

PLC-gamma phosphorylation catalyzes the hydrolysis of phosphoinositide 4,5-bisphosphate (PIP₂) into diacylglycerol (DAG) and inositol 1,4,5-trisphosphate (IP₃).^[4,10] DAG activates protein kinase C, which phosphorylates downstream substrates, leading to activation of the nuclear transcription factor NF-kappaB.^[4] The generation of IP₃ leads to an increase in intracellular Ca²⁺ followed by activation of calcineurin, a calmodulin-dependent phosphatase that dephosphorylates the T cell-specific nuclear transcription factor NF-AT.^[4,10] In the nucleus, NF-AT dimerizes with AP1, and the complex binds to the interleukin 2 (IL-2) enhancer.^[4] Both NF-kappaB and NF-AT serve to activate several genes, especially IL-2.^[4,10] IL-2 is an autocrine growth factor whose secretion from the T cell and subsequent binding to the IL-2 receptor is required for proliferation and differentiation into either long-lived memory cells or effector cells capable of dictating an appropriate immune response.^[4] However, without the CD28-B7 costimulatory signal, little IL-2 is produced and the T cell is not able to induce self-proliferation.^[4,10] In addition to possibly increasing the half-life of the messenger RNA encoding IL-2, the costimulatory signal is believed to act in concert with the TCR-mediated signals to activate JNK, which phosphorylates c-Jun.^[4]

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As can be imagined, infection of T cells by HIV-1 can have deleterious effects on these signaling pathways, leading initially to activation of the pathways, for productive infection, followed by a defective response to stimulation by antigen via the TCR.^[10] The HIV-1 envelope glycoprotein gp120 binds with high affinity to the CD4 receptor, which is found on numerous cell types, including CD4⁺ T cells, DCs, monocytes/macrophages, microglia, and others.^[6,11] The additional envelope glycoprotein gp41 is noncovalently associated with gp120 and mediates fusion of the viral envelope with the plasma membrane of the targeted cell, whereby the HIV-1 genome and several proteins and enzymes are delivered to the cytoplasm.^[4,6,11] Without the presence of a coreceptor on the host cell, however, this active process of infection is not possible. During the primary and asymptomatic stages of infection, the predominant HIV-1 strain is one that uses the CCR5 chemokine coreceptor, which is found primarily on macrophages (M-tropic HIV-1).^[6,11-13] As the infection proceeds, a switch in the viral phenotype gives rise to a CD4⁺ T-lymphocyte-tropic isolate that uses the CXCR4 coreceptor (T-tropic HIV-1), resulting in a steep decline in CD4⁺ T cells and an AIDS diagnosis.^[6,11-13]

Individuals with a homozygous polymorphism in the CCR5 gene consisting of a 32-base pair deletion (delta32 CCR5) in the coding region do not express CCR5 on cell surfaces and are generally resistant to infection by M-tropic HIV-1.^[12] A few HIV-positive delta32 CCR5 homozygotes have been identified; however, they have been shown to carry the CXCR4-using T-tropic isolate.^[12] In addition, the defective delta32 CCR5 gene product in heterozygotes is capable of forming oligomers with wild-type CCR5 in the endoplasmic reticulum, resulting in less than 50% of cell surface CCR5.^[12] Disease progression tends to be significantly slower in HIV-positive patients that are delta32 CCR5 heterozygotes.^[6,11-13] Although these observations underscore the importance of CCR5 in HIV pathogenesis, polymorphisms of similar significance have not been described with CXCR4.^[4]

The natural ligands for CCR5 are the chemokines macrophage inflammatory protein 1 (MIP-1) alpha, MIP-1beta, RANTES, and monocyte chemoattractant protein 2, and they have been shown to inhibit replication of M-tropic HIV-1 viral isolates that use CCR5 as a coreceptor.^[7,14] Stromal cell-derived factor 1 is the natural ligand for CXCR4.^[3,6,7,11] Both the CCR5 and CXCR4 coreceptors belong to the family of G protein-coupled 7-transmembrane-segment receptors, which all contain an N-terminal region that is acidic and tyrosine rich.^[12,14] It has been shown that CCR5 is posttranslationally modified by O-linked glycosylation and by sulfation of its N-terminal tyrosines; CXCR4 also appears to be sulfated.^[12,14] In addition, sulfated tyrosines contribute in the binding of CCR5 to MIP-1 alpha, MIP-1 beta, and gp120/CD4 complexes.^[14] Finally, sulfated tyrosines are known to contribute to the efficiency of HIV-1 entry into cells that express CCR5 and CD4.^[12,14]

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The observation that DC-SIGN is also a receptor for the HIV-1 envelope glycoprotein gp120 has led investigators to question the implications of this discovery on HIV-1 pathogenesis. Immature DCs, while in the nonlymphoid mucosal tissues waiting to capture antigen, express the CCR5 receptor.^[1,7] As the DC matures, CCR5 downregulation is coupled with upregulation of the CXCR4 receptor.^[7] Chemotaxis of immature DCs is observed toward M-tropic but not T-tropic HIV-1, and exposure of immature DCs to M-tropic HIV-1 prevents migration toward CCR5 ligands.^[7] This recruitment of immature DCs in the anogenital tract during sexual transmission of HIV-1 may result in both productive infection of immature DCs, since they additionally express CD4, and binding of HIV-1 virions to DC-SIGN.^[7,19] It has been shown that DC-SIGN does not mediate HIV-1 entry into cells, as with the CD4/CCR5 receptor complex, even in the presence of either CD4 or CCR5 individually.^[1]

Following gp120-DC-SIGN binding and as the DC matures, HIV-1 is shuttled to the secondary lymphoid tissues by DCs, where DC-SIGN presents the bound viral particles to CD4/CCR5 complexes on T cells by a trans-receptor mechanism yet to be elucidated.^{[1,2,9,13,19]} Presumably, a conformational change in DC-SIGN-bound gp120 results in a more efficient interaction with CD4 and/or CCR5.^[1] In experiments using low viral titers to mimic in vivo conditions, CD4/CCR5-expressing cells were not infected without the help of DC-SIGN in trans.^[1] Furthermore, DC-SIGN was able to bind HIV-1 for more than 4 days, preserving its infectivity and possibly even protecting it via internalization within endocytic vesicles during the DC journey through the lymphatics.^[1]

The process by which HIV-1 exploits the migration of DCs to the T-cell compartments of lymphoid tissues to infect replication-permissive T cells has led to DCs being described as "Trojan horses" in this setting.^[7,13] The recruitment of immature DCs along virion gradients toward infected T cells, and vice versa, complicates matters even more, and it is believed to result in T-cell loss as a result of DC-T-cell syncytia formation.^[7] Factors required for upregulation of viral transcription are brought together by these syncytia, including Sp1 by T cells and NF-kappaB and Rel proteins by DCs, leading to efficient "factories" thought to be the primary source of viral production in vivo.^[7]

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Further study regarding DC-SIGNR and their spliced variants is imperative, because the discovery of this novel mechanism of DC presentation of HIV-1 to permissive cells may lead to a fuller understanding of HIV-1 transmission while offering new targets for blocking infection. Medications designed either to inhibit the ability of gp120 to bind DC-SIGN in the anogenital mucosa or to interfere with DC-T-cell interactions in the lymph nodes should be explored. Vaccine candidates designed to elicit mucosal antibodies against gp120-DC-SIGN binding should also be considered. Finally, precise details governing the molecular mechanisms involved in DC-SIGN-gp120 binding and infection of CD4⁺ T-helper cells in trans must be determined, allowing the search for possible treatments and vaccines that target these interactions to begin.