Curated Optogenetic Publication Database

Abstract: Membrane receptors play a crucial role in transmitting external signals inside cells. Signal molecule-bound receptors activate multiple downstream pathways, the dynamics of which are modulated by intracellular trafficking. A significant contribution of β-arrestin to intracellular trafficking has been suggested, but the underlying mechanism is poorly understood. Here, we describe a protocol for manipulating β-arrestin-regulated membrane receptor trafficking using photo-induced dimerization of cryptochrome-2 from Arabidopsis thaliana and its binding partner CIBN. Additionally, the protocol guides analytical methods to quantify the changes in localization and modification of membrane receptors during trafficking.

Abstract: Intracellular trafficking of G protein-coupled receptors (GPCRs) controls their localization and degradation, which affects a cell's ability to adapt to extracellular stimuli. Although the perturbation of trafficking induces important diseases, these trafficking mechanisms are poorly understood. Herein, we demonstrate an optogenetic method using an optical dimerizer, cryptochrome (CRY) and its partner protein (CIB), to analyze the trafficking mechanisms of GPCRs and their regulatory proteins. Temporally controlling the interaction between β-arrestin and β2-adrenergic receptor (ADRB2) reveals that the duration of the β-arrestin-ADRB2 interaction determines the trafficking pathway of ADRB2. Remarkably, the phosphorylation of ADRB2 by G protein-coupled receptor kinases is unnecessary to trigger clathrin-mediated endocytosis, and β-arrestin interacting with unphosphorylated ADRB2 fails to activate mitogen-activated protein kinase (MAPK) signaling, in contrast to the ADRB2 agonist isoproterenol. Temporal control of β-arrestin-GPCR interactions will enable the investigation of the unique roles of β-arrestin and the mechanism by which it regulates β-arrestin-specific trafficking pathways of different GPCRs.