NKG2D is a key component of cytotoxic antitumor and antiviral responses. Multiple viruses evade NKG2D recognition by blocking NKG2D ligand expression on infected cells. In contrast, cowpox virus targets NKG2D directly by encoding a secreted antagonist, Orthopoxvirus MHC Class I-like Protein (OMCP). We have previously reported that OMCP also binds to the orphan receptor FcRL5 on innate B cells. Here, we demonstrate that mammalian-derived, glycosylated OMCP binds NKG2D but not FcRL5. Cowpox viruses either lacking OMCP, or expressing an NKG2D-binding deficient mutant, are significantly attenuated in wild type and FcRL5-deficient mice but not NKG2D-deficient mice, demonstrating that OMCP is critical in subverting NKG2D-mediated immunity in vivo. Next we determined the structure of OMCP bound to human NKG2D. Despite a structure similar to that of host NKG2D ligands, OMCP uses a drastically different orientation for NKG2D binding. The re-orientation of OMCP is associated with dramatically higher affinity for human NKG2D and the targeted interface is highly conserved in mammalian NKG2Ds, increasing the zoonotic potential of cowpox virus. We also show that cell surface presented OMCP can trigger NKG2D effector functions equivalently to host NKG2D ligands, demonstrating that NKG2D-mediated signaling requires clustering but is insensitive to binding orientation. Thus, in contrast to TCR/MHC interactions, the docking topology of NKG2D with its ligands does not appear to regulate its activation.

Structural basis of cowpox evasion of NKG2D immunosurveillance