The lamellipodium is a myosin independent mechanosensor

Abstract

The ability of adherent cells to sense changes in the mechanical properties of their extracellular environments is critical to numerous aspects of their physiology. It has been well documented that cell attachment and spreading are sensitive to substrate stiffness. Here we demonstrate that this behavior is actually biphasic, with a transition that occurs around a Young's modulus of ~7 kPa. Furthermore, we demonstrate that, contrary to established assumptions, this property is independent of myosin II activity. Rather, we find that cell spreading on soft substrates is inhibited due to reduced nascent adhesion formation within the lamellipodium. Cells on soft substrates display normal leading edge protrusion activity, but these protrusions are not stabilized due to impaired adhesion assembly. Enhancing integrin-ECM affinity through addition of Mn 2+ recovers nascent adhesion assembly and cell spreading on soft substrates. Using a computational model to simulate nascent adhesion assembly, we find that biophysical properties of the integrin-ECM bond are optimized to stabilize interactions above a threshold matrix stiffness that is consistent with the experimentally observations. Together these results suggest that myosin II-independent forces in the lamellipodium are responsible for mechanosensation by regulating new adhesion assembly, which in turn, directly controls cell spreading. This myosin II-independent mechanism of substrate stiffness sensing could potentially regulate a number of other stiffness sensitive processes. Significance StatementCell physiology can be regulated by the mechanics of the extracellular environment. Here, we demonstrate that cell spreading is a mechanosensitive process regulated by weak forces generated at the cell periphery and independent of motor activity. We show that stiffness sensing depends on the kinetics of the initial adhesion bonds that are subjected to forces driven by protein polymerization. This work demonstrates how the binding kinetics of adhesion molecules are sensitively tuned to a range of forces that enable mechanosensation.All rights reserved. No reuse allowed without permission.was not peer-reviewed) is the author/funder, who has granted bioRxiv a license to display the preprint in perpetuity.

Cellular contractile forces are nonmechanosensitive

“…The starting point for our investigation is the widespread view that cellular contractile forces are mechanosensitive, i.e., that cells regulate these forces through sensing the rigidity of the ECM. This view is based on previous extensive observations that show that contractile forces depend on ECM rigidity: The larger the rigidity is, the larger cellular contractile forces are (10,23,24). We show that cellular forces are generated through intrinsic nonmechanosensitive contractile displacements.…”

Switching between blebbing and lamellipodia depends on the degree of non-muscle myosin II activity

“…Altogether these reports support a notion that cytoskeleton network machinery such as actin polymerization &NMII activity can change the migratory plasticity. (Oakes et al, 2018) proposed that NMII was not needed for lamellipodia. On the other hand, the involvement of NMII in lamellipodia was very much explained by a well-studied steadystate model system, in which the actin network at the leading edge of migrating cell (LP) is maintained by the balance between polymerization and depolymerization of actin filaments.…”

“…Previous studies established that actomyosin contractility and actin protrusivity at the edge of cells drive blebbing and lamellipodial activities of the plasma membrane, respectively (Bergert et al, 2012). Presence of folded NMIIs monomers and their mini filaments at the lamellipodium and lamella (Shutova et al, 2014), and the debate on whether NMII activity is needed for blebbing and lamellipodia (Chikina et al, 2019b, Oakes et al, 2018 prompted us to re-assess the importance of NMII in such essential membrane protrusive activities of a cell.…”

SLIT2/ROBO1-signaling inhibits macropinocytosis by opposing cortical cytoskeletal remodeling

“…In murine fibroblasts, nonmuscle myosin IIa inhibits spreading by increasing retrograde F-actin flow 86 . However, recent work by Oakes et al suggests that cell spreading is regulated by forces not generated by myosin motors 87 . Our results suggest that RhoA-mediated formin activation is necessary for NSlit2-induced actomyosin changes and that NSlit2-induced myosin II activation is needed for cell rounding but not for spreading.…”