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PMID: 18953334 Published · ppublish English Journal Article Research Support, N.I.H., Extramural Research Support, Non-U.S. Gov't

The Janus-faced nature of the C(2)B domain is fundamental for synaptotagmin-1 function.

Nature structural & molecular biology ·Vol. 15 ·No. 11 ·2008-11-00 ·Pages 1160-8

Xue M, Ma C, Craig TK, Rosenmund C, Rizo J

Abstract

Synaptotagmin-1 functions as a Ca2+ sensor in neurotransmitter release and was proposed to act on both the synaptic vesicle and plasma membranes through interactions involving the Ca2+ binding top loops of its C(2) domains and the Ca2+-independent bottom face of the C(2)B domain. However, the functional importance of the C(2)B domain bottom face is unclear. We now show that mutating two conserved arginine residues at the C(2)B domain bottom face practically abolishes synchronous release in hippocampal neurons. Reconstitution experiments reveal that Ca2+-synaptotagmin-1 can dramatically stimulate the rate of SNARE-dependent lipid mixing, and that the two-arginine mutation strongly impairs this activity. These results demonstrate that synaptotagmin-1 function depends crucially on the bottom face of the C(2)B domain and strongly support the notion that synaptotagmin-1 triggers membrane fusion and neurotransmitter release by bringing the vesicle and plasma membranes together, much like the SNAREs do but in a Ca2+-dependent manner.

