Abstract 19413: A Genetically Encoded Tension Sensor for Desmoplakin
Introduction: Desmoplakin (DP) is a key component of the cardiac intercalated disk that links components of the cell-cell junction to the intermediate filament cytoskeleton. The formation, regulation, and maintenance of mechanical integrity at these contacts is important for tissue integrity and cell-cell junction stability. Dysfunction of this critical mechanical component leads to arrythmogenic cardiomyopathy, highlighting the importance of these adhesions to cardiac health.
Methods: To better understand the process of establishing and maintaining mechanical integrity at cell-cell junctions we have placed a Förster resonance energy transfer (FRET) based tension sensing module (TSMod) into desmoplakin (DP-TSMod). The tension sensing module converts changes in molecular tension to changes in fluorescence, allowing us to localize and quantify molecular forces across DP-TSMod in living cells. Here we express DP-TSMod in the model epithelial Madin-Darby Canine Kidney (MDCK) cell line.
Results: The DP-TSMod protein is recruited to cell-cell contacts in a punctate manner and co-localizes well with endogenous DP. Preliminary evaluation finds no detectable molecular force in the confluent monolayer, but an increased level of force in a sub-confluent cell colony. Current work focuses on evaluating the mechanical states of DP at cell-cell junctions and evaluating load during remodeling and under mechanical strain.
Conclusions: We have developed and characterized a novel tool that enables the visualization of molecular load across cell-cell intermediate filament junctions. Using this tool we find that the load across these junctions decreases as monolayers reach confluency, suggesting that the mechanical state of this adhesion changes dynamically during growth. Future work will determine how pathogenic variants of DP alter force transmission at cell-cell junctions.
Author Disclosures: A.J. Price: None. A.R. Dunn: None.
- © 2016 by American Heart Association, Inc.