The Role of Myosin-II in Cell Spreading on Soft Hyaluronan-Fibronectin Substrates

Abstract

Studies using polyacrylamide (PAA) substrates show that cells respond to the resistance of the substrate by modifying spread area, focal adhesions (FA) and cytoskeletal morphology. Cells on a soft substrate (E∼300 Pa) generate low traction forces, which do not support cell growth, FA formation and maturation or the assembly of actin stress fibers. We show that soft hyaluronan-fibronectin (HA-Fn) substrates support cell growth and survival similarly to rigid substrates, suggesting that the link between increased tension at the cell-substrate interface and cell growth could be altered by the substrate composition.To address this question we measured epithelial and endothelial cell growth and morphology on soft HA-Fn substrates in the presence of agents that affect the myosin II-dependent contractile mechanism, such as the ROCK inhibitor Y-27632, and using myosin II knockdown cells.Inhibition of myosin II, which decreases traction forces, causes single cells to alter morphology, decrease their adherent area, and disassemble FA and actin stress fibers. In contrast, cells in contact with one another show no significant effect.The spreading, shape and cytoskeletal morphology of single cells on soft HA-Fn substrates seems to be influenced by internal traction forces; however, this link between internal traction forces and cell spreading is eliminated when cells make cell-cell contact. These results suggest that traction forces within single cells on soft HA-Fn can be stimulated by HA as on stiffer PAA-Fn substrates which lead to similar cell spreading area and morphology response. Moreover, it is demonstrated that mechanical signaling is specific not only to substrate stiffness but also to substrate composition and cell state.