MeSH Terms
Amino Acid Sequence Animals Arginine/metabolism Calcium/metabolism Cell Membrane/chemistry,metabolism Cells, Cultured Hippocampus/cytology Humans Lipids/chemistry Membrane Fusion/physiology Mice Mice, Knockout Models, Molecular Molecular Sequence Data Mutation Neurons/cytology,physiology Patch-Clamp Techniques Protein Structure, Tertiary Rats SNARE Proteins/metabolism Sequence Alignment Synaptotagmin I/chemistry,genetics,metabolism
Chemicals
Lipids SNARE Proteins Synaptotagmin I Arginine Calcium
Authors & Affiliations
5 authors, click to expand affiliations / ORCID
Xue Mingshan
Department of Neuroscience, Baylor College of Medicine, One Baylor Plaza, Houston, Texas 77030, USA.
Ma Cong
Craig Timothy K
Rosenmund Christian
Rizo Josep
References (52)
52 references, click to expand
  1. Rapid and efficient fusion of phospholipid vesicles by the alpha-helical core of a SNARE complex in the absence of an N-terminal regulatory domain.
    Proc Natl Acad Sci U S A. 1999 Oct 26;96(22):12565-70 PMID: 10535962
  2. Close membrane-membrane proximity induced by Ca(2+)-dependent multivalent binding of synaptotagmin-1 to phospholipids.
    Nat Struct Mol Biol. 2006 Mar;13(3):209-17 PMID: 16491093
  3. Synaptotagmin I functions as a calcium regulator of release probability.
    Nature. 2001 Mar 1;410(6824):41-9 PMID: 11242035
  4. The C2B domain of synaptotagmin I is a Ca2+-binding module.
    Biochemistry. 2001 May 22;40(20):5854-60 PMID: 11352720
  5. Mutations in the second C2 domain of synaptotagmin disrupt synaptic transmission at Drosophila neuromuscular junctions.
    J Comp Neurol. 2001 Jul 16;436(1):4-16 PMID: 11413542
  6. Three-dimensional structure of the synaptotagmin 1 C2B-domain: synaptotagmin 1 as a phospholipid binding machine.
    Neuron. 2001 Dec 20;32(6):1057-69 PMID: 11754837
  7. Germline transmission and tissue-specific expression of transgenes delivered by lentiviral vectors.
    Science. 2002 Feb 1;295(5556):868-72 PMID: 11786607
  8. The C(2)B Ca(2+)-binding motif of synaptotagmin is required for synaptic transmission in vivo.
    Nature. 2002 Jul 18;418(6895):340-4 PMID: 12110842
  9. Synaptotagmins I and IV promote transmitter release independently of Ca(2+) binding in the C(2)A domain.
    Nature. 2002 Jul 18;418(6895):336-40 PMID: 12110845
  10. Structure/function analysis of Ca2+ binding to the C2A domain of synaptotagmin 1.
    J Neurosci. 2002 Oct 1;22(19):8438-46 PMID: 12351718
  11. The synaptotagmin C2A domain is part of the calcium sensor controlling fast synaptic transmission.
    Neuron. 2003 Jul 17;39(2):299-308 PMID: 12873386
  12. Reconstitution of Ca2+-regulated membrane fusion by synaptotagmin and SNAREs.
    Science. 2004 Apr 16;304(5669):435-8 PMID: 15044754
  13. The synaptic vesicle cycle.
    Annu Rev Neurosci. 2004;27:509-47 PMID: 15217342
  14. Dual roles of the C2B domain of synaptotagmin I in synchronizing Ca2+-dependent neurotransmitter release.
    J Neurosci. 2004 Sep 29;24(39):8542-50 PMID: 15456828
  15. Perturbation of membrane structure by uranyl acetate labeling.
    Biophys J. 1981 Mar;33(3):475-7 PMID: 6164411
  16. An experimental test of the discreteness-of-charge effect in positive and negative lipid bilayers.
    Biochemistry. 1986 Dec 16;25(25):8206-14 PMID: 3814579
  17. Synaptotagmin I: a major Ca2+ sensor for transmitter release at a central synapse.
    Cell. 1994 Nov 18;79(4):717-27 PMID: 7954835
  18. Structure of the first C2 domain of synaptotagmin I: a novel Ca2+/phospholipid-binding fold.
    Cell. 1995 Mar 24;80(6):929-38 PMID: 7697723
  19. Definition of the readily releasable pool of vesicles at hippocampal synapses.
    Neuron. 1996 Jun;16(6):1197-207 PMID: 8663996
  20. Bipartite Ca2+-binding motif in C2 domains of synaptotagmin and protein kinase C.
    Science. 1996 Jul 12;273(5272):248-51 PMID: 8662510
  21. Synaptotagmin-syntaxin interaction: the C2 domain as a Ca2+-dependent electrostatic switch.
    Neuron. 1997 Jan;18(1):133-42 PMID: 9010211
  22. SNAREpins: minimal machinery for membrane fusion.
    Cell. 1998 Mar 20;92(6):759-72 PMID: 9529252
  23. Direct interaction of a Ca2+-binding loop of synaptotagmin with lipid bilayers.
    J Biol Chem. 1998 May 29;273(22):13995-4001 PMID: 9593749
  24. Ca2+ binding to synaptotagmin: how many Ca2+ ions bind to the tip of a C2-domain?
    EMBO J. 1998 Jul 15;17(14):3921-30 PMID: 9670009
  25. Mechanism of phospholipid binding by the C2A-domain of synaptotagmin I.
    Biochemistry. 1998 Sep 8;37(36):12395-403 PMID: 9730811
