3D images of Neuronal Adhesion Molecule Contactin-2 Reveal an Unanticipated Two-State Architecture

Contactins (CNTNs) are important cell adhesion molecules that mediate neuronal and axoglial contacts, and lesions in these molecules are linked to neuropsychiatric disorders. The extracellular domain of CNTNs contains six Ig domains and four FNIII domains. Crystal structures have shown that Ig1-Ig4 forms a horseshoe-shaped headpiece, in which the N-terminal domains might fold back on the C-terminal domains to form molecular super-U shaped architecture. The arrangement of these domains has been controversial, which may due to the structural dynamics and conformation heterogeneity of the protein. Here, we used a single-molecule 3D imaging method, individual-particle electron tomography (IPET), to study the extracellular domain of CNTN2 that forms monomers with a broad spectrum of conformations, and obtained 60 three-dimensional (3D) reconstructions. In addition to the known horseshoe-shaped headpiece, ~75% headpieces unexpectedly adopt an open (elongated) or a semi-open conformations contributed to our understanding about structural dynamics. The ectodomains formed curve but not double-back in any uniform way, with an averaged molecular dimension of ~255 A. The first-time demonstration of the dynamic nature and conformational preferences of the full-length CNTN2 ectodomain suggest that the headpiece exists in equilibrium in the 9closed9 or 9not-closed9 states. The important architecture may provide a structural platform for protein partners to influence this balance regulating the function of CNTN2. Encoding the ability of this neural adhesion molecule to form both homomers with itself, as well as recruit different protein partner to neuronal and axoglial contact points play the key role in mediating cell-cell interactions.