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Microdomains of the C-type lectin DC-SIGN are portals for virus entry into dendritic cells.

Cambi A, de Lange F, van Maarseveen NM, Nijhuis M, Joosten B, van Dijk EM, de Bakker BI, Fransen JA, Bovee-Geurts PH, van Leeuwen FN, Van Hulst NF, Figdor CG - J. Cell Biol. (2004)

Bottom Line: The C-type lectin dendritic cell (DC)-specific intercellular adhesion molecule grabbing non-integrin (DC-SIGN; CD209) facilitates binding and internalization of several viruses, including HIV-1, on DCs, but the underlying mechanism for being such an efficient phagocytic pathogen-recognition receptor is poorly understood.During development of human monocyte-derived DCs, DC-SIGN becomes organized in well-defined microdomains, with an average diameter of 200 nm.Biochemical experiments and confocal microscopy indicate that DC-SIGN microdomains reside within lipid rafts.

View Article: PubMed Central - PubMed

Affiliation: Dept. of Tumor Immunology, Nijmegen Center for Molecular Life Sciences, University Medical Center Nijmegen, P.O. Box 9101, 6500 HB Nijmegen, Netherlands.

ABSTRACT
The C-type lectin dendritic cell (DC)-specific intercellular adhesion molecule grabbing non-integrin (DC-SIGN; CD209) facilitates binding and internalization of several viruses, including HIV-1, on DCs, but the underlying mechanism for being such an efficient phagocytic pathogen-recognition receptor is poorly understood. By high resolution electron microscopy, we demonstrate a direct relation between DC-SIGN function as viral receptor and its microlocalization on the plasma membrane. During development of human monocyte-derived DCs, DC-SIGN becomes organized in well-defined microdomains, with an average diameter of 200 nm. Biochemical experiments and confocal microscopy indicate that DC-SIGN microdomains reside within lipid rafts. Finally, we show that the organization of DC-SIGN in microdomains on the plasma membrane is important for binding and internalization of virus particles, suggesting that these multimolecular assemblies of DC-SIGN act as a docking site for pathogens like HIV-1 to invade the host.

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DC-SIGN microdomains enhances binding of virus-sized particles with respect to isolated DC-SIGN molecules. Beads of 1-μm diam are saturated with numerous coated ligand molecules that can engage simultaneous interactions with several individual DC-SIGN molecules. These multiple interactions may strengthen the binding both with random and clustered DC-SIGN. In contrast, when virus-sized particles are used, the contact surface and therefore, the number of ligand molecules is much smaller. Consequently, only interactions with DC-SIGN molecules in highly organized multiprotein assemblies may result in stable binding of virus particles.
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fig8: DC-SIGN microdomains enhances binding of virus-sized particles with respect to isolated DC-SIGN molecules. Beads of 1-μm diam are saturated with numerous coated ligand molecules that can engage simultaneous interactions with several individual DC-SIGN molecules. These multiple interactions may strengthen the binding both with random and clustered DC-SIGN. In contrast, when virus-sized particles are used, the contact surface and therefore, the number of ligand molecules is much smaller. Consequently, only interactions with DC-SIGN molecules in highly organized multiprotein assemblies may result in stable binding of virus particles.

