Quantitative and Multiplexed Fluorescence Lifetime Imaging of Intercellular Tensile Forces

Abstract

Mechanical interactions between cells have been shown to play critical roles in regulating cell signaling and communications.H owever,t he precise measurement of intercellular forces is still quite challenging,e specially considering the complex environment at cell-cell junctions.Inthis study,we report af luorescence lifetime-based approach to image and quantify intercellular molecular tensions.U sing this method, tensile forces among multiple ligand-receptor pairs can be measured simultaneously.W efirst validated our approach and developed lifetime measurement-based DNAtension probes to image E-cadherin-mediated tension on epithelial cells.T hese probes were then further applied to quantify the correlations between E-cadherin and N-cadherin tensions during an epithelial-mesenchymal transition process.T he modular design of these probes can potentially be used to study the mechanical features of various physiological and pathological processes.

Fluorescent Flippers: Small‐Molecule Probes to Image Membrane Tension in Living Systems

“…Membrane tension is the counteracting force against mechanical stretching of lipid bilayer membranes [1–16] . FLIM imaging of GUVs and cells was used for the quantitative calibration of fluorescence lifetimes to membrane tension [33] …”

“…Fluorescence imaging generally dominates as a method to elucidate biological processes in living systems, and elegant fluorescent probes have been devised for many purposes. However, the detection of physical forces in living systems is generally limited to strongly interfering physical tools or fluorescent probes integrated into larger bioengineered systems [1–16] . What would be needed to enable much research in the life sciences are small‐molecule fluorescent probes that can simply be added to living systems to report on physical forces.…”

Digging into the biophysical features of cell membranes with lipid-DNA conjugates

“…To further enable the quantitative measurement of intercellular forces, we have lately engineered two ‘next-generation’ lipid-DNA-based mechanical probes (Zhao et al ., 2020 a ; Keshri et al ., 2021). In our first system, a reference fluorophore was added to normalize the cell membrane distribution variations of the DNA probes (Fig.…”

“…Based on a ratiometric fluorescence measurement, the cellular distributions of E-cadherin-mediated forces during epithelial adhesion and migration were quantitatively determined. In our second approach, quantification of intercellular forces was achieved through measuring fluorescence lifetime rather than fluorescence intensity (Keshri et al ., 2021). These fluorescence lifetime signals are independent on the probe concentration, and as a result, force measurement can be achieved without the requirement of a reference fluorophore.…”

Controllable mitochondrial aggregation and fusion by a programmable DNA binder

“…For example, cholesterolfunctionalized DNA nanostructures have been applied for cell surface engineering to realize adaptive cellular interactions [12][13][14] . DNA nanomachines demonstrate precise controllability and robust operation with effective biological outputs in extracellular environment 15,16 . However, a large number of biological processes occurs inside cells, the development of DNA-based manipulation of intracellular network still remains a big challenge.…”