Structural Flexibility of the Macrophage Dengue Virus Receptor CLEC5A

Abstract

The human C-type lectin-like molecule CLEC5A is a critical macrophage receptor for dengue virus. The binding of dengue virus to CLEC5A triggers signaling through the associated adapter molecule DAP12, stimulating proinflammatory cytokine release. We have crystallized an informative ensemble of CLEC5A structural conformers at 1.9-Å resolution and demonstrate how an on-off extension to a ␤-sheet acts as a binary switch regulating the flexibility of the molecule. This structural information together with molecular dynamics simulations suggests a mechanism whereby extracellular events may be transmitted through the membrane and influence DAP12 signaling. We demonstrate that CLEC5A is homodimeric at the cell surface and binds to dengue virus serotypes 1-4. We used blotting experiments, surface analyses, glycan microarray, and docking studies to investigate the ligand binding potential of CLEC5A with particular respect to dengue virus. This study provides a rational foundation for understanding the dengue virus-macrophage interaction and the role of CLEC5A in dengue virusinduced lethal disease.

Post-translational regulation and modifications of flavivirus structural proteins

“…As an evolutionary strategy, the use of CLRs as receptors for glycosylated flaviviruses appears to be a trade-off. Although the viruses gain the use of a subset of otherwise unavailable attachment molecules, they risk potentially triggering host lectin-based innate immunity such as activation of the com-plement cascade through mannose-binding lectin (Fuchs et al, 2010(Fuchs et al, , 2011 or inflammasome activation and cytokine stimulation through CLEC5A (Chen et al, , 2012Fernandez-Garcia et al, 2009;Watson et al, 2011;Wu et al, 2013).…”

Space-group and origin ambiguity in macromolecular structures with pseudo-symmetry and its treatment with the programZanuda

“…The structure of CLEC5A has been described previously (Watson et al, 2011;PDB entry 2yhf). Here, we focus on the critical steps of structure solution, reassignment of the origin of a substructure and the use of partial structure phases in MOLREP.…”

“…have very close orientations and their joint RF peak would have a multiplicity of six in a perfect twin, whereas the multiplicity of the joint RF peak from H and I (and from H 0 and I 0 ) would remain equal to two and the multiplicity for J (and J 0 ) would remain one. Four of the six monomers representing the two dominating orientations were found by conventional MR (MOLREP) and the remaining two were found using an MR search in the electron density calculated from the refined partial model as described in Watson et al (2011). The search included three steps: spherically averaged phased translation function, phased rotation function and phased translation function (SAPTF + PRF + PTF implemented in MOLREP; Vagin & Isupov, 2001).…”

“…As opposed to conventional RF, which is in effect a Patterson search, the orientation search using combination of SAPTF and PRF works with electron-density maps; therefore, it is local and is not affected by dominating orientations. The full model thus built was refined to R cryst /R free of 0.216/0.267 at 1.9 Å resolution (Watson et al, 2011). 4. Zanuda…”

Broad Antiviral Activity of Carbohydrate-Binding Agents Against Dengue Virus Infection

“…Mannan and fucose can inhibit the interaction between CLEC5A and DENV, indicating that the interaction is carbohydrate-dependent [64]. However, a glycan array demonstrated no binding signal between CLEC5A and N-glycans of mammals or insects [65]. The molecular interaction between CLEC5A and DENV remains to be elucidated.…”