Unbreakable DNA tension probes show that cell adhesion receptors detect the molecular force-extension curve of their ligands

Abstract

Integrin receptors transduce the mechanical properties of the extracellular matrix. Past studies using DNA probes showed that integrins sense the magnitude of ligand forces with pN resolution. An open question is whether integrin receptors also sense the force-extension trajectory of their ligands. The challenge in addressing this question pertains to the lack of molecular probes that can control force-extension trajectories independently of force magnitude. To address this limitation, we synthesized two reversible DNA probes that fold with identical self-complementary domains but with different topologies. Thus, these probes unfold at the same steady-state force magnitude but following different kinetic pathways to reach the fully extended ssDNA state. Hairpin-like probes unzip with a low barrier of 14 pN while the pseudo-knot-like probes shear at 59 pN. Confirming that we had created probes with different barriers of unfolding, we quantified platelet integrin forces and measured 50-fold more tension signal with the unzipping probes over the shearing probes. In contrast, fibroblasts opened both probes to similar levels indicating more static forces. Surprisingly, fibroblast mechanotransduction markers, such as YAP levels, fibronectin production, actin organization, and integrin activation were significantly elevated on unzipping probes. This demonstrates that integrin receptors within focal adhesions sense the molecular force-extension profile of their ligands and not only the magnitude of equilibrium mechanical resistance.

Develop Tandem Tension Sensor to Gauge Integrin-Transmitted Molecular Forces

“…We speculate that talin and vinculin recruitment may occur concomitantly in wild-type cells. This finding aligns with a recent publication, demonstrating the concurrent emergence of vinculin-GFP signals and integrin tension in cells …”

“…This finding aligns with a recent publication, demonstrating the concurrent emergence of vinculin-GFP signals and integrin tension in cells. 37 …”

The T-cell niche tunes immune function through modulation of the cytoskeleton and TCR-antigen forces

“…TCO-glass surfaces were prepared as described previously 91 . Coverslips were prepared identical to the AuNP coverslips through the APTES step.100 μL of 4 mg/mL TCO-NHS in DMSO was added to each amine-functionalized coverslip and another coverslip was placed on top to create a “sandwich.” Coverslips were incubated for at least 12h or up to 1 week at RT.…”

“…Labeling DNA anchor strand with tetrazine using a copper click reaction Tetrazine labeling of oligonucleotodies was done as described previously 91 . Oligonucleotides were custom synthesized by IDT to include a BHQ2 quencher and a terminal alkyne for azidetetrazine coupling.…”