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Mechanical regulation of cell shape by actomyosin stress fibers

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Abstract

Cell migration is a key process underlying embryogenesis, wound healing, and cancer progression. The actomyosin stress fiber (SF) network enables mechanosensing of the extracellular matrix (ECM) environment by generating forces to probe substrate stiffness, topography, and adhesive properties, which in turn, collectively influence SF organization. Migrating cells actively rearrange three SF subtypes—dorsal SFs, transverse arcs, and ventral SFs (including apical SFs and basal SFs)—to maintain a polarized shape needed for directional movement. There have been several efforts to dissect contributions of specific SF subtypes in generating and maintaining tension, which have produced important new insights into the field’s understanding of SF subtype function. However, they remain indirect measures of SF mechanical properties. Furthermore, many of these studies were conducted on cells cultured on flat, rigid substrates which do not recapitulate many of the salient features of the complex microenvironments found in vivo.

In this dissertation, we seek to understand how cells regulate tension in their SF network during polarization and migration, and how substrate geometry influences SF organization. Using laser nanosurgery, we systematically sever single SFs belonging to each subtype in order to measure their retraction kinetics. We find that SF subtypes are arranged in a mechanically integrated network and that SF subtypes have distinct mechanical properties that are dependent on intrinsic structure, external connections to other SFs, and formation history. Next, we examine the role of cofilin in remodeling the SF network during polarization, the first step in directed cell migration. We find that cofilin remodels SF tension by facilitating the fusion of thin, weakly contractile SFs into thicker, more contractile structures that break tensional symmetry and enable front-back polarization. Finally, we explore the influence of substrate curvature on SF network organization and mechanics. Apical and basal SFs are oriented perpendicular to one another, and bear different amounts of prestress, suggesting that they each have distinct roles in shaping the cell.

In summary, this dissertation systematically examines the mechanical contributions of SF subtypes to determining cell shape and establishing tensional asymmetry in the cell. We also examine the influence of substrate topography and adhesive patterns on cell shape and SF tension, which are important in understanding how cells interact with ECMs that vary in topography and adhesion. These findings enhance our understanding of how cells mechanically organize their SF network to build a contractile, integrated network for migration.

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This item is under embargo until August 5, 2023.