Mechanisms of activation and desensitization of full-length glycine receptor in lipid nanodiscs.
Journal
Nature communications
ISSN: 2041-1723
Titre abrégé: Nat Commun
Pays: England
ID NLM: 101528555
Informations de publication
Date de publication:
27 07 2020
27 07 2020
Historique:
received:
07
02
2020
accepted:
25
06
2020
entrez:
29
7
2020
pubmed:
29
7
2020
medline:
9
9
2020
Statut:
epublish
Résumé
Glycinergic synapses play a central role in motor control and pain processing in the central nervous system. Glycine receptors (GlyRs) are key players in mediating fast inhibitory neurotransmission at these synapses. While previous high-resolution structures have provided insights into the molecular architecture of GlyR, several mechanistic questions pertaining to channel function are still unanswered. Here, we present Cryo-EM structures of the full-length GlyR protein complex reconstituted into lipid nanodiscs that are captured in the unliganded (closed), glycine-bound (open and desensitized), and allosteric modulator-bound conformations. A comparison of these states reveals global conformational changes underlying GlyR channel gating and modulation. The functional state assignments were validated by molecular dynamics simulations, and the observed permeation events are in agreement with the anion selectivity and conductance of GlyR. These studies provide the structural basis for gating, ion selectivity, and single-channel conductance properties of GlyR in a lipid environment.
Identifiants
pubmed: 32719334
doi: 10.1038/s41467-020-17364-5
pii: 10.1038/s41467-020-17364-5
pmc: PMC7385131
doi:
Substances chimiques
Lipids
0
Neurotransmitter Agents
0
Receptors, Glycine
0
Zebrafish Proteins
0
glra1 protein, zebrafish
0
Glycine
TE7660XO1C
Types de publication
Journal Article
Research Support, N.I.H., Extramural
Research Support, Non-U.S. Gov't
Langues
eng
Sous-ensembles de citation
IM
Pagination
3752Subventions
Organisme : NIH HHS
ID : S10 OD021600
Pays : United States
Organisme : NIGMS NIH HHS
ID : R35 GM134896
Pays : United States
Organisme : NIGMS NIH HHS
ID : R01 GM108921
Pays : United States
Organisme : NIGMS NIH HHS
ID : R01 GM131216
Pays : United States
Organisme : NIGMS NIH HHS
ID : P41 GM103832
Pays : United States
Références
Harvey, R. J. et al. GlyR alpha3: an essential target for spinal PGE2-mediated inflammatory pain sensitization. Science 304, 884–887 (2004).
pubmed: 15131310
Bode, A. & Lynch, J. W. The impact of human hyperekplexia mutations on glycine receptor structure and function. Mol. Brain 7, 2 (2014).
pubmed: 24405574
pmcid: 3895786
Zeilhofer, H. U., Acuna, M. A., Gingras, J. & Yevenes, G. E. Glycine receptors and glycine transporters: targets for novel analgesics? Cell Mol. Life Sci. 75, 447–465 (2018).
pubmed: 28791431
Langlhofer, G. & Villmann, C. The intracellular loop of the glycine receptor: it’s not all about the size. Front. Mol. Neurosci. 9, 41 (2016).
pubmed: 27330534
pmcid: 4891346
Morales-Perez, C. L., Noviello, C. M. & Hibbs, R. E. X-ray structure of the human alpha4beta2 nicotinic receptor. Nature 538, 411–415 (2016).
pubmed: 27698419
pmcid: 5161573
Polovinkin, L. et al. Conformational transitions of the serotonin 5-HT3 receptor. Nature 563, 275–279 (2018).
pubmed: 30401839
pmcid: 6614044
Basak, S., Gicheru, Y., Rao, S., Sansom, M. S. P. & Chakrapani, S. Cryo-EM reveals two distinct serotonin-bound conformations of full-length 5-HT3A receptor. Nature 563, 270–274 (2018).
pubmed: 30401837
pmcid: 6237196
Du, J., Lu, W., Wu, S., Cheng, Y. & Gouaux, E. Glycine receptor mechanism elucidated by electron cryo-microscopy. Nature https://doi.org/10.1038/nature14853 (2015).
doi: 10.1038/nature14853
pubmed: 26675730
pmcid: 4641525
Laverty, D. et al. Cryo-EM structure of the human alpha1beta3gamma2 GABAA receptor in a lipid bilayer. Nature 565, 516–520 (2019).
pubmed: 30602789
Zhu, S. et al. Structure of a human synaptic GABAA receptor. Nature 559, 67–72 (2018).
