May 10, 2023 — A team of researchers led by Wendy Gordon, a Cancer Bioengineering Initiative faculty member and associate professor in the Department of Biochemistry, Molecular Biology and Biophysics, developed a new high-throughput biosensor assay, Rupture and Deliver Tension Gauge Tethers (RAD-TGTs), that determines the amount of force a single cell exerts on its surroundings or its mechanotype.
Probes that can be measured through DNA sequencing
The study, “RAD-TGTs: high-throughput measurement of cellular mechanotype via rupture and delivery of DNA tension probes,” was published in Nature Communications.
The team enhanced the throughput of mechanical assays using traditional Tension Gauge Tethers (TGTs), or short, double-stranded lengths of DNA that separate with a specific amount of force. The new test detects a probe the cell internalizes after exerting enough force to rupture its DNA tether. These probes can be measured using high-throughput flow cytometry or DNA sequencing instead of a microscope.
Bringing mechanobiology into the -omics era
The test can be used to assess the mechanotypes of mixed populations of cells, and has important applications in the fields of cell differentiation, migration, and disease progression, including cancer. The assay can also increase the throughput of CRISPR assays aimed at altering cell mechanotype.
“When mixed populations of cells can be distinguished by their mechanotype, it opens up the possibility of sorting cells by mechanotype to identify genes underlying disease-relevant mechanotypes in CRISPR screens or performing -omics analysis of cells with a given mechanotype,” says Prof. Gordon. “We hope this platform will be easy to employ by many labs and will help launch mechanobiology into the -omics era.”
RAD-TGTs are short, double-stranded lengths of DNA that require a specific amount of force to rupture. RAD-TGT anchor DNA strands (blue) are immobilized on a cell-culture plate. Ligand strands (pink) bind to integrins located on the cell surface. If a cell exerts enough force to rupture the bond between the anchor and ligand stands, the ligand strand is internalized by the cell and a probe (Cy5, pink) is detected in the cell through flow cytometry.