Different Function of α5β1 and αVβ3 Integrins: Elucidating their Interactions and Spatial Requirements for Adhesion and Migration

The coordination of specific functions of α5β1 and αvβ3 integrins is crucial for cell spreading and migration. For the spatio-temporal transduction of signals, cells interacting with their environment regulate the localization of α5β1 and αvβ3 integrins at adhesive contacts. The understanding of integrin function has progressed because of new techniques and analytical tools; however, the analysis of integrin specific contribution and crosstalk during adhesion remains a challenge.To determine αvβ3 and α5β1 integrin specific functions we used nanoarrays of gold particles presenting immobilized integrin-selective peptidomimetics. For measuring forces generated upon adhesion-mediated by these integrins, we produced pillar arrays decorated with gold particles presenting the peptidomimetics. Finally, to characterize integrin crosstalk, we achieved the controlled sumultaneous presentation of αvβ3 and α5β1 integrin peptidomimetics by using binary nanoarrays of alternating gold and iron oxide particles.We show that integrin binding to the peptidomimetics is highly selective and cells spread on both ligands. However, spreading is faster and cell area and intracellular forces are increased on the α5β1 mimetic. Analysis of integrin clusters indicates that αvβ3 clusters are more pronounced on αvβ3 ligand and are present in cells adhering to α5β1 ligand. Quantitative analysis of adhesion plaques shows that focal adhesion size is increased in cells adhering to αvβ3 ligand. Integrin α5β1clusters are present in cells adhering to α5β1 ligand and colocalize with αvβ3 clusters. There are numerous fibrillar structures in cells adhering to α5β1 ligand, while clusters are mostly localized at the cell margins in cells adhering to αvβ3 ligand.These findings suggest that the activation by ligand binding of αvβ3 integrin doesn't play an active role in promoting intial adhesion and spreading, but it is essential for the formation of stable focal adhesions.