Mentions: In vitro experiments with isolated recombinant C-type lectins suggested that these can oligomerize providing multiple surfaces to bind (multivalent) ligands (Drickamer, 1999). DC-SIGN has been shown to form tetramers stabilized by an α-helical stalk (Mitchell et al., 2001), and purified truncated forms of DC-SIGN containing the complete ECD were also shown to be able to bind ligands by surface plasmon resonance (Lozach et al., 2003). In these works, the affinity of isolated CRDs or complete ECDs of DC-SIGN for the ligand was measured and compared with the affinity of the membrane-bound form. Although no interactions with the ligand could be found using isolated CRDs, significant binding was detected if single CRDs were closely seeded (Kd = 48 nM) or if CRDs were part of a complete oligomeric-soluble ECD of DC-SIGN (Kd = 30 nM). The highest affinity was seen with membrane-bound DC-SIGN expressed on the surface of transfected cells (Kd = 3 nM), suggesting that the natural plasma membrane environment strongly influences the function of this receptor. To some extent, this mirrors our observations: DC-SIGN on the cell surface of intermediate DCs is randomly distributed and hardly binds virus-sized particles, whereas on immature DCs, the protein is organized in well-defined microdomains, which allows binding of 40-nm virus-sized particles (Fig. 6, A–F). In apparent contrast, we showed that on intermediate DCs, randomly distributed DC-SIGN was able to bind 1-μm beads, coated with ICAM-3 or gp120 (Fig. 3 B). This apparent discrepancy between 40-nm virus-sized particles and 1-μm beads can be explained by the fact that beads of 1-μm diam have ∼600-fold larger interaction surface saturated with numerous ligand molecules that can engage simultaneous interactions with several individual DC-SIGN molecules. These multiple interactions may signal into the cell and lead to a rapid recruitment of new DC-SIGN molecules, thus strengthening the binding. In contrast, when virus-sized particles are used, the contact surface and therefore, the number of ligand molecules is much smaller. Consequently, only interactions with DC-SIGN molecules in highly organized multiprotein assemblies can result in stable binding of virus-sized particles (Fig. 8). When a computer-aided simulation was performed to predict the capacity of round objects of different sizes to establish interactions with either random or clustered DC-SIGN molecules, we found that objects with a diameter in the range of virus sizes (40–200-nm diam) preferentially bind to clusters of DC-SIGN having the same size (unpublished data). In agreement with this model, no significant differences were detected in binding of intermediate and immature DCs to soluble gp120 or to ICAM-3-Fc chimeras, indicating that the capacity of each single DC-SIGN molecule to recognize and bind the ligand is comparable on both DC types (Fig. 6 G).


Microdomains of the C-type lectin DC-SIGN are portals for virus entry into dendritic cells.

Cambi A, de Lange F, van Maarseveen NM, Nijhuis M, Joosten B, van Dijk EM, de Bakker BI, Fransen JA, Bovee-Geurts PH, van Leeuwen FN, Van Hulst NF, Figdor CG - J. Cell Biol. (2004)

DC-SIGN microdomains enhances binding of virus-sized particles with respect to isolated DC-SIGN molecules. Beads of 1-μm diam are saturated with numerous coated ligand molecules that can engage simultaneous interactions with several individual DC-SIGN molecules. These multiple interactions may strengthen the binding both with random and clustered DC-SIGN. In contrast, when virus-sized particles are used, the contact surface and therefore, the number of ligand molecules is much smaller. Consequently, only interactions with DC-SIGN molecules in highly organized multiprotein assemblies may result in stable binding of virus particles.
© Copyright Policy
Related In: Results  -  Collection

