Molecular mechanisms of cell adhesion

Cell-cell adhesion is fundamental to the development of multi-cellular organisms and the maintenance of their tissue architecture. Cell adhesion is mediated by specific proteins called cell adhesion molecules (CAMs) that bind to each other. In this work, we use biophysical techniques including surface force measurements, single molecule force measurements and molecular dynamics simulations to study the molecular mechanisms by which cell adhesion molecules mediate adhesion.;Classic cadherins are a subtype of the cadherin superfamily of cell adhesion molecules and are the primary mediators of cell-cell adhesion. However, the molecular mechanism by which cadherins mediate adhesion is still not resolved. On the one hand, the second tryptophan residue (W2) at the outermost domain (EC1) has been shown to be critical. On the other hand, multiple domains are essential for generating the full adhesive activity of cadherins. We use surface and single molecule force measurements with Xenopus cleavage stage cadherin (C-cadherin), its W2A mutant and domain deletion mutants to resolve this apparent contradiction. The surface force measurements show that W2A abolishes the N-terminal interaction of cadherins as well as attenuates multiple inner adhesive interactions. The latter were previously shown to require the EC3 domain. The single molecule force measurements also show that W2A C-cadherin can undergo adhesion to yield a single bond that is weaker than both the prominent bonds exhibited by C-cadherin. These combined results suggest that the W2 residue allosterically affects domains as far as EC3 along the cadherin extra-cellular region. This has enormous implications for the modulation of cadherin adhesion and the possible means by which cadherin can transduce signals into the cell.;Neural Cell Adhesion Molecule (NCAM) belongs to the immunoglobulin (Ig) superfamily of CAMs and mediates cell-cell adhesion predominantly in the nervous system. Surface force measurements previously performed to elucidate its mechanism of adhesion could only be interpreted by assuming the presence of a hinge in its extra-cellular region. In this work, the presence of such a hinge was tested using neutron reflectivity measurements of the thickness of oriented monolayers of NCAM and its domain deletion mutants. The thickness of NCAM monolayers obtained is consistent with the presence of a hinge between its Ig5 and FnIII1 domains. The reflectivity measurements with the domain deletion mutants were inconclusive as the reflectivity profiles could not be fit to models satisfactorily. This also suggests the necessity of obtaining a clean monolayer for the purpose of such reflectivity studies.;NCAM forms a complex between its terminal domains Ig1 and Ig2. When NCAM of cell A and cell B connect to each other through complexes Ig12(A)/Ig12(B), the relative mobility of cells A and B and membrane tension exerts a force on the Ig12(A)/Ig12(B) complex. Here we investigate the response of the complex to force, using steered molecular dynamics. Starting from the structure of the complex from the Ig1-Ig2-Ig3 fragment, we first equilibrate the complex in solvent and show that its actual end-to-end length is markedly larger than in the crystal structure. We then show that the Ig12/Ig12 complex can behave as a molecular spring of spring constant ∼0.03 N/m in response to forces of tens of pico-Newton. Such tertiary structure elasticity can be expected to be pervasive considering the large number of multi-modular CAMs. Finally, we rupture the complex using higher forces to identify E16, F19, K98, and L175 as key residues stabilizing the complex.