ABSTRACT
G-protein-coupled receptors (GPCRs), also known as seven transmembrane receptors (7TMRs), typically interact with two distinct signal-transducers, i.e., G proteins and ß-arrestins (ßarrs). Interestingly, there are some non-canonical 7TMRs that lack G protein coupling but interact with ßarrs, although an understanding of their transducer coupling preference, downstream signaling, and structural mechanism remains elusive. Here, we characterize two such non-canonical 7TMRs, namely, the decoy D6 receptor (D6R) and the complement C5a receptor subtype 2 (C5aR2), in parallel with their canonical GPCR counterparts. We discover that D6R and C5aR2 efficiently couple to ßarrs, exhibit distinct engagement of GPCR kinases (GRKs), and activate non-canonical downstream signaling pathways. We also observe that ßarrs adopt distinct conformations for D6R and C5aR2, compared to their canonical GPCR counterparts, in response to common natural agonists. Our study establishes D6R and C5aR2 as ßarr-coupled 7TMRs and provides key insights into their regulation and signaling with direct implication for biased agonism.
Subject(s)
Cell Membrane/metabolism , Protein Conformation , Signal Transduction , beta-Arrestins/chemistry , Animals , GTP-Binding Proteins/chemistry , HEK293 Cells , Humans , Mice , Mice, Inbred C57BL , Mice, Knockout , Phosphorylation , Protein Binding , Protein Domains , Protein Structure, Secondary , Protein Transport , Receptor, Anaphylatoxin C5a/metabolismABSTRACT
G-coupled protein receptors (GPCRs) comprise the largest class of druggable targets. Signaling by GPCRs is initiated from subcellular hot spots including the plasma membrane, signalosomes, and endosomes to contribute to vascular inflammation. GPCR-G protein signaling at the plasma membrane causes endothelial barrier disruption and also cross-talks with growth factor receptors to promote proinflammatory signaling. A second surge of GPCR signaling is initiated by cytoplasmic NFκB activation mediated by ß-arrestins and CARMA-BCL10-MALT1 signalosomes. Once internalized, ubiquitinated GPCRs initiate signaling from endosomes via assembly of the transforming growth factor-ß-activated kinase binding protein-1 (TAB1)-TAB2-p38 MAPK complex to promote vascular inflammation. Understanding the complexities of GPCR signaling is critical for development of new strategies to treat vascular inflammation such as that associated with COVID-19.