The minimal essential unit for cadherin-mediated intercellular adhesion comprises extracellular domains 1 and 2

N-cadherin comprises five homologous extracellular domains, a transmembrane, and a cytoplasmic domain. The extracellular domains of N-cadherin play important roles in homophilic cell adhesion, but the contribution of each domain to this phenomenon has not been fully evaluated. In particular, the following questions remain unanswered: what is the minimal domain combination that can generate cell adhesion, how is domain organization related to adhesive strength, and does the cytoplasmic domain serve to facilitate extracellular domain interaction? To address these issues, we made serial constructs of the extracellular domains of N-cadherin and produced various cell lines to examine adhesion properties. We show that the first domain of N-cadherin alone on the cell surface fails to generate adhesive activity and that the first two domains of N-cadherin form the minimal essential unit to mediate cell adhesion. Cell lines expressing longer extracellular domains or N-cadherin wild type cells formed larger cellular aggregates than those expressing shorter aggregates. However, adhesion strength, as measured by a shearing test, did not reveal any differences among these aggregative cell lines, suggesting that the first two domains of N-cadherin cells generate the same strength of adhesive activity as longer extracellular domain cells. Furthermore, truncations of the first two domains of N-cadherin are also sufficient to form cis-dimerization at an adhesive junction. Our findings suggest that the extracellular domains of N-cadherin have distinct roles in cell adhesion, i.e. the first two domains are responsible for homophilic adhesion activity, and the other domains promote adhesion efficiency most likely by positioning essential domains relatively far out from the cell surface.