Cortical Actin Tension, Elastic Modulus and Cytosolic Pressure in Fibroblasts Determined using Atomic Force Microscopy

Abstract

The cortical actin cytoskeleton lies just beneath the cell membrane to define cell shape and mechanical properties and thus plays a key role in cell biology processes such as migration and morphogenesis. The organization of actin filaments and actomyosin contractility are known to contribute to modifying the mechanical properties of the cortex. However, recent work report how these properties contribute to cortex tension and intracellular pressure. Here we propose a new method for using an atomic force microscope to determine actin cortex mechanical properties of non-adherent human foreskin fibroblasts including the cortex tension and intracellular pressure, but additionally, the cortex elastic modulus which has not been measured before. First, we validated the method by measuring the surface tension of water-in-oil microdrops deposited on a glass surface. We extracted an average tension of T∼20.2 nN/µm, which agrees with macroscopic experimental methods. We then proceeded to measure cortical actin mechanical properties in non-adherent fibroblasts, and compare this to the properties after inducing two perturbations (i) adding blebbistatin which inhibits myosin II molecular motor activity, and (ii) adding CK-666 which inhibits Arp2/3-mediated actin branching. Our results show that perturbing the actin cortex had significant changes in each of the cortical mechanical properties: blebbistatin reduced them by ∼50%, while CK-666 increased them by ∼2-fold. These results validate our novel method for determining the quantitative mechanics of the actin cortex in eukaryotic cells.