Abstract
BACKGROUND: Reports of adhesions in cells growing in 3D vary widely-from nonexistent to very large and elongated-and are often in apparent conflict, due largely to our minimal understanding of the underlying mechanisms that determine 3D cell phenotype. We address this problem directly by systematically identifying mechanisms that determine adhesion in 3D matrices and, from our observations, develop principles widely applicable across 2D and 3D substrates. RESULTS: We demonstrate that nonmuscle myosin II activity guides adhesion phenotype in 3D as it does in 2D; however, in contrast to 2D, decreasing bulk matrix stiffness does not necessarily inhibit the formation of elongated adhesions. Even in soft 3D matrices, cells can form large adhesions in areas with appropriate local matrix fiber alignment. We further show that fiber orientation, apart from influencing local stiffness, modulates the available adhesive area and thereby determines adhesion size. CONCLUSIONS: Thus adhesion in 3D is determined by both myosin activity and the immediate microenvironment of each adhesion, as defined by the local matrix architecture. Important parameters include not only the resistance of the fiber to pulling (i.e., stiffness) but also the orientation and diameter of the fiber itself. These principles not only clarify conflicts in the literature and point to adhesion modulating factors other than stiffness, but also have important implications for tissue engineering and studies of tumor cell invasion.