Dual-mode intramolecular agonist-dependent mechanoactivation of the adhesion GPCR ADGRG1/GPR56

Mechanical stimuli instruct cardinal cellular decisions pertaining to their fate, proliferation, morphology, and movement. How adhesion G protein-coupled receptors (aGPCRs), a large family of mechanosensors with more than 30 members in humans, transduce mechanical cues into metabotropic commands, has been a matter of debate due to the lack of suitable approaches to analyze receptor activation during mechanotransduction in live cells. Here we use human ADGRG1/GPR56 (G1), an aGPCR with roles in brain development, skeletal muscle, and platelet function, to study the events during aGPCR mechanotransduction. We show that G1 dissociation, taking place at ∼18 pN, occurs at retracting fibers during cell migration over an adhesive substrate, an effect that can be enhanced by cell stretching. Simultaneous live recording of G protein recruitment through total internal reflection fluorescence (TIRF) imaging and pharmacological assays during force transmission onto G1 show graded receptor responses with sub-maximal signaling before receptor dissociation, and maximal activation thereafter. Both modes require the intramolecular agonist/Stachel element of the receptor. Our findings demonstrate that non-dissociative and dissociative aGPCR signaling can emerge from the same aGPCR depending on mechanical stimulus magnitude, demonstrating the capability of aGPCRs for graded receptor responses upon force detection.