Structural insight into the recognition of pathogen-derived phosphoglycolipids by C-type lectin receptor DCAR

Abstract

The C-type lectin receptors (CLRs) form a family of pattern recognition receptors that recognize numerous pathogens, such as bacteria and fungi, and trigger innate immune responses. The extracellular carbohydrate-recognition domain (CRD) of CLRs forms a globular structure that can coordinate a Ca2+ ion, allowing receptor interactions with sugar-containing ligands. Although well-conserved, the CRD fold can also display differences that directly affect the specificity of the receptors for their ligands. Here, we report crystal structures at 1.8–2.3 Å resolutions of the CRD of murine dendritic cell-immunoactivating receptor (DCAR, or Clec4b1), the CLR that binds phosphoglycolipids such as acylated phosphatidyl-myo-inositol mannosides (AcPIMs) of mycobacteria. Using mutagenesis analysis, we identified critical residues, Ala136 and Gln198, on the surface surrounding the ligand-binding site of DCAR, as well as an atypical Ca2+-binding motif (Glu-Pro-Ser/EPS168–170). By chemically synthesizing a water-soluble ligand analog, inositol-monophosphate dimannose (IPM2), we confirmed the direct interaction of DCAR with the polar moiety of AcPIMs by biolayer interferometry and co-crystallization approaches. We also observed a hydrophobic groove extending from the ligand-binding site that is in a suitable position to interact with the lipid portion of whole AcPIMs. These results suggest that the hydroxyl group-binding ability and hydrophobic groove of DCAR mediate its specific binding to pathogen-derived phosphoglycolipids such as mycobacterial AcPIMs.

Helicobacter pylori metabolites exacerbate gastritis through C-type lectin receptors

“…S4 G ), indicating that αCPG may also exert their effect in humans via an unidentified receptor. Our recent report of murine DCAR–ligand complex structure ( Omahdi et al, 2020 ) will help with the structure-based identification of human counterpart. Collectively, these results suggest that blockade of Hp0421 may prevent gastritis in humans by reducing the level of αCAG and αCPG.…”

Fc‐conjugated C‐type lectin receptors: Tools for understanding host–pathogen interactions

“…Table 1). The CRDs of hDCIR, mDCIR1, -3, -4, and DCAR contain an atypical EPS motif, while DCIR2 has a typical EPN motif influencing their ligand binding profiles, although other further elements within the CRDs contribute to specificity and affinity (Lee et al, 2011;Nagae et al, 2016;Omahdi et al, 2020).…”

“…M. tuberculosis Ac1PIM2 is a DCAR ligand, with the Fc fusion protein binding with higher affinity than either Fc Dectin-2 or Fc Mincle, although binding affinity decreases as the number of mannose residues increase (Omahdi et al, 2020;Toyonaga et al, 2016). Using mutated Fc DCAR fusion proteins the role of the EPS motif in binding AcPIM2 was examined, and the requirement for a neutral Ala136 residue surrounding the binding site was identified.…”

“…A hydrophobic grove leading away from the glycan-binding site contributes to the binding of the phosphate moiety of AcPIMs. Furthermore, exchanging the EPS for an EPN motif reduces affinity for AcPIM2, highlighting the importance of the non-typical sequence (Arai et al, 2020;Omahdi et al, 2020).…”

3D Structural View of Pathogen Recognition by Mammalian Lectin Receptors

“…Analogous to Mincle, the C-type lectin DCAR (dendritic cell immunoactivating receptor) is a receptor for acylated phosphatidyl-myo-inositol mannosides, which are expressed on the surface of mycobacteria. There is a crystal structure of DCAR lectin domain in complex with acyl chain-free, inositol-monophosphate dimannose at 1.8 Å ( Omahdi et al, 2020 ). The 3D structure shows that a mannose residue interacts with a calcium ion in the primary sugar-binding site in the canonical way.…”