DNA Nanotechnology to Map and Manipulate Adhesion Forces at Fluid Interfaces

Cells transmit piconewton (pN) receptor forces to ligands in the extracellular matrix (ECM) and on the surface of adjacent cells. These forces regulate functions ranging from adhesion to clotting and the immune response. Whereas adhesion mechanics on rigid substrates are well characterized, understanding mechanotransduction at cell-cell junctions remains challenging due to a lack of tools. We develop and apply new classes of DNA-based force probes to map and manipulate receptor forces on supported lipid bilayers (SLBs), planar membranes that mimic an adjacent cell. We use these probes to elucidate force balance in podosomes, which are multipurpose protrusive structures that form at cell-cell and cell-ECM interfaces. Podosomes have a core-ring architecture, and previous works demonstrated that the podosome’s actin core generates nanonewton protrusive forces. However, the podosome’s contractile landscape remained poorly understood. In Aim 1 (Chapter 3), we develop and apply Molecular Tension- Fluorescence Lifetime Imaging Microscopy to map integrin receptor forces and clustering on SLBs. We demonstrate that integrin receptors apply pN tension in podosome rings. We then introduce photocleavable probes to site-specifically perturb adhesion forces and apply rupturable DNA-based force probes to test the role of receptor tension in podosome formation and maintenance. These studies confirm a local mechanical feedback between podosome core protrusion and integrin receptor tension. In Aim 2 (Chapter 4), we evaluate structure and energy transfer across a library of DNA-based tension probes using spectroscopy and microscopy. We then demonstrate the functional implications of probe design on cellular imaging. This work expands our understanding of receptor forces in podosome mechanobiology and contributes new insight and tools for studying juxtacrine receptor interactions.Ph.D