UC Berkeley

The E-cadherin cell adhesion protein is important for embryo development and cell-cell junction stability, and furthermore plays a role in tumor suppression. It binds apposing cells by forming trans-dimers, which can additionally be stabilized by cis-interactions. However, given the low affinity of the trans-dimer, questions persist about the mechanisms that drive the assembly of such biologically important junctions. One recent theory describes the cooperation of trans and cis-affinity in stabilizing contacts of freely diffusing E-cadherin. Another theory is instead based on the force dependence of different trans-dimer configurations. Not to be overlooked, any such hypothesis must also consider the influence of actin dynamics in forming stable junctions. A synthetic cell membrane platform was used to address these concerns by creating a controlled environment for observing E-cadherin interactions on the planar surface of a supported lipid bilayer. This allowed for the precise adjustment of physicochemical attributes and direct observation of E-cadherin mediated assembly, which uncovered new aspects of adhesion regulation.

The behavior of cells with E-cadherin has been studied previously using lipid bilayers, but such experiments have lacked even a minimal physical characterization of the membrane surface and direct imaging of cell interactions. In this study, an E-cadherin bilayer platform was characterized with respect to fluidity and density. With the introduction of the epithelial cell line MKN28, cells were observed to interact and transport the E-cadherin, causing enrichment of the protein above initial bilayer surface densities. Immunostaining these cell-bilayer interfaces exposed an architecture similar to those found in vivo, suggesting the formation of mature E-cadherin mediated adhesions.

The current system led to the observation that cells assembled junctions on a low-mobility protein surface much more frequently than with a fluid bilayer (adhesions assembled in 54% of cells vs. 1%). Limiting protein fluidity in other ways, such as using nano-patterned diffusion barriers, non-fluid bilayers, and cross-linking antibodies, only showed enrichment events with 1 - 3% of the cells added. The reason for this unusual behavior comes from the physical response between the surface and the cell's retraction of filopodia during junction formation. Cells on both fluid and low-mobility surfaces exhibited active membrane protrusions, but only on the low-mobility surface was protein enrichment observed underneath retracting filopodia.

This discovery shows that E-cadherin adhesions are formed by an active process, which is dependent on the force and densities resulting from interactions with the low-mobility lipid surface. These lipid bilayer studies address the theories of adhesion assembly, with results suggesting that the decreased diffusion of E-cadherin on the low-mobility lipid surface provides the stability and resistance needed for the formation of robust, force-dependent junctions.