Show All Figures
getmorefigures.php?uid=PMC2171967&req=5

fig8: DC-SIGN microdomains enhances binding of virus-sized particles with respect to isolated DC-SIGN molecules. Beads of 1-μm diam are saturated with numerous coated ligand molecules that can engage simultaneous interactions with several individual DC-SIGN molecules. These multiple interactions may strengthen the binding both with random and clustered DC-SIGN. In contrast, when virus-sized particles are used, the contact surface and therefore, the number of ligand molecules is much smaller. Consequently, only interactions with DC-SIGN molecules in highly organized multiprotein assemblies may result in stable binding of virus particles.
Mentions: In vitro experiments with isolated recombinant C-type lectins suggested that these can oligomerize providing multiple surfaces to bind (multivalent) ligands (Drickamer, 1999). DC-SIGN has been shown to form tetramers stabilized by an α-helical stalk (Mitchell et al., 2001), and purified truncated forms of DC-SIGN containing the complete ECD were also shown to be able to bind ligands by surface plasmon resonance (Lozach et al., 2003). In these works, the affinity of isolated CRDs or complete ECDs of DC-SIGN for the ligand was measured and compared with the affinity of the membrane-bound form. Although no interactions with the ligand could be found using isolated CRDs, significant binding was detected if single CRDs were closely seeded (Kd = 48 nM) or if CRDs were part of a complete oligomeric-soluble ECD of DC-SIGN (Kd = 30 nM). The highest affinity was seen with membrane-bound DC-SIGN expressed on the surface of transfected cells (Kd = 3 nM), suggesting that the natural plasma membrane environment strongly influences the function of this receptor. To some extent, this mirrors our observations: DC-SIGN on the cell surface of intermediate DCs is randomly distributed and hardly binds virus-sized particles, whereas on immature DCs, the protein is organized in well-defined microdomains, which allows binding of 40-nm virus-sized particles (Fig. 6, A–F). In apparent contrast, we showed that on intermediate DCs, randomly distributed DC-SIGN was able to bind 1-μm beads, coated with ICAM-3 or gp120 (Fig. 3 B). This apparent discrepancy between 40-nm virus-sized particles and 1-μm beads can be explained by the fact that beads of 1-μm diam have ∼600-fold larger interaction surface saturated with numerous ligand molecules that can engage simultaneous interactions with several individual DC-SIGN molecules. These multiple interactions may signal into the cell and lead to a rapid recruitment of new DC-SIGN molecules, thus strengthening the binding. In contrast, when virus-sized particles are used, the contact surface and therefore, the number of ligand molecules is much smaller. Consequently, only interactions with DC-SIGN molecules in highly organized multiprotein assemblies can result in stable binding of virus-sized particles (Fig. 8). When a computer-aided simulation was performed to predict the capacity of round objects of different sizes to establish interactions with either random or clustered DC-SIGN molecules, we found that objects with a diameter in the range of virus sizes (40–200-nm diam) preferentially bind to clusters of DC-SIGN having the same size (unpublished data). In agreement with this model, no significant differences were detected in binding of intermediate and immature DCs to soluble gp120 or to ICAM-3-Fc chimeras, indicating that the capacity of each single DC-SIGN molecule to recognize and bind the ligand is comparable on both DC types (Fig. 6 G).

Bottom Line: The C-type lectin dendritic cell (DC)-specific intercellular adhesion molecule grabbing non-integrin (DC-SIGN; CD209) facilitates binding and internalization of several viruses, including HIV-1, on DCs, but the underlying mechanism for being such an efficient phagocytic pathogen-recognition receptor is poorly understood.During development of human monocyte-derived DCs, DC-SIGN becomes organized in well-defined microdomains, with an average diameter of 200 nm.Biochemical experiments and confocal microscopy indicate that DC-SIGN microdomains reside within lipid rafts.

View Article: PubMed Central - PubMed

Affiliation: Dept. of Tumor Immunology, Nijmegen Center for Molecular Life Sciences, University Medical Center Nijmegen, P.O. Box 9101, 6500 HB Nijmegen, Netherlands.

ABSTRACT
The C-type lectin dendritic cell (DC)-specific intercellular adhesion molecule grabbing non-integrin (DC-SIGN; CD209) facilitates binding and internalization of several viruses, including HIV-1, on DCs, but the underlying mechanism for being such an efficient phagocytic pathogen-recognition receptor is poorly understood. By high resolution electron microscopy, we demonstrate a direct relation between DC-SIGN function as viral receptor and its microlocalization on the plasma membrane. During development of human monocyte-derived DCs, DC-SIGN becomes organized in well-defined microdomains, with an average diameter of 200 nm. Biochemical experiments and confocal microscopy indicate that DC-SIGN microdomains reside within lipid rafts. Finally, we show that the organization of DC-SIGN in microdomains on the plasma membrane is important for binding and internalization of virus particles, suggesting that these multimolecular assemblies of DC-SIGN act as a docking site for pathogens like HIV-1 to invade the host.

Show MeSH
Related in: MedlinePlus