Molecular-Scale Tools for Studying Mechanotransduction

Abstract

Mechanical stimuli are known to be potent regulators of the form and function of cells and organisms. Although biological regulation has classically been understood in terms of principles from solution biochemistry, advancements in many fields have led to the development of a suite of techniques that are able to reveal the interplay between mechanical loading and changes in the biochemical properties of proteins in systems ranging from single molecules to living organisms. Here, we review these techniques and highlight the emergence of a new molecular-scale understanding of the mechanisms mediating the detection and response of cells to mechanical stimuli, a process termed mechanotransduction. Specifically, we focus on the role of subcellular adhesion structures in sensing the stiffness of the surrounding environment because this process is pertinent to applications in tissue engineering as well the onset of several mechanosensitive disease states, including cancer.

Improving Quality, Reproducibility, and Usability of FRET‐Based Tension Sensors

“…One tool for studying mechanical load across proteins is a Förster resonance energy transfer (FRET)‐based tension sensor, which enables the measurement of molecular‐scale forces based on changes in emitted light . The first calibrated, genetically encoded FRET‐based tension sensor was developed to study vinculin , a mechanical linker protein that localizes to both adhesion structures, and solidified the importance of vinculin in mechanosensing and migration .…”

“…Whereas FAs bind to extracellular matrix proteins, AJs adhere to receptors on adjacent cell membranes (7). Both structures involve hundreds of distinct protein types and exhibit mechanosensitivity, meaning they sense and respond to changes in mechanical loading (4-6).One tool for studying mechanical load across proteins is a Förster resonance energy transfer (FRET)-based tension sensor, which enables the measurement of molecular-scale forces based on changes in emitted light (4,(8)(9)(10)(11)(12)(13). The first calibrated, genetically encoded FRET-based tension sensor was developed to study vinculin (14), a mechanical linker protein that localizes to both adhesion structures, and solidified the importance of vinculin in mechanosensing and migration (14)(15)(16)(17)(18)(19).…”

Molecular Force Sensors: From Fundamental Concepts toward Applications in Cell Biology

“…Mechanical signals, i.e., forces, are constantly transmitted from the extracellular to the intracellular side and vice versa, allowing cells to probe the physical properties of their surroundings . The perceived mechanical information is translated into biochemical activity and serves as input for intracellular signaling cascades in a process called mechanotransduction . Receptor‐mediated force transduction regulates many cellular processes such as adhesion, migration, proliferation, differentiation as well as tumor formation and progression .…”

“…Focusing on the general mechanism of receptor‐mediated force transduction, a detailed molecular picture is required for understanding the highly sophisticated response of cells to different physical parameters in their environment, such as geometry and material properties. Proteomic studies have allowed for identifying a larger number of proteins involved in these mechanotransduction cascades and it is now widely accepted that many of these proteins undergo force‐induced conformational changes that alter their activity . Little is known, however, about the magnitude of the molecular forces that are required for triggering these conformational changes.…”

“…We then introduce the methods that have been developed for measuring these forces, with a special focus on molecular force sensors (MFSs). Several excellent reviews have appeared recently, focusing mostly on the applications of MFSs in cell biology . Here, we take a molecular point of view with the aim of categorizing currently used MFSs by their working mechanism.…”