Journal Article

Allosteric modulation of GPCR-induced β-arrestin trafficking and signaling by a synthetic intrabody

Hébert, Terence E., Baruah, Minakshi, Laporte, Stéphane A., Maharana, Jagannath, Chaturvedi, Madhu, Pandey, Shubhi, Stepniewski, Tomasz Maciej, Devost, Dominic, Banerjee, Ramanuj, Selent, Jana, Sarma, Parishmita, Inoue, Asuka, Baidya, Mithu, Dwivedi-Agnihotri, Hemlata, Ranjan, Ashutosh, Namkung, Yoon, Yadav, Manish K., Kawakami, Kouki, Shukla, Arun K., Panigrahi, Bhanupriya

2022

Overview

Agonist-induced phosphorylation of G protein-coupled receptors (GPCRs) is a primary determinant of β-arrestin (βarr) recruitment and trafficking. For several GPCRs such as the vasopressin receptor subtype 2 (V 2 R), agonist-stimulation first drives the translocation of βarrs to the plasma membrane, followed by endosomal trafficking, which is generally considered to be orchestrated by multiple phosphorylation sites. We have previously shown that mutation of a single phosphorylation site in the V 2 R (i.e., V 2 R T360A ) results in near-complete loss of βarr translocation to endosomes despite robust recruitment to the plasma membrane, and compromised ERK1/2 activation. Here, we discover that a synthetic intrabody (Ib30), which selectively recognizes activated βarr1, efficiently rescues the endosomal trafficking of βarr1 and ERK1/2 activation for V 2 R T360A . Molecular dynamics simulations reveal that Ib30 enriches active-like βarr1 conformation with respect to the inter-domain rotation, and cellular assays demonstrate that it also enhances βarr1-β 2 -adaptin interaction. Our data provide an experimental framework to positively modulate the receptor-transducer-effector axis for GPCRs using intrabodies, which can be potentially integrated in the paradigm of GPCR-targeted drug discovery. G protein-coupled receptors (GPCRs) are integral membrane proteins and the largest class of drug targets in the human genome. Here, Baidya et al. show that a synthetic antibody can be used to modulate GPCR trafficking and signaling in live cells.

• Publisher: Nature Publishing Group UK,Nature Publishing Group,Nature Portfolio
• Subject: 631/45/612/194 / 631/80/86/2363 / 82/1 / 82/83 / 9/10 / 96/95 / Adaptin / Agonists / Allosteric properties / Antibodies / Arrestin / beta-Arrestin 1 - genetics / beta-Arrestin 1 - metabolism / beta-Arrestin 2 - metabolism / beta-Arrestins - metabolism / Endosomes / G protein-coupled receptors / Genomes / Humanities and Social Sciences / Intrabodies / Kinases / Membrane proteins / Membrane trafficking / Membranes / Molecular dynamics / multidisciplinary / Mutation / Phosphorylation / Protein structure / Proteins / Receptors / Receptors, G-Protein-Coupled - metabolism / Recruitment / Science / Science (multidisciplinary) / Signal Transduction / Signaling / Therapeutic targets / Trafficking / Translocation / Vasopressin / Vasopressin V2 receptors