Super-resolved visualization of single DNA-based tension sensors in cell adhesion

Thomas Schlichthaerle,Ralf Jungmann,Caroline Lindner

doi:10.1038/s41467-021-22606-1

Abstract

Cell-extracellular matrix sensing plays a crucial role in cellular behavior and leads to the formation of a macromolecular protein complex called the focal adhesion. Despite their importance in cellular decision making, relatively little is known about cell-matrix interactions and the intracellular transduction of an initial ligand-receptor binding event on the single-molecule level. Here, we combine cRGD-ligand-decorated DNA tension sensors with DNA-PAINT super-resolution microscopy to study the mechanical engagement of single integrin receptors and the downstream influence on actin bundling. We uncover that integrin receptor clustering is governed by a non-random organization with complexes spaced at 20–30 nm distances. The DNA-based tension sensor and analysis framework provide powerful tools to study a multitude of receptor-ligand interactions where forces are involved in ligand-receptor binding.

Highlights

Cell-extracellular matrix sensing plays a crucial role in cellular behavior and leads to the formation of a macromolecular protein complex called the focal adhesion
We here developed a DNA-based molecular tension sensor, which carries a sequestered DNA binding site for DNAPAINT28–30 super-resolution microscopy, which is revealed upon mechanical unfolding as a result of binding of the cyclic arginine-glycine-aspartatic motif to an integrin receptor (Fig. 1)
We have developed a DNA-based molecular tension sensor, which carries a sequestered DNA binding site for DNA-PAINT super-resolution microscopy that is revealed upon mechanical engagement of a cRGD motif through binding to an integrin receptor

Summary

Introduction

Cell-extracellular matrix sensing plays a crucial role in cellular behavior and leads to the formation of a macromolecular protein complex called the focal adhesion. While increasing spatial resolution, this approach still integrates forces over several tens of nanometers and falls short of the ultimate goal to interrogate and resolve forces between true single ligand-receptor pairs To address this issue, extracellular protein-based Förster resonance energy transfer (FRET) sensors were developed to measure the mechanical tension of integrins interacting with their extracellular matrix ligands[18,19,20]. Extracellular protein-based Förster resonance energy transfer (FRET) sensors were developed to measure the mechanical tension of integrins interacting with their extracellular matrix ligands[18,19,20] This advance enabled the quantification of mechanical forces on the single-molecule level and map subpopulations bearing different loads within adhesion structures[21,22]. We show how this receptor clustering translates to the force-generating intracellular cytoskeletal architecture by multiplexed visualization of both the mechanically strained force sensors and the cellular actin network

Methods

Results

Conclusion