Persistence of fan-shaped keratocytes is a matrix-rigidity-dependent mechanism that requires α5β1 integrin engagement (original) (raw)

Despite the importance of matrix rigidity on cell functions, many aspects of the mechanosensing process in highly migratory cells remain elusive. Here, we studied the migration of highly motile keratocytes on culture substrates with similar biochemical properties and rigidities spanning the range between soft tissues (~kPa) and stiff culture substrates (~GPa). We show that morphology, polarization and persistence of motile keratocytes are regulated by the matrix stiffness over seven orders of magnitude, without changing the cell spreading area. Increasing the matrix rigidity leads to more F-actin in the lamellipodia and to the formation of mature contractile actomyosin fibers that control the cell rear retraction. Keratocytes remain rounded and form nascent adhesions on compliant substrates, whereas large and uniformly distributed focal adhesions are formed on fan-shaped keratocytes migrating on rigid surfaces. By combining poly-L-lysine, fibronectin and vitronectin coatings with selective blocking of α v β 3 or α 5 β 1 integrins, we show that α V β 3 integrins permit the spreading of keratocytes but are not sufficient for polarization and rigidity sensing that require the engagement of α 5 β 1 integrins. Our study demonstrates a matrix rigidity-dependent regulation of the directional persistence in motile keratocytes and refines the role of α v β 3 and α 5 β 1 integrins in the molecular clutch model. Cell migration is an essential process in embryonic development, wound repair, and immune responses resulting from multiple interactions among intracellular organelles 1. The directional movement of motile cells, which is critical to many physiological 2 and pathological 3 situations, is a highly integrated process guided by gradients of environmental cues. These cues may be diffusible or substrate-bounded, as in chemotaxis 4 and haptotaxis 5 respectively, or physical, including topography, 6 electric fields 7 or extracellular-matrix (ECM) rigidity 8. The directed migration of a cell along an ECM-rigidity gradient, known as "durotaxis", was originally observed in fibroblastic cells migrating along a soft-to-stiff interface 9. Although durotaxis is thought to be critical to development of the nervous system 10 , epithelial-to-mesenchymal transition 11 , or cancer metastasis 12 , the cellular machinery used by motile cells to sense matrix rigidity and to migrate towards stiffer zones remains poorly understood. Motile cells are thought to transmit forces to their surroundings by coupling their actin cytoskeleton to the ECM through adhesion sites 13. Low motile cells, in general, exhibit highly organized actin stress fibers and larger spreading areas on stiff substrate, both leading to an accumulation of internal prestress 14,15. Recent attentions to dynamic cellular processes on compliant environment have proposed the motor-clutch hypothesis 16. In this model, cell adhesion molecules act as molecular clutches that create a frictional slippage interface modulating the degree of force transmission from actomyosin fibers. However, the role of the ECM rigidity in the regulation of motile cell properties such as cell shape, polarization, speed and trajectories is poorly characterized. In addition, the identification of the precise molecular interactions involved in the regulation of the molecular clutch on compliant matrices is critical to further understanding the rigidity-sensing mechanism of motile cells.