Re-examining how complexin inhibits neurotransmitter release: SNARE complex insertion or electrostatic hindrance?

  1. Thorsten Trimbuch
  2. Junjie Xu
  3. David Flaherty
  4. Diana R Tomchick
  5. Josep Rizo  Is a corresponding author
  6. Christian Rosenmund
  1. Charité-Universitätsmedizin Berlin, Germany
  2. University of Texas Southwestern Medical Center, United States

Abstract

Complexins play activating and inhibitory functions in neurotransmitter release. The complexin accessory helix inhibits release and was proposed to insert into SNARE complexes to prevent their full assembly. This model was supported by 'superclamp' and 'poor-clamp' mutations that enhanced or decreased the complexin-I inhibitory activity in cell-cell fusion assays, and by the crystal structure of a superclamp mutant bound to a synaptobrevin-truncated SNARE complex. NMR studies now show that the complexin-I accessory helix does not insert into synaptobrevin-truncated SNARE complexes in solution, and electrophysiological data reveal that superclamp mutants have slightly stimulatory or no effects on neurotransmitter release, whereas a poor-clamp mutant inhibits release. Importantly, increasing or decreasing the negative charge of the complexin-I accessory helix inhibits or stimulates release, respectively. These results suggest a new model whereby the complexin accessory helix inhibits release through electrostatic (and perhaps steric) repulsion enabled by its location between the vesicle and plasma membranes.

Article and author information

Author details

  1. Thorsten Trimbuch

    Charité-Universitätsmedizin Berlin, Berlin, Germany
    Competing interests
    No competing interests declared.
  2. Junjie Xu

    University of Texas Southwestern Medical Center, Dallas, United States
    Competing interests
    No competing interests declared.
  3. David Flaherty

    University of Texas Southwestern Medical Center, Dallas, United States
    Competing interests
    No competing interests declared.
  4. Diana R Tomchick

    University of Texas Southwestern Medical Center, Dallas, United States
    Competing interests
    No competing interests declared.
  5. Josep Rizo

    University of Texas Southwestern Medical Center, Dallas, United States
    For correspondence
    jose@arnie.swmed.edu
    Competing interests
    No competing interests declared.
  6. Christian Rosenmund

    Charité-Universitätsmedizin Berlin, Berlin, Germany
    Competing interests
    Christian Rosenmund, Reviewing editor, eLife.

Ethics

Animal experimentation: Animals were handled according to the animal welfare committee of the Charit� Berlin, Germany. Time pregnant females were anesthetized and euthanized at E18 according to the german animal law ({section sign}4 section 1) and permit by the Berlin authorities (Landesamt f�r Gesundheit und Soziales, LaGeSo) under the permit number T0220/09.

Reviewing Editor

  1. Axel T Brunger, Stanford University, United States

Publication history

  1. Received: January 24, 2014
  2. Accepted: May 6, 2014
  3. Accepted Manuscript published: May 8, 2014 (version 1)
  4. Version of Record published: June 3, 2014 (version 2)

Copyright

© 2014, Trimbuch et al.

This article is distributed under the terms of the Creative Commons Attribution License, which permits unrestricted use and redistribution provided that the original author and source are credited.

Metrics

  • 2,290
    Page views
  • 292
    Downloads
  • 51
    Citations

Article citation count generated by polling the highest count across the following sources: Scopus, Crossref, PubMed Central.

Download links

A two-part list of links to download the article, or parts of the article, in various formats.

Downloads (link to download the article as PDF)

Open citations (links to open the citations from this article in various online reference manager services)

Cite this article (links to download the citations from this article in formats compatible with various reference manager tools)

  1. Thorsten Trimbuch
  2. Junjie Xu
  3. David Flaherty
  4. Diana R Tomchick
  5. Josep Rizo
  6. Christian Rosenmund
(2014)
Re-examining how complexin inhibits neurotransmitter release: SNARE complex insertion or electrostatic hindrance?
eLife 3:e02391.
https://doi.org/10.7554/eLife.02391

Further reading

    1. Biochemistry and Chemical Biology
    2. Structural Biology and Molecular Biophysics
    Shyam S Krishnakumar et al.
    Short Report

    We have previously proposed that complexin cross-links multiple pre-fusion SNARE complexes via a trans interaction to function as a clamp on SNARE-mediated neurotransmitter release. A recent NMR study was unable to detect the trans clamping interaction of complexin and therefore questioned the previous interpretation of the fluorescence resonance energy transfer and isothermal titration calorimetry data on which the trans clamping model was originally based. Here we present new biochemical data that underscore the validity of our previous interpretation and the continued relevancy of the trans insertion model for complexin clamping.

    1. Structural Biology and Molecular Biophysics
    Christian JG Tessier et al.
    Research Article Updated

    Human adult muscle-type acetylcholine receptors are heteropentameric ion channels formed from two α-subunits, and one each of the β-, δ-, and ε-subunits. To form functional channels, the subunits must assemble with one another in a precise stoichiometry and arrangement. Despite being different, the four subunits share a common ancestor that is presumed to have formed homopentamers. The extent to which the properties of the modern-day receptor result from its subunit complexity is unknown. Here, we discover that a reconstructed ancestral muscle-type β-subunit can form homopentameric ion channels. These homopentamers open spontaneously and display single-channel hallmarks of muscle-type acetylcholine receptor activity. Our findings attest to the homopentameric origin of the muscle-type acetylcholine receptor, and demonstrate that signature features of its function are both independent of agonist and do not necessitate the complex heteropentameric architecture of the modern-day protein.