Nobles, K. N. et al. Distinct phosphorylation sites on the β2-adrenergic receptor establish a barcode that encodes differential functions of β-arrestin. Sci. Signal 4, ra51 (2011).
Schafer, C. T., Chen, Q., Tesmer, J. J. G. & Handel, T. M. Atypical chemokine receptor 3 “senses” CXC chemokine receptor 4 activation through GPCR kinase phosphorylation. Mol. Pharmacol. 104, 174–186 (2023).
Latorraca, N. R. et al. How GPCR phosphorylation patterns orchestrate arrestin-mediated signaling. Cell 183, 1813–1825.e1818 (2020).
Maharana, J. et al. Structural snapshots uncover a key phosphorylation motif in GPCRs driving β-arrestin activation. Mol. Cell 83, 2091–2107.e2097 (2023).
Dwivedi-Agnihotri, H. et al. Distinct phosphorylation sites in a prototypical GPCR differently orchestrate β-arrestin interaction, trafficking, and signaling. Sci. Adv. https://doi.org/10.1126/sciadv.abb8368 (2020).
Nash, C. A. et al. Differential regulation of β2-adrenoceptor and adenosine A2B receptor signalling by GRK and arrestin proteins in arterial smooth muscle. Cell. Signalling 51, 86–98 (2018).
Morris, G. E. et al. Arrestins 2 and 3 differentially regulate ETA and P2Y2 receptor-mediated cell signaling and migration in arterial smooth muscle. Am. J. Physiol. Cell Physiol. 302, C723–C734 (2012).
Ahn, S., Wei, H., Garrison, T. R. & Lefkowitz, R. J. Reciprocal regulation of angiotensin receptor-activated extracellular signal-regulated kinases by β-arrestins 1 and 2. J. Biol. Chem. 279, 7807–7811 (2004).
Bohn, L. M. et al. Enhanced morphine analgesia in mice lacking β-arrestin 2. Science 286, 2495–2498 (1999).
Sun, D., Ma, J. Z., Payne, T. J. & Li, M. D. β-Arrestins 1 and 2 are associated with nicotine dependence in European American smokers. Mol. Psychiatry 13, 398–406 (2008).
Haider, R. S. et al. β-arrestin1 and 2 exhibit distinct phosphorylation-dependent conformations when coupling to the same GPCR in living cells. Nat. Commun. 13, 5638 (2022).
Lee, Y. et al. Molecular basis of β-arrestin coupling to formoterol-bound β1-adrenoceptor. Nature 583, 862–866 (2020).
Yang, F. et al. Phospho-selective mechanisms of arrestin conformations and functions revealed by unnatural amino acid incorporation and 19F-NMR. Nat. Commun. 6, 8202 (2015).
Murphy, P. M. & Heusinkveld, L. Multisystem multitasking by CXCL12 and its receptors CXCR4 and ACKR3. Cytokine 109, 2–10 (2018).
Hattermann, K. & Mentlein, R. An infernal trio: the chemokine CXCL12 and its receptors CXCR4 and CXCR7 in tumor biology. Ann. Anat. 195, 103–110 (2013).
Rajagopal, S. et al. β-arrestin- but not G protein-mediated signaling by the “decoy” receptor CXCR7. Proc. Natl Acad. Sci. USA 107, 628–632 (2010).
Canals, M. et al. Ubiquitination of CXCR7 controls receptor trafficking. PLoS ONE 7, e34192 (2012).
Levoye, A., Balabanian, K., Baleux, F., Bachelerie, F. & Lagane, B. CXCR7 heterodimerizes with CXCR4 and regulates CXCL12-mediated G protein signaling. Blood 113, 6085–6093 (2009).
Torossian, F. et al. CXCR7 participates in CXCL12-induced CD34+ cell cycling through β-arrestin-dependent Akt activation. Blood 123, 191–202 (2014).
Thelen, M. & Thelen, S. CXCR7, CXCR4 and CXCL12: an eccentric trio? J. Neuroimmunol. 198, 9–13 (2008).
Naumann, U. et al. CXCR7 functions as a scavenger for CXCL12 and CXCL11. PLoS ONE 5, e9175 (2010).
Saaber, F. et al. ACKR3 regulation of neuronal migration requires ACKR3 phosphorylation, but not β-arrestin. Cell Rep. 26, 1473–1488.e1479 (2019).
Luker, K. E. et al. Scavenging of CXCL12 by CXCR7 promotes tumor growth and metastasis of CXCR4-positive breast cancer cells. Oncogene 31, 4750–4758 (2012).
