Abstract
Contact guidance—cell polarization by anisotropic substrate features—is integral to numerous physiological processes; however the complexities of its regulation are only beginning to be discovered. In particular, cells polarize to anisotropic features under non-muscle myosin II (MII) inhibition, despite MII ordinarily being essential for polarized cell migration. Here, we investigate the ability of cells to sense and respond to fiber alignment in the absence of MII activity. We find that contact guidance is determined at the level of individual protrusions, which are individually guided by local fiber orientation, independent of MII. Protrusion stability and persistence are functions of adhesion lifetime, which depends on fiber orientation. Under MII inhibition, adhesion lifetime no longer depends on fiber orientation; however the ability of protrusions to form closely spaced adhesions sequentially without having to skip over gaps in adhesive area, biases protrusion formation along fibers. The co-alignment of multiple protrusions polarizes the entire cell; if the fibers are not aligned, contact guidance of individual protrusions still occurs, but does not produce overall cell polarization. These results describe how aligned features polarize a cell independently of MII and demonstrate how cellular contact guidance is built on the local alignment of adhesions and individual protrusions.
Highlights
Directed cell migration is an important element of numerous physiological processes including cancer metastasis, inflammation, and wound healing, as well as a critical parameter in the design of engineered tissues for regenerative medicine[1,2,3,4]
Cells incubated with a combination of the Rho kinase (ROCK) inhibitor Y-27632 and the myosin II (MII) light chain kinase (MLCK) inhibitor ML-7 exhibited a greater number of protrusions than control cells, but still showed persistent protrusion in the direction of fiber alignment (Fig. 1b, Supplementary Movie 2)
This study demonstrates that contact guidance is regulated locally: individual protrusions are guided by local fiber orientation independently of one another and independent of MII activity
Summary
Directed cell migration is an important element of numerous physiological processes including cancer metastasis, inflammation, and wound healing, as well as a critical parameter in the design of engineered tissues for regenerative medicine[1,2,3,4] Cells determine their migration direction based on one or a combination of extracellular guidance cues, including chemical gradients (chemotaxis), adhesion gradients (haptotaxis), stiffness gradients (durotaxis), cell-cell contacts (collective cell migration; contact inhibition), and anisotropic physical features (contact guidance). Cells on microcontact printed adhesive lines, 2D arrays of aligned fibers, and in 3D fibrillar ECMs organize non-muscle myosin (MII) along their lateral edges, focusing protrusive activity in the direction of substrate feature alignment[22,27,29,30,31]. While MII activity controls the polarity of protrusion initiation by establishing non-protrusive zones[22,28,29], it is not essential for biasing protrusion stability in the direction of feature alignment and not essential for contact guidance
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