Tethered agonist exposure in intact adhesion/class B2 GPCRs through intrinsic structural flexibility of the GAIN domain

Mol Cell. 2021 Mar 4;81(5):905-921.e5. doi: 10.1016/j.molcel.2020.12.042. Epub 2021 Jan 25.


Adhesion G protein-coupled receptors (aGPCRs)/family B2 GPCRs execute critical tasks during development and the operation of organs, and their genetic lesions are associated with human disorders, including cancers. Exceptional structural aGPCR features are the presence of a tethered agonist (TA) concealed within a GPCR autoproteolysis-inducing (GAIN) domain and their non-covalent heteromeric two-subunit layout. How the TA is poised for activation while maintaining this delicate receptor architecture is central to conflicting signaling paradigms that either involve or exclude aGPCR heterodimer separation. We investigated this matter in five mammalian aGPCR homologs (ADGRB3, ADGRE2, ADGRE5, ADGRG1, and ADGRL1) and demonstrate that intact aGPCR heterodimers exist at the cell surface, that the core TA region becomes unmasked in the cleaved GAIN domain, and that intra-GAIN domain movements regulate the level of tethered agonist exposure, thereby likely controlling aGPCR activity. Collectively, these findings delineate a unifying mechanism for TA-dependent signaling of intact aGPCRs.

Keywords: FRET microscopy; GAIN domain; adhesion GPCR; auto-proteolysis; bioorthogonal click labeling; dSTORM; genetic code expansion; super-resolution microscopy; tethered agonism.

Publication types

  • Research Support, Non-U.S. Gov't

MeSH terms

  • Amino Acid Sequence
  • Animals
  • Antigens, CD / chemistry*
  • Antigens, CD / genetics
  • Antigens, CD / metabolism
  • Binding Sites
  • COS Cells
  • Chlorocebus aethiops
  • Crystallography, X-Ray
  • Gene Expression
  • HEK293 Cells
  • Humans
  • Molecular Dynamics Simulation
  • Nerve Tissue Proteins / chemistry*
  • Nerve Tissue Proteins / genetics
  • Nerve Tissue Proteins / metabolism
  • Peptides / chemistry*
  • Peptides / genetics
  • Peptides / metabolism
  • Protein Binding
  • Protein Conformation, alpha-Helical
  • Protein Conformation, beta-Strand
  • Protein Interaction Domains and Motifs
  • Protein Multimerization
  • Proteolysis
  • Receptors, G-Protein-Coupled / chemistry*
  • Receptors, G-Protein-Coupled / genetics
  • Receptors, G-Protein-Coupled / metabolism
  • Receptors, Peptide / chemistry*
  • Receptors, Peptide / genetics
  • Receptors, Peptide / metabolism
  • Recombinant Proteins / chemistry
  • Recombinant Proteins / genetics
  • Recombinant Proteins / metabolism
  • Sequence Alignment
  • Sequence Homology, Amino Acid
  • Signal Transduction


  • ADGRB3 protein, human
  • ADGRE2 protein, human
  • ADGRE5 protein, human
  • ADGRG1 protein, human
  • ADGRL1 protein, human
  • Antigens, CD
  • Nerve Tissue Proteins
  • Peptides
  • Receptors, G-Protein-Coupled
  • Receptors, Peptide
  • Recombinant Proteins