Frizzled BRET sensors based on bioorthogonal labeling of unnatural amino acids reveal WNT-induced dynamics of the cysteine-rich domain.
basic_science · Level V
Where this comes from
- Record sourced from PubMed, PMID 34757789.
- Also identified by DOI 10.1126/sciadv.abj7917 and PMC identifier 8580317.
- Licence recorded as CC BY.
- The licence permits redistribution, so the abstract is shown in full and the full text is available from the publisher.
Abstract
Frizzleds (FZD<sub>1–10</sub>) are G protein–coupled receptors containing an extracellular cysteine-rich domain (CRD) binding Wingless/Int-1 lipoglycoproteins (WNTs). Despite the role of WNT/FZD signaling in health and disease, our understanding of how WNT binding is translated into receptor activation and transmembrane signaling remains limited. Current hypotheses dispute the roles for conformational dynamics. To clarify how WNT binding to FZD translates into receptor dynamics, we devised conformational FZD-CRD biosensors based on bioluminescence resonance energy transfer (BRET). Using FZD with N-terminal nanoluciferase (Nluc) and fluorescently labeled unnatural amino acids in the linker domain and extracellular loop 3, we show that WNT-3A and WNT-5A induce similar CRD conformational rearrangements despite promoting distinct signaling pathways and that CRD dynamics are not required for WNT/β-catenin signaling. Thus, these FZD-CRD biosensors provide insights into binding, activation, and signaling processes in FZDs. The sensor design is broadly applicable to explore ligand-induced dynamics also in other membrane receptors.