RAD-TGTs: high-throughput measurement of cellular mechanotype via rupture and delivery of DNA tension probes

Matthew R. Pawlak, Adam T. Smiley, Maria Paz Ramirez, Marcus D. Kelly, Ghaidan A. Shamsan, Sarah M. Anderson, Branden A. Smeester, David A. Largaespada, David J. Odde and Wendy R. Gordon ()
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Matthew R. Pawlak: University of Minnesota
Adam T. Smiley: University of Minnesota
Maria Paz Ramirez: University of Minnesota
Marcus D. Kelly: University of Minnesota
Ghaidan A. Shamsan: University of Minnesota
Sarah M. Anderson: University of Minnesota
Branden A. Smeester: University of Minnesota
David A. Largaespada: University of Minnesota
David J. Odde: University of Minnesota
Wendy R. Gordon: University of Minnesota

Nature Communications, 2023, vol. 14, issue 1, 1-11

Abstract: Abstract Mechanical forces drive critical cellular processes that are reflected in mechanical phenotypes, or mechanotypes, of cells and their microenvironment. We present here “Rupture And Deliver” Tension Gauge Tethers (RAD-TGTs) in which flow cytometry is used to record the mechanical history of thousands of cells exerting forces on their surroundings via their propensity to rupture immobilized DNA duplex tension probes. We demonstrate that RAD-TGTs recapitulate prior DNA tension probe studies while also yielding a gain of fluorescence in the force-generating cell that is detectable by flow cytometry. Furthermore, the rupture propensity is altered following disruption of the cytoskeleton using drugs or CRISPR-knockout of mechanosensing proteins. Importantly, RAD-TGTs can differentiate distinct mechanotypes among mixed populations of cells. We also establish oligo rupture and delivery can be measured via DNA sequencing. RAD-TGTs provide a facile and powerful assay to enable high-throughput mechanotype profiling, which could find various applications, for example, in combination with CRISPR screens and -omics analysis.

Date: 2023
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DOI: 10.1038/s41467-023-38157-6

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