Description
Adhesion G protein-coupled receptors (GPCRs) are proposed to play pivotal adhesive roles in neurons. We investigate mechanotransduction as a possible operating principle for adhesion GPCRs using an optical tweezer-based strategy at two different scales. At the cellular level, we use an optically trapped bead to apply forces directly to many ADGRL3 receptors (a prototypical adhesion GPCR) on the membrane of living HEK cells, while applying forces and deformation velocities comparable to physiological traction and migration velocities, in parallel with confocal imaging of fluorescent mini G proteins (mG12) reporting on receptor signaling activation. At the single-molecule (SM) level on supported lipid bilayers (SLBs), we investigate ADGRL3 interactions with its endogenous transsynaptic ligand FLRT3, both present on opposing fluid SLBs of membrane-coated optically trapped beads. Recent SM studies on functionalized ADGRL3 GAIN domains suggest a force-dependent reversible transition between a compact and an unfolded state, as well as an irreversible transition between an unfolded and a dissociated state. The unfolded and dissociated states are believed to drive receptor signaling activation, yet there is little evidence for this in living cells. Using our SM strategy with SLB-coated beads we examine whether the force per receptor applied in our cell assay would promote the structural transitions reported by others, thereby enabling a link to signaling activity (mG12 recruitment). The data suggest receptor-ligand SM interactions, with a fourfold increase in interaction frequency relative to negative controls, as well as ADGRL3 activation under tensile forces, but not under compression forces. A draft for a force-dependant kinetic model for the mechanical activation of ADGRL3 on live cells is proposed
| Field of study | Biophysics |
|---|---|
| Supervisor | Prof. Pól Martin Bendix and Prof. Signe Mathiasen |