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Visualizing Intracellular SNARE Trafficking by Fluorescence Lifetime Imaging Microscopy
Published on: December 29, 2017
Subunit structure of a mammalian ER/Golgi SNARE complex.
D Xu1, A P Joglekar, A L Williams
1Department of Biology and Program in Biomedical Sciences, University of Michigan, Ann Arbor, Michigan 48109-1048, USA.
The Journal of Biological Chemistry
|October 18, 2000
Summary
Researchers identified a new quaternary SNARE complex essential for ER to Golgi transport. This complex, crucial for membrane fusion, reveals conserved structural principles across different cellular transport steps.
Area of Science:
- Cell Biology
- Molecular Biology
- Membrane Trafficking
Background:
- Soluble NSF Attachment Protein (SNAP) receptor (SNARE) complexes mediate membrane fusion in cellular transport pathways.
- Detailed characterization has been limited to exocytic SNARE complexes, leaving structural similarities across fusion steps unknown.
Purpose of the Study:
- To elucidate the subunit structure, assembly, and regulation of a quaternary SNARE complex involved in early endoplasmic reticulum (ER) to Golgi transport.
- To compare the structure of this ER/Golgi complex with known exocytic SNARE complexes.
Main Methods:
- Purification and assembly of recombinant syntaxin 5, membrin, rbet1 (Q-SNAREs), and sec22b (R-SNARE).
- Analysis of SNARE complex assembly kinetics and regulation by the syntaxin 5 amino-terminal domain.
- Validation of the quaternary complex's presence in living cells.
Main Results:
- A quaternary SNARE complex comprising syntaxin 5, membrin, rbet1, and sec22b was identified for ER to Golgi transport.
- Syntaxin 5's N-terminal domain inhibits SNARE complex assembly.
- The ER/Golgi complex shares structural similarities with synaptic complexes, suggesting a conserved helical assembly pattern (three Q-helices binding one R-helix).
- Sec22b binding requires its presence during the assembly of the other SNAREs, not to a pre-assembled complex.
Conclusions:
- A conserved structural motif (three Q-helices forming a binding site for an R-helix) is utilized across different membrane fusion events.
- The identified quaternary SNARE complex is a functional entity in living cells, not just an in vitro artifact.
- Understanding this complex provides insights into the fundamental mechanisms of intracellular membrane trafficking.
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