Extended Neck Regions Stabilize Tetramers of the Receptors DC-SIGN and DC-SIGNR

Abstract

The human cell surface receptors DC-SIGN (dendritic cell-specific intercellular adhesion molecule-grabbing nonintegrin) and DC-SIGNR (DC-SIGN-related) bind to oligosaccharide ligands found on human tissues as well as on pathogens including viruses, bacteria, and parasites. The extracellular portion of each receptor contains a membrane-distal carbohydrate-recognition domain (CRD) and forms tetramers stabilized by an extended neck region consisting of 23 amino acid repeats. Cross-linking analysis of full-length receptors expressed in fibroblasts confirms the tetrameric state of the intact receptors. Hydrodynamic studies on truncated receptors demonstrate that the portion of the neck of each protein adjacent to the CRD is sufficient to mediate the formation of dimers, whereas regions near the N terminus are needed to stabilize the tetramers. Some of the intervening repeats are missing from polymorphic forms of DC-SIGNR. Two different crystal forms of truncated DC-SIGNR comprising two neck repeats and the CRD reveal that the CRDs are flexibly linked to the neck, which contains ␣-helical segments interspersed with non-helical regions. Differential scanning calorimetry measurements indicate that the neck and CRDs are independently folded domains. Based on the crystal structures and hydrodynamic data, models for the full extracellular domains of the receptors have been generated. The observed flexibility of the CRDs in the tetramer, combined with previous data on the specificity of these receptors, suggests an important role for oligomerization in the recognition of endogenous glycans, in particular those present on the surfaces of enveloped viruses recognized by these proteins.The dendritic cell receptor DC-SIGN 1 and the closely related DC-SIGNR found on endothelial cells have been of considerable interest because of their ability to enhance infection of T cells by the human immunodeficiency virus and because of their interactions with glycoproteins found on the surface of other enveloped viruses (1, 2). The physiological functions of these receptors are not known with certainty, but DC-SIGN has been ascribed roles in binding to intercellular adhesion molecule 3 on T cells and intercellular adhesion molecule 2 on endothelia, as well as in uptake of pathogens (3-7). Although DC-SIGN binds to a broad spectrum of glycans, it has highest affinity for N-linked high mannose oligosaccharides and fucose-containing structures that are found both as blood group antigens in human tissues and on the surfaces of certain parasites (7,8). DC-SIGNR binds only to the high mannose oligosaccharides, and unlike DC-SIGN, it does not mediate uptake and degradation of glycoconjugates (8).DC-SIGN and DC-SIGNR share nearly 80% sequence identity and are closely similar in overall architecture. Both receptors are type II transmembrane proteins in which C-terminal C-type carbohydrate-recognition domains (CRDs) are projected from the cell surface by a neck comprising a series of highly conserved 23-amino acid repeats. There are seven complete re...

Multiple Modes of Binding Enhance the Affinity of DC-SIGN for High Mannose N-Linked Glycans Found on Viral Glycoproteins

“…The presence of Man␣1-2Man enhances the affinity of oligomannose toward these receptors, even though by itself this disaccharide binds only slightly more strongly than mannose. The CRD has an intrinsically high affinity for oligomannose structures, and tetramerization likely provides further avidity enhancements for arrays of such structures (6,26). The ability of DC-SIGN and DC-SIGNR to bind high mannose glycans in multiple orientations may facilitate this multivalent binding of clusters of CRDs to glycans displayed in various arrangements on the surface of the virus, as proposed previously for cell surface recognition by mannose-binding proteins (22).…”

The evolutionary history of the CD209 (DC-SIGN) family in humans and non-human primates

“…The type and number of the neck-region repeats are important in dimer formation and stabilization of protein tetramers. 5 Thus, neck-length variation could influence the pathogen-binding properties of these lectins. In previous analyses in primates, 1 CD209 and CD209L have variable numbers of repeats.…”

HIV-1 Transmission by Dendritic Cell-specific ICAM-3-grabbing Nonintegrin (DC-SIGN) Is Regulated by Determinants in the Carbohydrate Recognition Domain That Are Absent in Liver/Lymph Node-SIGN (L-SIGN)

“…studies demonstrated that repeat domains of DC-SIGN and L-SIGN contribute to the overall integrity of the tetramer on the cell surface (16,17). To investigate the impact of the repeat region of L-SIGN on HIV-1 trans-mission, we obtained Raji cells expressing L-SIGN with five, six, or seven repeats and tested these cells in HIV-1 infection assays.…”

“…Although no structure of dimeric or tetrameric DC-SIGN is currently available, inferences regarding the positioning of Trp-258 relative to other DC-SIGN subunits can be made using a model of the tetrameric L-SIGN receptor (Fig. 9B), which was constructed from the crystal structure of the L-SIGN dimer (16,45). In this model, Lys-270, the L-SIGN homologue of Trp-258, is positioned near the interface between ␣-helix 1 (in portion 1, red, of our CRD swaps) and ␣-helix 2 (in portion 2, green) on adjacent subunits, suggesting a possible role for either or both these residues in dimer formation and/or stabilization.…”