Structure function study of L-CAM: Insight into the role of the first extracellular domain in the trans cell-cell interaction

The state of cell-cell adhesion is important to processes such as cell differentiation, tissue function, and oncogenic progression. Cadherin proteins mediate adhesion via calcium dependent intercellular homophilic binding and intracellular linkage to the actin cytoskeleton. To test whether function of the putative binding site depends on the orientation and organization of the highly conserved tripeptide His-Ala-Val of the first domain, we mutated it to Val-Ala-His. To determine how different structural elements of the protein contribute to the adhesive interaction a deletion mutant of the first extracellular domain was constructed. Switching HAV to VAH did not eliminate adhesion, indicating that HAV is not an essential component of the adhesive interface. The HAV mutation may have disrupted the cadherin's ability to bind the unmutated L-CAM, therefore HAV/VAH was tested for its capacity to adhere to unmutated L-CAM. L-CAM adhered strongly with HAV/VAH, refuting the theory that HAV organization is important to the adhesive interface. Deletion of the first extracellular domain eliminated cell-cell adhesion, confirming previous observations that homophilic cadherin adhesion requires Domain I. However, the ΔI mutants were able to interact with the unmutated L-CAM, in contrast to the theory that cadherin adhesion is restricted to the first extracellular domain.