Crystal Structure of Cell Adhesion Molecule Nectin-2/CD112 and Its Binding to Immune Receptor DNAM-1/CD226

Abstract

The nectin and nectin-like molecule (Necl) family includes important cell adhesion molecules (CAMs) characterized by their Ig-like nature. Such CAMs regulate a broad spectrum of cell–cell interactions, including the interaction between NK cells and cytotoxic T lymphocytes (CTLs) and their target cells. CAM members nectin-2 (CD112) and Necl-5 (CD155) are believed to form homodimers (for nectin-2) or heterodimers in their functions for cell adhesion, as well as to interact with immune costimulatory receptor DNAX accessory molecule 1 (DNAM-1) (CD226) to regulate functions of both NK and CTL cells. However, the structural basis of the interactive mode of DNAM-1 with nectin-2 or Necl-5 is not yet understood. In this study, a soluble nectin-2 Ig-like V-set domain (nectin-2v) was successfully prepared and demonstrated to bind to both soluble ectodomain and cell surface-expressed full-length DNAM-1. The 1.85-Å crystal structure of nectin-2v displays a perpendicular homodimer arrangement, revealing the homodimer characteristics of the nectin and Necls. Further mutational analysis indicated that disruption of the homodimeric interface of nectin-2v led to a failure of the homodimer formation, as confirmed by crystal structure and biochemical properties of the mutant protein of nectin-2v. Interestingly, the monomer mutant also loses DNAM-1 binding, as evidenced by cell staining with tetramers and surface plasmon resonance assays. The data indicate that interaction with DNAM-1 requires either the homodimerization or engagement of the homodimeric interface of nectin-2v. These results have implications for immune intervention of tumors or autoimmune diseases in the DNAM-1/nectin-2–dependent pathway.

Structure of Nectin-2 reveals determinants of homophilic and heterophilic interactions that control cell–cell adhesion

“…However, it remains a possibility that the cis-interaction between IgV domains proposed on the basis of the nectin-1 structure could act as a regulatory mechanism to control the extent of transinteractions. In summary, on the basis of our structural and biochemical data, as well as previously reported data (16)(17)(18)(19), we believe that our nectin-2 dimer represents the biologically relevant homophilic transinteraction shown in Fig. 5.…”

Crystal structures for short-chain pentraxin from zebrafish demonstrate a cyclic trimer with new recognition and effector faces

“…SeMet Dare-PTX was prepared using a reported method (Blackburn et al, 1999;Chen et al, 2010;Garboczi et al, 1996). The purification procedures for native and SeMet Dare-PTX were performed in accordance with the method described in our previous reports (Chen et al, 2010;Liu et al, 2012). The Dare-PTX inclusion bodies were extracted, and Dare-PTX was renatured in refolding buffer.…”

“…Structure determination was performed using the singlewavelength anomalous diffraction (SAD) method, as previously reported (Liu et al, 2012). Briefly, the expected heavy atoms were determined by SHELXD (Schneider and Sheldrick, 2002), and then initial phases were determined using Phaser (McCoy et al, 2007).…”

B Virus (Macacine Herpesvirus 1) Divergence: Variations in Glycoprotein D from Clinical and Laboratory Isolates Diversify Virus Entry Strategies

“…Nectin-2 is also a ligand of the activating natural killer (NK) cell receptor DNAX accessory molecule 1 (DNAM-1) (43,(89)(90)(91)(92). Work in PRV and HSV-2 showed that following cellular transfection with gD or viral infection, DNAM-1 was downregulated to prevent NK cell-mediated cytolysis of infected cells (93).…”