t-SNARE protein conformations patterned by the lipid microenvironment.
The spatial distribution of the target (t-)SNARE proteins (syntaxin and SNAP-25) on the plasma membrane has been extensively characterized. However, the protein conformations and interactions of the two t-SNAREs in situ remain poorly defined. By using super-resolution optical techniques and fluorescence lifetime imaging microscopy, we observed that within the t-SNARE clusters syntaxin and SNAP-25 molecules interact, forming two distinct conformations of the t-SNARE binary intermediate. These are spatially segregated on the plasma membrane with each cluster exhibiting predominantly one of the two conformations, representing the two- and three-helical forms previously observed in vitro. We sought to explain why these two t-SNARE intermediate conformations exist in spatially distinct clusters on the plasma membrane. By disrupting plasma membrane lipid order, we found that all of the t-SNARE clusters now adopted a single conformational state corresponding to the three helical t-SNARE intermediates. Together, our results define spatially distinct t-SNARE intermediate states on the plasma membrane and how the conformation adopted can be patterned by the underlying lipid environment.
Rickman
Colin
C
Centre for Integrative Physiology, School of Informatics, University of Edinburgh, Edinburgh EH8 9XD, Scotland, United Kingdom.
Medine
Claire N
CN
Dun
Alison R
AR
Moulton
David J
DJ
Mandula
Ondrej
O
Halemani
Nagaraj D
ND
Rizzoli
Silvio O
SO
Chamberlain
Luke H
LH
Duncan
Rory R
RR
D000818:Animals; D002462:Cell Membrane / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism; D008563:Membrane Lipids / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism; D016716:PC12 Cells; D020836:Protein Structure, Quaternary; D017433:Protein Structure, Secondary; D050765:Qa-SNARE Proteins / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism; D051381:Rats; D050825:Synaptosomal-Associated Protein 25 / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism
D008563:Membrane Lipids; D050765:Qa-SNARE Proteins; C494210:Snap25 protein, rat; D050825:Synaptosomal-Associated Protein 25