Kinetics of Mechanoresponse in the Mammalian Actin Cytoskeleton

Abstract

Cells have a remarkable ability to sense and respond to their mechanical environment; this ability directs a host of cellular and global processes, such as metastasis, engulfment, blood vessel constriction, and tissue development. However, the direct mechanism by which the dynamic network of actin filaments, myosin motors, and actin crosslinking proteins assemble and interact to exert force on the membrane in response to mechanical perturbations remains unclear. Here, we define the in vivo kinetics of 20 cytoskeletal and signaling proteins in response an applied external stress and identify a number of structural elements that accumulate to regions of the cell under shear or dilational forces. Prominent among these are the three non-muscle myosin II paralogs, the actin bundling protein α-actinin 4, and the orthogonal actin crosslinking protein filamin B. All three myosin paralogs, as well as α-actinin 4, accumulate to the tip region of the aspirated cell, which experiences primarily dilational force. Filamin B, however, accumulates to the tip and the neck region, demonstrating that it responds to both sheared and dilated microenvironments. These differences can be directly related to the actin binding geometries and distinct mechanism of force response for each protein. In support of a proposed model for an increased actin affinity state under load, a disease-relevant point mutation K255E in α-actinin 4, which abolishes the structural low affinity state, greatly slows its accumulation kinetics to dilational stress. The accumulation of cytoskeletal motors and crosslinking proteins under mechanical load represents a mechanism by which cells can concentrate structural components to reduce strain or retract from a local environment of high stress. Understanding how cortical protein dynamics intrinsically give rise to the self-tuning nature of the cell will offer insight into the nature of mechanically-dependent cell processes.