Nectins are Ca2؉ -independent immunoglobulin (Ig) superfamily proteins that participate in the organization of epithelial and endothelial junctions. Nectins have three Ig-like domains in the extracellular region, and the first one is essential in cell-cell adhesion and plays a central role in the interaction with the envelope glycoprotein D of several viruses. Five Nectin-like molecules (Necl-1 through -5) with similar domain structures to those of Nectins have been identified. Necl-1 is specifically expressed in neural tissue, has Ca 2؉ -independent homophilic and heterophilic cell-cell adhesion activity, and plays an important role in the formation of synapses, axon bundles, and myelinated axons. Here we report the first crystal structure of its N-terminal Ig-like V domain at 2.4 Å , providing insight into trans-cellular recognition mediated by Necl-1. The protein crystallized as a dimer, and the dimeric form was confirmed by size-exclusion chromatography and chemical cross-linking experiments, indicating this V domain is sufficient for homophilic interaction. Mutagenesis work demonstrated that Phe 82 is a key residue for the adhesion activity of Necl-1. A model for homophilic adhesion of Necl-1 at synapses is proposed based on its structure and previous studies.The cell-cell adhesion systems play important roles in neuronal cell migration, axon-bundle formation, target-cell recognition, activity-dependent plasticity of synapses, and formation of complex glial networks, which surround axons and synapses (1). Nectins are a novel family of Ca 2ϩ -independent immunoglobulin-like cell-cell adhesion molecules and comprise four members, termed Nectin-1 to Nectin-4 (2, 3). All Nectins have an extracellular region of three Ig-like domains, a single transmembrane region, and a cytoplasmic tail region (see Fig. 1) (2, 3). Each Nectin forms a homo-cis-dimer, in which the monomers are aligned in a parallel orientation, which is then followed by formation of homo-trans-dimers in which cis-dimers from opposing cell surfaces interact in an anti-parallel orientation, causing cell-cell adhesion (4 -8).The first Ig-like domain is necessary and sufficient for the formation of homo-trans-dimers, whereas the second Ig-like domain is necessary for the formation of homo-cis-dimers (5, 7-9). Furthermore, Nectin-3 can also form a hetero-trans-dimer with either Nectin-1 or -2, whereas Nectin-4 also forms a hetero-trans-dimer with Nectin-1 (4). The heterotrans-dimers are also mediated by the V domains (9). Nectin-1 has been confirmed to serve as a receptor for all ␣-herpes viruses tested so far, including herpes simplex virus (HSV) 3 type 1, HSV type 2, and pseudorabies virus, facilitating their entry and cell-cell spread (10, 11). Furthermore, the V domain of Nectin-1 has been demonstrated to play a central role in the interaction with the envelope glycoprotein D of HSV, pseudorabies virus, and bovine herpes virus-1 (12, 13). Therefore, the V domain plays an essential role not only in homophilic and heterophilic trans-interactions but ...