Identification of PVR (CD155) and Nectin-2 (CD112) as Cell Surface Ligands for the Human DNAM-1 (CD226) Activating Molecule

TLDR

Human NK cells express activating receptors and coreceptors that recognize and kill target cells. The study sought to identify cellular ligands for NK activating receptors by immunizing mice with NK‑susceptible target cells. Functional screening of monoclonal antibodies against the immunized target cells was used to identify these ligands. PVR (CD155) and Nectin‑2 (CD112) were identified as DNAM‑1 ligands, and their expression on target cells enhanced NK‑cell lysis, which could be blocked by specific antibodies.

Abstract

Human natural killer (NK) cells express a series of activating receptors and coreceptors that are involved in recognition and killing of target cells. In this study, in an attempt to identify the cellular ligands for such triggering surface molecules, mice were immunized with NK-susceptible target cells. On the basis of a functional screening, four mAbs were selected that induced a partial down-regulation of the NK-mediated cytotoxicity against the immunizing target cells. As revealed by biochemical analysis, three of such mAbs recognized molecules of ∼70 kD. The other mAb reacted with two distinct molecules of ∼65 and 60 kD, respectively. Protein purification followed by tryptic digestion and mass spectra analysis, allowed the identification of the 70 kD and the 65/60 kD molecules as PVR (CD155) and Nectin-2 δ/α (CD112), respectively. PVR-Fc and Nectin-2-Fc soluble hybrid molecules brightly stained COS-7 cells transfected with the DNAM-1 (CD226) construct, thus providing direct evidence that both PVR and Nectin-2 represent specific ligands for the DNAM-1 triggering receptor. Finally, the surface expression of PVR or Nectin-2 in cell transfectants resulted in DNAM-1–dependent enhancement of NK-mediated lysis of these target cells. This lysis was inhibited or even virtually abrogated upon mAb-mediated masking of DNAM-1 (on NK cells) or PVR or Nectin-2 ligands (on cell transfectants).