pubmed: 29950725
pmcid: 6220708
Huang, X., Chen, H., Michelsen, K., Schneider, S. & Shaffer, P. L. Crystal structure of human glycine receptor-alpha3 bound to antagonist strychnine. Nature 526, 277–280 (2015).
pubmed: 26416729
Gonzalez-Gutierrez, G., Wang, Y., Cymes, G. D., Tajkhorshid, E. & Grosman, C. Chasing the open-state structure of pentameric ligand-gated ion channels. J. Gen. Physiol. 149, 1119–1138 (2017).
pubmed: 29089419
pmcid: 5715906
Cerdan, A. H., Martin, N. E. & Cecchini, M. An ion-permeable state of the glycine receptor captured by molecular dynamics. Structure 26, 1555–1562 e1554 (2018).
pubmed: 30220542
Damgen, M. A. & Biggin, P. C. A refined open state of the glycine receptor obtained via molecular dynamics simulations. Structure 28, 130–139 e132 (2020).
pubmed: 31753620
pmcid: 6945115
Carland, J. E. et al. Characterization of the effects of charged residues in the intracellular loop on ion permeation in alpha1 glycine receptor channels. J. Biol. Chem. 284, 2023–2030 (2009).
pubmed: 19049967
Song, Y. M. & Huang, L. Y. Modulation of glycine receptor chloride channels by cAMP-dependent protein kinase in spinal trigeminal neurons. Nature 348, 242–245 (1990).
pubmed: 2172840
Papke, D. & Grosman, C. The role of intracellular linkers in gating and desensitization of human pentameric ligand-gated ion channels. J. Neurosci. 34, 7238–7252 (2014).
pubmed: 24849357
pmcid: 4028499
Ivica, J. et al. The intracellular domain of homomeric glycine receptors modulates agonist efficacy. J. Biol. Chem. https://doi.org/10.1074/jbc.RA119.012358 (2020).
doi: 10.1074/jbc.RA119.012358
pubmed: 32075914
Ritchie, T. K. et al. Chapter 11—reconstitution of membrane proteins in phospholipid bilayer nanodiscs. Methods Enzymol. 464, 211–231 (2009).
pubmed: 19903557
pmcid: 4196316
Low, S. E., Ito, D. & Hirata, H. Characterization of the zebrafish glycine receptor family reveals insights into glycine receptor structure function and stoichiometry. Front. Mol. Neurosci. 11, 286 (2018).
pubmed: 30323738
pmcid: 6130310
Lynch, J. W., Rajendra, S., Barry, P. H. & Schofield, P. R. Mutations affecting the glycine receptor agonist transduction mechanism convert the competitive antagonist, picrotoxin, into an allosteric potentiator. J. Biol. Chem. 270, 13799–13806 (1995).
pubmed: 7775436
Pribilla, I., Takagi, T., Langosch, D., Bormann, J. & Betz, H. The atypical M2 segment of the beta subunit confers picrotoxinin resistance to inhibitory glycine receptor channels. EMBO J. 11, 4305–4311 (1992).
pubmed: 1385113
pmcid: 557003
Gielen, M., Thomas, P. & Smart, T. G. The desensitization gate of inhibitory Cys-loop receptors. Nat. Commun. 6, 6829 (2015).
pubmed: 25891813
pmcid: 4410641
Griffon, N. et al. Molecular determinants of glycine receptor subunit assembly. EMBO J. 18, 4711–4721 (1999).
pubmed: 10469650
pmcid: 1171544
Unwin, N. Refined structure of the nicotinic acetylcholine receptor at 4A resolution. J. Mol. Biol. 346, 967–989 (2005).
pubmed: 15701510
Mowrey, D. D. et al. Open-channel structures of the human glycine receptor alpha1 full-length transmembrane domain. Structure 21, 1897–1904 (2013).
pubmed: 23994010
Keramidas, A., Moorhouse, A. J., French, C. R., Schofield, P. R. & Barry, P. H. M2 pore mutations convert the glycine receptor channel from being anion- to cation-selective. Biophys. J. 79, 247–259 (2000).
pubmed: 10866951
pmcid: 1300929
Saul, B. et al. Novel GLRA1 missense mutation (P250T) in dominant hyperekplexia defines an intracellular determinant of glycine receptor channel gating. J. Neurosci. 19, 869–877 (1999).
pubmed: 9920650
pmcid: 6782149
Cymes, G. D. & Grosman, C. Tunable pKa values and the basis of opposite charge selectivities in nicotinic-type receptors. Nature 474, 526–530 (2011).
