Schematic illustration of the assembly strategy of the DNA-based Nano-winch. Cropped figure. Credit: A. Mills et al., "A modular spring-loaded actuator for mechanical activation of membrane proteins," Nature Communications, CC BY 4.0
Cellular mechanosensitivity plays a key role in regulating critical biological processes such as blood vessel constriction, pain perception, breathing and many more. Numerous diseases, including cancer, involve dysfunction of cellular mechanosensitivity, but closely studying this mechanism has been mostly limited to the use of expensive technologies that do not allow for several mechanoreceptors to be studied at the same time. Researchers from the Institut National de la Santé et de la Recherche Médicale (Inserm), the Centre national de la recherche scientifique (CNRS) and the Université de Montpellier have now introduced a new way to study cellular mechanosensitivity, in the form of a DNA origami-based “nano-robot” capable of applying fine-tuned forces down to piconewton resolution.
DNA origami refers to the assembling of 3D nanostructures in a pre-defined form using DNA as a construction material. The device created by the researchers, dubbed the “Nano-winch,” is composed of three DNA origami structures, including a central piston-cylinder “body” that applies force through the use of a molecular spring, and two landing “legs” that allow the Nano-winch to stay upright and anchor itself onto the cell membrane. Single-stranded DNA connector loops with a constant stiffness act as entropic springs to exert a defined force that is mechanically translated to the tip of the piston in order to manipulate targeted mechanoreceptors, the authors wrote. The piston tip can position up to three ligand moieties in order to target specific receptors on the cell surface.
The nano-sized molecular actuators are relatively easy and inexpensive to assemble compared to other methods used for studying cellular mechanosensitivity, and the design of the assembly can be tuned to target specific mechanoreceptors and exert specific forces. Additionally, multiple receptors could be activated in parallel using multiple nano-devices, increasing the speed and throughput of mechanosensitivity studies. The Nano-winch was successfully used to autonomously activate integrin receptors on MCF-7 human breast cancer cells, stimulating detectable downstream phosphorylation of focal adhesion kinase (FAK), which demonstrates its applicability to studying cellular mechanical processes, the authors wrote. A modified, remotely activated version of the Nano-winch, which allows for finer extension control and great force exertion, was also used to directly observe the opening of a channel by mechanical force in the protein BtuB in single-channel bilayer experiments. This study was published in Nature Communications.
“The design of a robot enabling the in vitro and in vivo application of piconewton forces meets a growing demand in the scientific community, and represents a major technological advance,” said corresponding author Gaëtan Bellot.
Bellot noted that one of the Nano-winch’s weaknesses is its sensitivity to enzymes that can degrade DNA, and said that the research team’s next step will be to modify the surface of the device so that it is less sensitive to the action of enzymes. Additionally, the team hopes to explore additional modes of activation for the Nano-winch, such as the use of magnetic fields, said Bellot.