Quantitatively distinct requirements for signaling-competent cell spreading on engineered versus natural adhesion ligands

To design synthetic microenvironments that elicit desired cell behaviors, we must better understand the molecular mechanisms by which cells interact with candidate biomaterials. Using cell lines with distinct alpha(5)beta(1) integrin expression profiles, we demonstrate that this integrin mediates cell spreading on substrata coated with genetically engineered artificial extracellular matrix (aECM) proteins containing the RGD sequence (RGD-containing aECM protein [(a)RGD]) but lacking the PHSRN synergy site. Furthermore, (a)RGD-mediated adhesion stimulates an intracellular focal adhesion kinase (FAK) signal that is indicative of integrin tethering. Although both (a)RGD and the natural ECM protein fibronectin (FN) support alpha(5)beta(1) integrin-mediated cell spreading, quantitative single-cell analysis revealed that (a)RGD-mediated spreading requires ten-fold greater threshold amount of integrin expression than FN-mediated spreading. Our analysis demonstrates that (a)RGD-based substrata mediate both biophysical (cell spreading) and biochemical (FAK signaling) events via the alpha(5)beta(1) integrin, albeit with efficacy quantitatively distinct from that of natural ECM proteins that possess the full spectrum of adhesion and synergy domains. (c) 2004 Elsevier B.V. All rights reserved.