Orthogonal Peptide-Templated Labeling Elucidates Lateral ET A R/ET B R Proximity and Reveals Altered Downstream Signaling

• Published in: Chembiochem : a European journal of chemical biology 2022-03, Vol.23 (6), p.e202100340
• Main Authors: Wolf, Philipp, Mohr, Alexander, Gavins, Georgina, Behr, Victoria, Mörl, Karin, Seitz, Oliver, Beck-Sickinger, Annette G
• Format: Article
• Language: English
• Subjects: GTP-Binding Proteins - metabolism; Peptides - metabolism; Receptor, Endothelin A - metabolism; Receptor, Endothelin B - metabolism; Signal Transduction - physiology
• ISSN: 1439-4227; 1439-7633
• DOI: 10.1002/cbic.202100340

Fine-tuning of G protein-coupled receptor (GPCR) signaling is important to maintain cellular homeostasis. Recent studies demonstrated that lateral GPCR interactions in the cell membrane can impact signaling profiles. Here, we report on a one-step labeling method of multiple membrane-embedded GPCRs. Based on short peptide tags, complementary probes transfer the cargo (e. g. a fluorescent dye) by an acyl transfer reaction with high spatial and temporal resolution within 5 min. We applied this approach to four receptors of the cardiovascular system: the endothelin receptor A and B (ET R and ET R), angiotensin II receptor type 1, and apelin. Wild type-like G protein activation after N-terminal modification was demonstrated for all receptor species. Using FRET-competent dyes, a constitutive proximity between hetero-receptors was limited to ET R/ET R. Further, we demonstrate, that ET R expression regulates the signaling of co-expressed ET R. Our orthogonal peptide-templated labeling of different GPCRs provides novel insight into the regulation of GPCR signaling.