Ligand Binding and Signaling of Dendritic Cell Immunoreceptor (DCIR) Is Modulated by the Glycosylation of the Carbohydrate Recognition Domain

Karien Bloem,Arend-Jan Van Der Plas,Yvette Van Kooyk,Johan Garssen,Léon M J Knippels,Sandra J Van Vliet,Juan J García-Vallejo,Ilona M Vuist,Martina Lahmann

doi:10.1371/journal.pone.0066266

Abstract

C-type lectins are innate receptors expressed on antigen-presenting cells that are involved in the recognition of glycosylated pathogens and self-glycoproteins. Upon ligand binding, internalization and/or signaling often occur. Little is known on the glycan specificity and ligands of the Dendritic Cell Immunoreceptor (DCIR), the only classical C-type lectin that contains an intracellular immunoreceptor tyrosine-based inhibitory motif (ITIM). Here we show that purified DCIR binds the glycan structures Lewisb and Man3. Interestingly, binding could not be detected when DCIR was expressed on cells. Since DCIR has an N-glycosylation site inside its carbohydrate recognition domain (CRD), we investigated the effect of this glycan in ligand recognition. Removing or truncating the glycans present on purified DCIR increased the affinity for DCIR-binding glycans. Nevertheless, altering the glycosylation status of the DCIR expressing cell or mutating the N-glycosylation site of DCIR itself did not increase glycan binding. In contrast, cis and trans interactions with glycans induced DCIR mediated signaling, resulting in a decreased phosphorylation of the ITIM sequence. These results show that glycan binding to DCIR is influenced by the glycosylation of the CRD region in DCIR and that interaction with its ligands result in signaling via its ITIM motif.

Highlights

C-type lectin receptors (CLRs) are glycan binding receptors present on the surface of immune cells
We first generated a Dendritic Cell Immunoreceptor (DCIR)-Fc chimeric construct, in which the carbohydrate recognition domain (CRD) of DCIR was fused to the Fc tail of human IgG and used this DCIRFc construct in our CLR binding assay to search for new ligands for DCIR
To compare the glycan specificity of DCIR with that of dendritic cells (DCs)-SIGN, which shares the ability to bind Lewisb and Man3, we investigated glycan binding to immobilized DC-SIGNFc (Figure 1C-D)

Summary

Introduction

C-type lectin receptors (CLRs) are glycan binding receptors present on the surface of immune cells. Most CLRs expressed on dendritic cells (DCs), like dendritic cell-specific intercellular adhesion molecule 3 (ICAM-3) grabbing non-integrin (DC-SIGN) [2], macrophage galactose-type lectin (MGL) [3] and the mannose receptor (MR) [4], can function as antigen uptake receptors. The CLR dendritic cell immunoreceptor (DCIR) is expressed on a variety of immune cells, such as DCs, B-cells and monocytes [6]. ITIMs can interact with Src Homology 2 (SH2) domain containing protein tyrosine phosphatase (SHP) 1 or 2 or the SH2 domain containing inositol 5phosphatase (SHIP). These phosphatases are able to dephosphorylate signaling molecules [7].

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Results

Conclusion