Piconewton forces mediate GAIN domain dissociation of the latrophilin-3 adhesion GPCR

Abstract

Latrophilins are adhesion G-protein coupled receptors (aGPCRs) that control excitatory synapse formation. aGPCRs, including latrophilins, are autoproteolytically cleaved at their GPCR-Autoproteolysis Inducing (GAIN) domain, but the two resulting fragments remain associated on the cell surface. It is thought that force-mediated dissociation of the fragments exposes a peptide that activates G-protein signaling of aGPCRs, but whether GAIN domain dissociation can occur on biologically relevant timescales and at physiological forces is unknown. Here, we show using magnetic tweezers that physiological forces dramatically accelerate the dissociation of the latrophilin-3 GAIN domain. Forces in the 1-10 pN range were sufficient to dissociate the GAIN domain on a seconds-to-minutes timescale, and the GAIN domain fragments reversibly reassociated after dissociation. Thus, mechanical force may be a key driver of latrophilin signaling during synapse formation, suggesting a physiological mechanism by which aGPCRs may mediate mechanically-induced signal transduction.

Step-wise mechanical unfolding and dissociation of the GAIN domains of ADGRG1/GPR56, ADGRL1/Latrophilin-1 and ADGRB3/BAI3: insights into the mechanical activation hypothesis of adhesion G protein-coupled receptors

“…However, until very recently, the required force range and time scale for these processes to occur in any aGPCR remained unknown. A recent study by Zhong et al, which was uploaded to BioRxiv [61] during the preparation of our manuscript, reports on a single-molecule study of the mechanical stability of a different aGPCR (ADGRL3/LPHN3). The authors show that constant forces in the 1-10 pN range are sufficient to dissociate the NTF from CTF within seconds to minutes.…”

“…According to Zhong et al [61], the GAIN domain of auto-cleaved LPHN3 was found to be remarkably stable and remained undissociated even after several days in solution. This is in stark contrast to the observation that when subjected to pN forces, dissociation of LPHN3’s NTF/CTF occurs within seconds to minutes.…”

Unveiling Mechanical Activation: GAIN Domain Unfolding and Dissociation in Adhesion GPCRs

“…24,28 Our study shows that a 1 pN/s loading rate results in conserved GAIN domain partial unfolding over pN forces for all three aGPCRs tested by comparing mechanical properties across subfamilies B, G, and L. Additionally, GPR56 and LPHN1 display extensive GAIN domain partial unfolding before NTF/CTF dissociation. In addition to the investigations of GPR56, LPHN1, and BAI3 presented in this work, the mechanical response of another aGPCR, LPHN3, has been studied by Zhong et al 59 They also observed a similar partial unfolding event that precedes the NTF/CTF dissociation at similar forces. These findings indicate that GAIN domain mechanosensitivity during physiological stretching is likely a conserved property among aGPCRs.…”

Unraveling the Dual-Stretch-Mode Impact on Tension Gauge Tethers’ Mechanical Stability

“…This characteristic holds promise for a variety of applications. In addition to shedding light on the time scales involved in specific mechanotransduction processes at cell−matrix adhesions, 1−8 cell−cell adherence junctions, 9 and mechanical activation of Tcells, 10,31 they may also be applied to study the tension duration of various mechanically sensitive membrane receptors, such as adhesion GPCRs 32,33 and tethered ion channels. 34 Moreover, the potential of dual-stretch-mode TGTs to significantly augment the utility of TGTs as mechanical instruments in DNA nanotechnology and bioengineering is considerable.…”