pubmed: 21602825
pmcid: 3121909
Aryal, P., Sansom, M. S. & Tucker, S. J. Hydrophobic gating in ion channels. J. Mol. Biol. 427, 121–130 (2015).
pubmed: 25106689
Rao, S., Klesse, G., Stansfeld, P. J., Tucker, S. J. & Sansom, M. S. P. A heuristic derived from analysis of the ion channel structural proteome permits the rapid identification of hydrophobic gates. Proc. Natl Acad. Sci. USA 116, 13989–13995 (2019).
pubmed: 31235590
Klesse, G., Rao, S., Sansom, M. S. P. & Tucker, S. J. CHAP: a versatile tool for the structural and functional annotation of ion channel pores. J. Mol. Biol. 431, 3353–3365 (2019).
pubmed: 31220459
pmcid: 6699600
Huang, X., Chen, H. & Shaffer, P. L. Crystal structures of human GlyRalpha3 bound to ivermectin. Structure 25, 945–950 e942 (2017).
pubmed: 28479061
Vandenberg, R. J., Handford, C. A. & Schofield, P. R. Distinct agonist- and antagonist-binding sites on the glycine receptor. Neuron 9, 491–496 (1992).
pubmed: 1326295
Hansen, S. B. et al. Structures of aplysia AChBP complexes with nicotinic agonists and antagonists reveal distinctive binding interfaces and conformations. Embo J. 24, 3635–3646 (2005).
pubmed: 16193063
pmcid: 1276711
Althoff, T., Hibbs, R. E., Banerjee, S. & Gouaux, E. X-ray structures of GluCl in apo states reveal a gating mechanism of Cys-loop receptors. Nature 512, 333–337 (2014).
pubmed: 25143115
pmcid: 4255919
Masiulis, S. et al. GABAA receptor signalling mechanisms revealed by structural pharmacology. Nature https://doi.org/10.1038/s41586-018-0832-5 (2019).
doi: 10.1038/s41586-018-0832-5
pubmed: 30733619
pmcid: 6370056
Shan, Q., Haddrill, J. L. & Lynch, J. W. A single beta subunit M2 domain residue controls the picrotoxin sensitivity of alphabeta heteromeric glycine receptor chloride channels. J. Neurochem. 76, 1109–1120 (2001).
pubmed: 11181831
Xu, M., Covey, D. F. & Akabas, M. H. Interaction of picrotoxin with GABAA receptor channel-lining residues probed in cysteine mutants. Biophys. J. 69, 1858–1867 (1995).
pubmed: 8580329
pmcid: 1236419
Etter, A. et al. Picrotoxin blockade of invertebrate glutamate-gated chloride channels: subunit dependence and evidence for binding within the pore. J. Neurochem. 72, 318–326 (1999).
pubmed: 9886084
Grutter, T. et al. Molecular tuning of fast gating in pentameric ligand-gated ion channels. Proc. Natl Acad. Sci. USA 102, 18207–18212 (2005).
pubmed: 16319224
daCosta, C. J., Dey, L., Therien, J. P. & Baenziger, J. E. A distinct mechanism for activating uncoupled nicotinic acetylcholine receptors. Nat. Chem. Biol. 9, 701–707 (2013).
pubmed: 24013278
Lynch, J. W. et al. Identification of intracellular and extracellular domains mediating signal transduction in the inhibitory glycine receptor chloride channel. EMBO J. 16, 110–120 (1997).
pubmed: 9009272
pmcid: 1169618
Mihic, S. J. et al. Sites of alcohol and volatile anaesthetic action on GABA(A) and glycine receptors. Nature 389, 385–389 (1997).
pubmed: 9311780
Lobo, I. A., Mascia, M. P., Trudell, J. R. & Harris, R. A. Channel gating of the glycine receptor changes accessibility to residues implicated in receptor potentiation by alcohols and anesthetics. J. Biol. Chem. 279, 33919–33927 (2004).
pubmed: 15169788
Hibbs, R. E. & Gouaux, E. Principles of activation and permeation in an anion-selective Cys-loop receptor. Nature 474, 54–60 (2011).
pubmed: 21572436
pmcid: 3160419
Legendre, P. Pharmacological evidence for two types of postsynaptic glycinergic receptors on the Mauthner cell of 52-h-old zebrafish larvae. J. Neurophysiol. 77, 2400–2415 (1997).
pubmed: 9163366
Zheng, S. Q. et al. MotionCor2: anisotropic correction of beam-induced motion for improved cryo-electron microscopy. Nat. Methods 14, 331–332 (2017).
pubmed: 5494038
pmcid: 5494038
Rohou, A. & Grigorieff, N. CTFFIND4: fast and accurate defocus estimation from electron micrographs. J. Struct. Biol. 192, 216–221 (2015).
pubmed: 26278980
pmcid: 6760662
Grant, T., Rohou, A. & Grigorieff, N. cisTEM, user-friendly software for single-particle image processing. Elife https://doi.org/10.7554/eLife.35383 (2018).
