Digital and Tunable Genetically Encoded Tension Sensors Based on Engineered Coiled-Coils
Shuhong Liu
Department of Chemistry, Emory University, Atlanta, Georgia, 30322 United States
Search for more papers by this authorJinchan Liu
Department of Molecular Biophysics and Biochemistry, Yale University, New Haven, Connecticut, 06520 United States
Search for more papers by this authorAlexander Foote
Department of Chemistry, Emory University, Atlanta, Georgia, 30322 United States
Search for more papers by this authorHiroaki Ogasawara
Department of Chemistry, Emory University, Atlanta, Georgia, 30322 United States
Search for more papers by this authorSarah Al Abdullatif
Department of Chemistry, Emory University, Atlanta, Georgia, 30322 United States
Search for more papers by this authorProf. Victor S. Batista
Department of Chemistry, Yale University, New Haven, Connecticut, 06520 United States
Search for more papers by this authorCorresponding Author
Prof. Khalid Salaita
Department of Chemistry, Emory University, Atlanta, Georgia, 30322 United States
Wallace H. Coulter Department of Biomedical Engineering, Georgia Institute of Technology and Emory University, Atlanta, Georgia, 30322 United States
Search for more papers by this authorShuhong Liu
Department of Chemistry, Emory University, Atlanta, Georgia, 30322 United States
Search for more papers by this authorJinchan Liu
Department of Molecular Biophysics and Biochemistry, Yale University, New Haven, Connecticut, 06520 United States
Search for more papers by this authorAlexander Foote
Department of Chemistry, Emory University, Atlanta, Georgia, 30322 United States
Search for more papers by this authorHiroaki Ogasawara
Department of Chemistry, Emory University, Atlanta, Georgia, 30322 United States
Search for more papers by this authorSarah Al Abdullatif
Department of Chemistry, Emory University, Atlanta, Georgia, 30322 United States
Search for more papers by this authorProf. Victor S. Batista
Department of Chemistry, Yale University, New Haven, Connecticut, 06520 United States
Search for more papers by this authorCorresponding Author
Prof. Khalid Salaita
Department of Chemistry, Emory University, Atlanta, Georgia, 30322 United States
Wallace H. Coulter Department of Biomedical Engineering, Georgia Institute of Technology and Emory University, Atlanta, Georgia, 30322 United States
Search for more papers by this authorAbstract
Genetically encoded tension sensors (GETSs) allow for quantifying forces experienced by intracellular proteins involved in mechanotransduction. The vast majority of GETSs are comprised of a FRET pair flanking an elastic “spring-like” domain that gradually extends in response to force. Because of ensemble averaging, the FRET signal generated by such analog sensors conceals forces that deviate from the average, and hence it is unknown if a subset of proteins experience greater magnitudes of force. We address this problem by developing digital GETSs comprised of coiled-coils (CCs) with tunable mechanical thresholds. We validate the mechanical response of CC digital probes using thermodynamic stability prediction, AlphaFold2 modeling, steered molecular dynamics simulations, and single-molecule force spectroscopy. Live cell measurements using optimized CC tension sensors that are inserted into vinculin demonstrate that 13 % of this mechanosensor experiences forces >9.9 pN within focal adhesions. This reveals greater magnitudes of vinculin force than had previously been reported and demonstrates that CC tension sensors enable more facile and precise tension measurements in living systems.
Conflict of Interests
The authors declare no conflict of interest.
Open Research
Data Availability Statement
The data that support the findings of this study are available from the corresponding author upon reasonable request.
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