Mechanical dynamics in live cells and fluorescence-based force/tension sensors

Abstract

Three signaling systems play the fundamental roles in modulating cell activities: chemical, electrical, and mechanical. While the former two are well studied, the mechanical signaling system is still elusive because of the lack of methods to measure structural forces in real time at cellular and subcellular levels. Indeed, almost all biological processes are responsive to modulation by mechanical forces that trigger dispersive downstream electrical and biochemical pathways. Communication among the three systems is essential to make cells and tissues receptive to environmental changes. Cells have evolved many sophisticated mechanisms for the generation, perception and transduction of mechanical forces, including motor proteins and mechanosensors. In this review, we introduce some background information about mechanical dynamics in live cells, including the ubiquitous mechanical activity, various types of mechanical stimuli exerted on cells and the different mechanosensors. We also summarize recent results obtained using genetically encoded FRET (fluorescence resonance energy transfer)-based force/tension sensors; a new technique used to measure mechanical forces in structural proteins. The sensors have been incorporated into many specific structural proteins and have measured the force gradients in real time within live cells, tissues, and animals.

FRET Pairs with Fixed Relative Orientation of Chromophores

“…In general, two kinds of forces can be distinguished based on their origin: Exogenous and endogenous. Exogenous forces are caused, for example, by gravity or fluid shear, whereas endogenous forces originate from osmotic pressure or from proteins (e.g., motor proteins) . The measurement of the impact of such diverse forces on cells in vivo is a major challenge.…”

“…The development/progress of force‐based microscopic techniques, such as atomic force microscopy (AFM), and the availability of genetically encoded (intrinsic) fluorescence probes (GFP‐based probes) have advanced in vivo detection at a single‐cell/molecule level . Based on the tremendous progress made in genetic engineering, the labeling of specific proteins inside the cell or in the membrane, which react to different external force stimuli initiating a downstream signaling cascade or act as an origin of force themselves (e.g., motor proteins), is now being developed . The latter approach is based on the concept of Förster resonance energy transfer (FRET) and transduces the force‐related alteration in the relative positions of fluorescent molecules into an optical signal [in the case of intrinsic probes two GFPs act as a donor (D)–acceptor (A) pair].…”

Talin promotes integrin activation accompanied by generation of tension in talin and an increase in osmotic pressure in neurite outgrowth

“…Neuronal polarization is closely related to intracellular forces, which can be transmitted by the talin rod (29, 68–71). We constructed a talin tension probe based on dipole‐oriented FRET, according to our previous reports (11, 25, 64–66). A pair of eCFP and eYFP (mTurquoise2/sYFP2) was inserted between the talin head and talin rod domains at aa 447.…”

Mechanical stress activates NMDA receptors in the absence of agonists

“…However, fluid drag applied to the ECM will pull on the cytoskeleton increasing its stress3132, and bilayer tension is known to be coupled to cytoskeletal tension3132. While our fluid velocity was precisely controlled, the body stress it produced in the cells depends upon cell geometry and the distribution of stress within the cytoskeleton; that is known to be heterogeneous3334 and this heterogeneity likely accounts for the variability between cells.…”