Fernandez-Leiro, R. & Scheres, S. H. W. A pipeline approach to single-particle processing in RELION. Acta Crystallogr D Struct. Biol. 73, 496–502 (2017).
pubmed: 28580911
pmcid: 5458491
Mindell, J. A. & Grigorieff, N. Accurate determination of local defocus and specimen tilt in electron microscopy. J. Struct. Biol. 142, 334–347 (2003).
pubmed: 12781660
Zivanov, J., Nakane, T. & Scheres, S. H. W. A Bayesian approach to beam-induced motion correction in cryo-EM single-particle analysis. IUCrJ 6, 5–17 (2019).
pubmed: 30713699
pmcid: 6327179
Kucukelbir, A., Sigworth, F. J. & Tagare, H. D. Quantifying the local resolution of cryo-EM density maps. Nat. Methods 11, 63–65 (2014).
pubmed: 24213166
Chen, V. B. MolProbity: all-atom structure validation for macromolecular crystallography. Acta Crystallogr. D 66, 12–21 (2010).
Collaborative Computational Project, N. The CCP4 suite: programs for protein crystallography. Acta Crystallogr. Sect. D Biol. Crystallogr. 50, 760–763 (1994).
Emsley, P. & Cowtan, K. Coot: model-building tools for molecular graphics. Acta Crystallogr. Sect. D Biol. Crystallogr. 60, 2126–2132 (2004).
Adams, P. D. et al. PHENIX: building new software for automated crystallographic structure determination. Acta Crystallogr. Sect. D Biol. Crystallogr. 58, 1948–1954 (2002).
Afonine, P. V. et al. New tools for the analysis and validation of cryo-EM maps and atomic models. Acta Crystallogr D Struct. Biol. 74, 814–840 (2018).
pubmed: 30198894
pmcid: 6130467
Smart, O. S., Neduvelil, J. G., Wang, X., Wallace, B. A. & Sansom, M. S. HOLE: a program for the analysis of the pore dimensions of ion channel structural models. J. Mol. Graph 14, 354–360, 376 (1996).
pubmed: 9195488
Le Guilloux, V., Schmidtke, P. & Tuffery, P. Fpocket: an open source platform for ligand pocket detection. BMC Bioinform. 10, 168 (2009).
Stansfeld, P. J. et al. MemProtMD: automated Insertion of membrane protein structures into explicit lipid membranes. Structure 23, 1350–1361 (2015).
pubmed: 26073602
pmcid: 4509712
Abraham, M. J., Murtola, T., Schulz, R., Pall, S. & Jeremy, C. GROMACS: high performance molecular simulations through multi-level parallelism from laptops to supercomputers. SoftwareX 1, 19–25 (2015).
Abascal, J. L. & Vega, C. A general purpose model for the condensed phases of water: TIP4P/2005. J. Chem. Phys. 123, 234505 (2005).
pubmed: 16392929
Jorgensen, W. L., Maxwell, D. S. & Tirado-Rives, J. Development and testing of the OPLS all-atom force field on conformational energetics and properties of organic liquids. J. Am. Chem. Soc. 118, 11225–11236 (1996).
Hess, B., Bekker, H., Berendsen, H. J. C. & Fraaije, J. G. E. M. LINCS: a linear constraint solver for molecular simulations. J. Comput. Chem. 18, 1463–1472 (1997).
Darden, T., York, D. & Pedersen, L. Particle mesh Ewald: an N·log(N) method for Ewald sums in large systems. J. Chem. Phys. 98, 10089–10092 (1993).
Bussi, G., Donadio, D. & Parrinello, M. Canonical sampling through velocity rescaling. J. Chem. Phys. 126, 14101 (2007).
Parrinello, M. Rahman, A. Polymorphic transitions in single crystals: a new molecular dynamics method. J. Appl. Phys. 52, 7182–7190 (1981).
Wallace, A. C., Laskowski, R. A. & Thornton, J. M. LIGPLOT: a program to generate schematic diagrams of protein-ligand interactions. Protein Eng. 8, 127–134 (1995).
pubmed: 7630882