Mechanically Induced Catalytic Amplification Reaction for Readout of Receptor‐Mediated Cellular Forces

Abstract

Mechanics play a fundamental role in cell biology, but detecting piconewton forces is challenging due to the lack of accessible and high throughput assays. Herein we report the mechanically-induced catalytic amplification reaction (MCR) for readout of receptor-mediated forces in cells. Mechanically labile DNA duplexes presenting ligands are surface immobilized such that specific receptor forces denature the duplex and thus expose a blocked primer. Amplification of primers is achieved using an isothermal polymerization reaction and quantified by fluorescence readout. As a proof-of-concept, the assay was used to test the activity of a mechano-modulatory drug and integrin adhesion receptor antibodies. To the best of our knowledge, this is the first example of a catalytic reaction triggered in response to molecular piconewton forces. The MCR may transform the field of mechanobiology by providing a new facile tool to detect receptor-specific mechanics with the convenience of the PCR.

Ratiometric Tension Probes for Mapping Receptor Forces and Clustering at Intermembrane Junctions

“…Second generation MTFM probes have further improved the dynamic force range, 27 sensitivity 28,29 and stability. 30–33 We recently developed a gold nanoparticle (AuNP)-based DNA tension probe to directly image the TCR-pMHC forces on immobilized ligands and revealed that T-cells may harness mechanics for enhanced antigen sampling and discrimination. 34 …”

DNA Nanotechnology as an Emerging Tool to Study Mechanotransduction in Living Systems

“…Inspired by the ELISA and PCR assays that revolutionized molecular biology, our lab developed a catalytic amplification strategy to convert mechanical forces into an amplified fluorescent readout, coined mechanically induced catalytic amplification reaction (MCR) . In initial proof‐of‐concept experiments, MCR leveraged the TGT as a peak force transducer.…”

“…C) Bar graph showing the ability of MCR to differentiate integrin antibodies that block different integrin subtypes. Reproduced with permission . Copyright 2016, Wiley‐VCH.…”

DNA Surface Technology: From Gene Sensors to Integrated Systems for Life and Materials Sciences

“…[48] Depending on the detection strategy used, these devices are divided into electrochemical, optical, and mass-sensitive sensors.A sa ne xample,a ptamers can be tagged with aredox-active moiety like methylene blue (MB) or ferrocene (Fc) to transduce conformational changes upon target binding into an electrochemical signal that originates from the labels closer proximity to the electrode ( Figure 3A). [52,55] Indeed, this approach proved extremely sturdy and useful to elucidate fundamental Angewandte Chemie Kurzaufsätze processes in cell biology,such as the determination of pulling forces exerted by integrin or T-cell receptors. In this approach, one reporter serves as the target responsive sensor,w hereas the other one acts as ar eference,w hich decreases signal drift events to less than 2% in complex media such as whole blood.…”

“…[51,54] While the above methods allow for in vitro measurement of biologically relevant forces in the pico-to nanonewton range,i ti sd ifficult to apply these techniques to the study of mechanical forces emanating from living cells,w hich often trigger their fundamental programs like adhesion, migration, and differentiation by mechanical forces.T os implify the quantification of cellular pulling forces,s urfaces bearing fluorescently labeled DNAhairpin structures were developed by Salaitasg roup (Figure 3D). [52,55] Indeed, this approach proved extremely sturdy and useful to elucidate fundamental Angewandte Chemie Kurzaufsätze processes in cell biology,such as the determination of pulling forces exerted by integrin or T-cell receptors.…”