Ultrahigh-resolution imaging reveals formation of neuronal SNARE/Munc18 complexes in situ Academic Article uri icon

Overview

MeSH Major

  • Aging
  • Nerve Degeneration
  • Neurons
  • Presynaptic Terminals
  • SNARE Proteins
  • alpha-Synuclein

abstract

  • Membrane fusion is mediated by complexes formed by SNAP-receptor (SNARE) and Secretory 1 (Sec1)/mammalian uncoordinated-18 (Munc18)-like (SM) proteins, but it is unclear when and how these complexes assemble. Here we describe an improved two-color fluorescence nanoscopy technique that can achieve effective resolutions of up to 7.5-nm full width at half maximum (3.2-nm localization precision), limited only by stochastic photon emission from single molecules. We use this technique to dissect the spatial relationships between the neuronal SM protein Munc18-1 and SNARE proteins syntaxin-1 and SNAP-25 (25 kDa synaptosome-associated protein). Strikingly, we observed nanoscale clusters consisting of syntaxin-1 and SNAP-25 that contained associated Munc18-1. Rescue experiments with syntaxin-1 mutants revealed that Munc18-1 recruitment to the plasma membrane depends on the Munc18-1 binding to the N-terminal peptide of syntaxin-1. Our results suggest that in a primary neuron, SNARE/SM protein complexes containing syntaxin-1, SNAP-25, and Munc18-1 are preassembled in microdomains on the presynaptic plasma membrane. Our superresolution imaging method provides a framework for investigating interactions between the synaptic vesicle fusion machinery and other subcellular systems in situ.

publication date

  • July 23, 2013

Research

keywords

  • Academic Article

Identity

Language

  • eng

PubMed Central ID

  • PMC3725100

Digital Object Identifier (DOI)

  • 10.1073/pnas.1221464110

PubMed ID

  • 23776236

Additional Document Info

start page

  • E2812

end page

  • 20

volume

  • 110

number

  • 30