Yen, Y. C. et al. Structures of atypical chemokine receptor 3 reveal the basis for its promiscuity and signaling bias. Sci. Adv. 8, eabn8063 (2022).
Zarca, A. et al. Differential involvement of ACKR3 C-tail in β-arrestin recruitment, trafficking and internalization. Cells https://doi.org/10.3390/cells10030618 (2021).
Gustavsson, M., Dyer, D. P., Zhao, C. & Handel, T. M. Kinetics of CXCL12 binding to atypical chemokine receptor 3 reveal a role for the receptor N terminus in chemokine binding. Sci. Signaling https://doi.org/10.1126/scisignal.aaw3657 (2019).
Hanes, M. S. et al. Dual targeting of the chemokine receptors CXCR4 and ACKR3 with novel engineered chemokines. J. Biol. Chem. 290, 22385–22397 (2015).
Chen, K. et al. Tail engagement of arrestin at the glucagon receptor. Nature 620, 904–910 (2023).
Cao, C. et al. Signaling snapshots of a serotonin receptor activated by the prototypical psychedelic LSD. Neuron https://doi.org/10.1016/j.neuron.2022.08.006 (2022).
Bous, J. et al. Structure of the vasopressin hormone–V2 receptor–β-arrestin1 ternary complex. Sci. Adv. 8, eabo7761 (2022).
Staus, D. P. et al. Structure of the M2 muscarinic receptor–β-arrestin complex in a lipid nanodisc. Nature 579, 297–302 (2020).
Yin, W. et al. A complex structure of arrestin-2 bound to a G protein-coupled receptor. Cell Res. 29, 971–983 (2019).
Kang, Y. et al. Crystal structure of rhodopsin bound to arrestin by femtosecond X-ray laser. Nature 523, 561–567 (2015).
Shukla, A. K. et al. Structure of active β-arrestin-1 bound to a G-protein-coupled receptor phosphopeptide. Nature 497, 137–141 (2013).
Sarma, P. et al. Molecular insights into intrinsic transducer-coupling bias in the CXCR4-CXCR7 system. Nat. Commun. 14, 4808 (2023).
Chen, Q. et al. Structural basis of arrestin-3 activation and signaling. Nat. Commun. 8, 1427 (2017).
Ereno-Orbea, J. et al. Structural basis of enhanced crystallizability induced by a molecular chaperone for antibody antigen-binding fragments. J. Mol. Biol. 430, 322–336 (2018).
Wang, X., Zhu, H., Terashi, G., Taluja, M. & Kihara, D. DiffModeler: large macromolecular structure modeling for cryo-EM maps using a diffusion model. Nat. Methods https://doi.org/10.1038/s41592-024-02479-0 (2024).
Huang, W. et al. Structure of the neurotensin receptor 1 in complex with β-arrestin 1. Nature 579, 303–308 (2020).
Zhou, X. E. et al. Identification of phosphorylation codes for arrestin recruitment by G protein-coupled receptors. Cell 170, 457–469.e413 (2017).
Chen, Q. & Tesmer, J. J. G. G protein-coupled receptor interactions with arrestins and GPCR kinases: the unresolved issue of signal bias. J. Biol. Chem. 298, 102279 (2022).
Min, K. et al. Crystal structure of β-arrestin 2 in complex with CXCR7 phosphopeptide. Structure 28, 1014–1023.e1014 (2020).
Aydin, Y. et al. Structural details of a class B GPCR-arrestin complex revealed by genetically encoded crosslinkers in living cells. Nat. Commun. 14, 1151 (2023).
Drube, J. et al. GPCR kinase knockout cells reveal the impact of individual GRKs on arrestin binding and GPCR regulation. Nat. Commun. 13, 540 (2022).
Zhan, X., Gimenez, L. E., Gurevich, V. V. & Spiller, B. W. Crystal structure of arrestin-3 reveals the basis of the difference in receptor binding between two non-visual subtypes. J. Mol. Biol. 406, 467–478 (2011).
Xiao, K. et al. Functional specialization of β-arrestin interactions revealed by proteomic analysis. Proc. Natl Acad. Sci. USA 104, 12011–12016 (2007).
Parruti, G. et al. Molecular analysis of human beta-arrestin-1: cloning, tissue distribution, and regulation of expression. Identification of two isoforms generated by alternative splicing. J. Biol. Chem. 268, 9753–9761 (1993).
