Structural Basis of Chemokine Sequestration by CrmD, a Poxvirus-Encoded Tumor Necrosis Factor Receptor

Abstract

Pathogens have evolved sophisticated mechanisms to evade detection and destruction by the host immune system. Large DNA viruses encode homologues of chemokines and their receptors, as well as chemokine-binding proteins (CKBPs) to modulate the chemokine network in host response. The SECRET domain (smallpox virus-encoded chemokine receptor) represents a new family of viral CKBPs that binds a subset of chemokines from different classes to inhibit their activities, either independently or fused with viral tumor necrosis factor receptors (vTNFRs). Here we present the crystal structures of the SECRET domain of vTNFR CrmD encoded by ectromelia virus and its complex with chemokine CX3CL1. The SECRET domain adopts a β-sandwich fold and utilizes its β-sheet I surface to interact with CX3CL1, representing a new chemokine-binding manner of viral CKBPs. Structure-based mutagenesis and biochemical analysis identified important basic residues in the 40s loop of CX3CL1 for the interaction. Mutation of corresponding acidic residues in the SECRET domain also affected the binding for other chemokines, indicating that the SECRET domain binds different chemokines in a similar manner. We further showed that heparin inhibited the binding of CX3CL1 by the SECRET domain and the SECRET domain inhibited RAW264.7 cell migration induced by CX3CL1. These results together shed light on the structural basis for the SECRET domain to inhibit chemokine activities by interfering with both chemokine-GAG and chemokine-receptor interactions.

Multipronged attenuation of macrophage-colony stimulating factor signaling by Epstein–Barr virus BARF1

“…Simply to block M-CSF:FMS signaling, a more direct approach and consequently a more efficient strategy would be for an EBV-encoded antagonist fully disguised as a receptor or ligand mimic to occupy the active surfaces required for signaling. This strategy has been used by various types of viruses in immune modulation, especially for sequestrating cytokines (27,28). In fact, when BARF1 first was identified as a binding partner for M-CSF, it was speculated to be an FMS homolog (13).…”

Structures of Orf Virus Chemokine Binding Protein in Complex with Host Chemokines Reveal Clues to Broad Binding Specificity

“…An additional group, more recently described, includes the SECRET (Smallpox virus Encoded Chemokine Receptor) domain from the variola virus protein, CrmB and the ectromelia virus (EV) protein, CrmD (Antonets et al, 2010;Xue et al, 2011). These proteins each contain two domains, one that binds tumor necrosis factor and another named SECRET that binds, with variable affinity, to selected CC and CXC chemokines (Antonets et al, 2010;Xue et al, 2011). Viral CBKPs are thought to facilitate successful viral replication by reducing leukocyte activation and chemotaxis to the site of infection.…”

“…Sequence conservation across the various vCKBPs is quite low (<15%) but structures, which have appeared for several members, often contain a characteristic b-sandwich topology, which has no resemblance to host proteins that bind chemokines (Alexander et al, 2002;Carfí et al, 1999;Bahar et al, 2008;Xue et al, 2011). The type-II CKBP family proteins, A41 and vCCI, have structures that are monomeric with a b-sandwich fold (Arnold and Fremont, 2006;Bahar et al, 2008;Carfí et al, 1999;Zhang et al, 2006).…”

“…The structure of mouse gMHV68 M3 protein in complex with a C chemokine has revealed that its chemokine binding site is distinct from the type-II family CKBPs, and that chemokine binding is associated with significant M3 domain rearrangements (Alexander et al, 2002). The structure of CrmD from EV also displays a b-sandwich fold, but binds to chemokines in a completely different fashion from both A41 and vCCI proteins (Xue et al, 2011) as well as the M3 family. There are currently no structures available for M-T7 or related CKBPs.…”

Immune modulation by virus-encoded secreted chemokine binding proteins

“…De novo modeling of VARV CrmB SECRET domain spatial structure revealed structural homology with CPXV 35-kDa and VACV A41 proteins (Antonets et al, 2010). The three dimensional structure of the SECRET domain alone or complexed with a low affinity binding chemokine, CX3CL1, was solved and confirmed that it shares the same ␤-sandwich topology with the 35-kDa and A41 poxvirus CKBPs (Xue et al, 2011) (Fig. 2C).…”