  26. Solution structures of the Ca2+-free and Ca2+-bound C2A domain of synaptotagmin I: does Ca2+ induce a conformational change?
    Biochemistry. 1998 Nov 17;37(46):16106-15 PMID: 9819203
  27. Single-molecule studies of synaptotagmin and complexin binding to the SNARE complex.
    Biophys J. 2005 Jul;89(1):690-702 PMID: 15821166
  28. Synaptotagmin isoforms couple distinct ranges of Ca2+, Ba2+, and Sr2+ concentration to SNARE-mediated membrane fusion.
    Mol Biol Cell. 2005 Oct;16(10):4755-64 PMID: 16093350
  29. Augmenting neurotransmitter release by enhancing the apparent Ca2+ affinity of synaptotagmin 1.
    Proc Natl Acad Sci U S A. 2005 Dec 20;102(51):18664-9 PMID: 16352718
  30. Ca(2+)-synaptotagmin directly regulates t-SNARE function during reconstituted membrane fusion.
    Nat Struct Mol Biol. 2006 Apr;13(4):323-30 PMID: 16565726
  31. Synaptotagmin: fusogenic role for calcium sensor?
    Nat Struct Mol Biol. 2006 Apr;13(4):301-3 PMID: 16715046
  32. Phosphatidylinositol phosphates as co-activators of Ca2+ binding to C2 domains of synaptotagmin 1.
    J Biol Chem. 2006 Jun 9;281(23):15845-52 PMID: 16595652
  33. Rabphilin regulates SNARE-dependent re-priming of synaptic vesicles for fusion.
    EMBO J. 2006 Jun 21;25(12):2856-66 PMID: 16763567
  34. Unraveling the mechanisms of synaptotagmin and SNARE function in neurotransmitter release.
    Trends Cell Biol. 2006 Jul;16(7):339-50 PMID: 16698267
  35. Hemifusion arrest by complexin is relieved by Ca2+-synaptotagmin I.
    Nat Struct Mol Biol. 2006 Aug;13(8):748-50 PMID: 16845390
  36. A clamping mechanism involved in SNARE-dependent exocytosis.
    Science. 2006 Aug 4;313(5787):676-80 PMID: 16794037
  37. N- to C-terminal SNARE complex assembly promotes rapid membrane fusion.
    Science. 2006 Aug 4;313(5787):673-6 PMID: 16888141
  38. Position of synaptotagmin I at the membrane interface: cooperative interactions of tandem C2 domains.
    Biochemistry. 2006 Aug 15;45(32):9668-74 PMID: 16893168
  39. SNAREs--engines for membrane fusion.
    Nat Rev Mol Cell Biol. 2006 Sep;7(9):631-43 PMID: 16912714
  40. A complexin/synaptotagmin 1 switch controls fast synaptic vesicle exocytosis.
    Cell. 2006 Sep 22;126(6):1175-87 PMID: 16990140
  41. A quaternary SNARE-synaptotagmin-Ca2+-phospholipid complex in neurotransmitter release.
    J Mol Biol. 2007 Mar 30;367(3):848-63 PMID: 17320903
  42. How synaptotagmin promotes membrane fusion.
    Science. 2007 May 25;316(5828):1205-8 PMID: 17478680
  43. A complexin fusion clamp regulates spontaneous neurotransmitter release and synaptic growth.
    Nat Neurosci. 2007 Oct;10(10):1235-7 PMID: 17873870
  44. Distinct domains of complexin I differentially regulate neurotransmitter release.
    Nat Struct Mol Biol. 2007 Oct;14(10):949-58 PMID: 17828276
  45. Synaptotagmin activates membrane fusion through a Ca2+-dependent trans interaction with phospholipids.
    Nat Struct Mol Biol. 2007 Oct;14(10):904-11 PMID: 17891149
  46. How does synaptotagmin trigger neurotransmitter release?
    Annu Rev Biochem. 2008;77:615-41 PMID: 18275379
  47. Synaptic vesicle fusion.
    Nat Struct Mol Biol. 2008 Jul;15(7):665-74 PMID: 18618940
  48. Synaptotagmin arrests the SNARE complex before triggering fast, efficient membrane fusion in response to Ca2+.
    Nat Struct Mol Biol. 2008 Aug;15(8):827-35 PMID: 18622390
  49. Conserved prefusion protein assembly in regulated exocytosis.
    Mol Biol Cell. 2006 Jan;17(1):283-94 PMID: 16267273
  50. Structure and function of SNARE and SNARE-interacting proteins.
    Q Rev Biophys. 2005 Feb;38(1):1-47 PMID: 16336742
  51. SNARE-mediated lipid mixing depends on the physical state of the vesicles.
    Biophys J. 2006 Mar 15;90(6):2062-74 PMID: 16361343
  52. Structure of the Janus-faced C2B domain of rabphilin.
    Nat Cell Biol. 1999 Jun;1(2):106-12 PMID: 10559882
Article Info
Journal
Nature structural & molecular biology
Abbr.
Nat Struct Mol Biol
ISSN
1545-9985
DOI
10.1038/nsmb.1508
Published
2008-11-00
Epub
2008-00-26
Pages
1160-8
Language
English
Region
United States
NLM ID
101186374
PMCID
PMC2587052
Subset
IM
Grants
NINDS NIH HHS · R01 NS040944-08 · United States
NINDS NIH HHS · R01 NS050655-05 · United States
NINDS NIH HHS · NS40944 · United States
NIGMS NIH HHS · GM65364 · United States
NINDS NIH HHS · R01 NS040944 · United States
NIGMS NIH HHS · R01 GM065364 · United States
NINDS NIH HHS · NS50655 · United States
NINDS NIH HHS · R01 NS050655 · United States
External Links
PubMed source DOI: 10.1038/nsmb.1508 PMC Full Text View journal details
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