Busillo, J. M. et al. Site-specific phosphorylation of CXCR4 is dynamically regulated by multiple kinases and results in differential modulation of CXCR4 signaling. J. Biol. Chem. 285, 7805–7817 (2010).
Kawakami, K. et al. Heterotrimeric Gq proteins act as a switch for GRK5/6 selectivity underlying β-arrestin transducer bias. Nat. Commun. 13, 487 (2022).
Underwood, O. et al. Key phosphorylation sites for robust β-arrestin2 binding at the MOR revisited. Commun. Biol. 7, 933 (2024).
Beyett, T. S. et al. Characterization of a hyperphosphorylated variant of G protein-coupled receptor kinase 5 expressed in E. coli. Protein Expression Purif. 168, 105547 (2020).
Vishnivetskiy, S. A., Zhan, X., Chen, Q., Iverson, T. M. & Gurevich, V. V. Arrestin expression in E. coli and purification. Curr. Protoc. Pharmacol. 67, 2.11.1–2.11.19 (2014).
Paduch, M. et al. Generating conformation-specific synthetic antibodies to trap proteins in selected functional states. Methods 60, 3–14 (2013).
Miller, K. R. et al. T cell receptor-like recognition of tumor in vivo by synthetic antibody fragment. PLoS ONE 7, e43746 (2012).
Bloch, J. S. et al. Development of a universal nanobody-binding Fab module for fiducial-assisted cryo-EM studies of membrane proteins. Proc. Natl Acad. Sci. USA https://doi.org/10.1073/pnas.2115435118 (2021).
Kimanius, D., Dong, L., Sharov, G., Nakane, T. & Scheres, S. H. W. New tools for automated cryo-EM single-particle analysis in RELION-4.0. Biochem. J. 478, 4169–4185 (2021).
Scheres, S. H. RELION: implementation of a Bayesian approach to cryo-EM structure determination. J. Struct. Biol. 180, 519–530 (2012).
Rohou, A. & Grigorieff, N. CTFFIND4: fast and accurate defocus estimation from electron micrographs. J. Struct. Biol. 192, 216–221 (2015).
Bepler, T. et al. Positive-unlabeled convolutional neural networks for particle picking in cryo-electron micrographs. Nat. Methods 16, 1153–1160 (2019).
Kimanius, D. et al. Data-driven regularization lowers the size barrier of cryo-EM structure determination. Nat. Methods 21, 1216–1221 (2024).
Liu, Y. T., Fan, H., Hu, J. J. & Zhou, Z. H. Overcoming the preferred-orientation problem in cryo-EM with self-supervised deep learning. Nat. Methods 22, 113–123 (2025).
Punjani, A., Rubinstein, J. L., Fleet, D. J. & Brubaker, M. A. CryoSPARC: algorithms for rapid unsupervised cryo-EM structure determination. Nat. Methods 14, 290–296 (2017).
Punjani, A., Zhang, H. & Fleet, D. J. Non-uniform refinement: adaptive regularization improves single-particle cryo-EM reconstruction. Nat. Methods 17, 1214–1221 (2020).
Liebschner, D. et al. Macromolecular structure determination using X-rays, neutrons and electrons: recent developments in Phenix. Acta Crystallogr., D: Struct. Biol. 75, 861–877 (2019).
Emsley, P., Lohkamp, B., Scott, W. G. & Cowtan, K. Features and development of Coot. Acta Crystallogr. D Biol. Crystallogr. 66, 486–501 (2010).
Pettersen, E. F. et al. UCSF ChimeraX: structure visualization for researchers, educators, and developers. Protein Sci. 30, 70–82 (2021).
Grant, B. J., Rodrigues, A. P., ElSawy, K. M., McCammon, J. A. & Caves, L. S. Bio3d: an R package for the comparative analysis of protein structures. Bioinformatics 22, 2695–2696 (2006).
Skjærven, L., Yao, X.-Q., Scarabelli, G. & Grant, B. J. Integrating protein structural dynamics and evolutionary analysis with Bio3D. BMC Bioinf. 15, 399 (2014).
Grant, B. J., Skjaerven, L. & Yao, X. Q. The Bio3D packages for structural bioinformatics. Protein Sci. 30, 20–30 (2021).
Edgar, R. C. MUSCLE: multiple sequence alignment with high accuracy and high throughput. Nucleic Acids Res. 32, 1792–1797 (2004).
Gerstein, M. & Altman, R. B. Average core structures and variability measures for protein families: application to the immunoglobulins. J. Mol. Biol. 251, 161–175 (1995).