The SNARE hypothesis states that the folding and assembly of four-helix SNARE complex bundles drives intracellular membrane fusion, including fusion in exocytosis of synaptic vesicles at the plasma membranes of synaptic terminals in neurons. The neuronal SNAREs are syntaxin 1 a and SNAP-25 on the plasma membrane and synaptobrevin 2 on the vesicle membrane. Syntaxin contributes one and SNAP-25 contributes two helices to the active acceptor SNARE complex on the target membrane. Synaptic vesicles dock to the presynaptic plasma membrane by contributing synaptobrevin to the nascent SNARE complex. How far this trans-SNARE complex that bridges the two membranes is folded is presently not known. The trans-SNARE complex is then thought to zipper-fold from the N- towards the C-terminus of the parallel fourhelix bundle. The bundle, thereby pulling the two membranes into close proximity. In a final step, SNARE complex folding proceeds into the transmembrane domains of syntaxin and synaptobrevin to form a cis- SNARE complex, i.e. a step which must be coupled with merging the lipid bilayers of the two membranes. The research proposed in this project will dissect the described folding steps and correlate them with different stages of vesicle docking and membrane fusion. Magnetic resonance approaches will be taken to determine (dynamic) structures of SNARE folding intermediates and micro-fluorescence approaches will be taken to determine the kinetics and higher architecture of the evolving fusion pore in artificial and cellular membrane preparations.

Public Health Relevance

Fusion of synaptic vesicles with the presynaptic membrane of neurons is a key element of neurotransmitter release in synaptic transmission. Defects in synaptic transmission lead to epilepsy, depression, and other neurological disorders. The research of this grant will elucidate the basic mechanism that leads to presynaptic membrane fusion and thus help define disorders in which this mechanism is disturbed.

Agency
National Institute of Health (NIH)
Institute
National Institute of General Medical Sciences (NIGMS)
Type
Research Program Projects (P01)
Project #
5P01GM072694-07
Application #
8376195
Study Section
Special Emphasis Panel (ZRG1-BCMB-S)
Project Start
Project End
Budget Start
2012-07-01
Budget End
2013-06-30
Support Year
7
Fiscal Year
2012
Total Cost
$335,458
Indirect Cost
$115,037
Name
University of Virginia
Department
Type
DUNS #
065391526
City
Charlottesville
State
VA
Country
United States
Zip Code
22904
Yang, Sung-Tae; Lim, Sung In; Kiessling, Volker et al. (2016) Site-specific fluorescent labeling to visualize membrane translocation of a myristoyl switch protein. Sci Rep 6:32866
Dawidowski, Damian; Cafiso, David S (2016) Munc18-1 and the Syntaxin-1 N Terminus Regulate Open-Closed States in a t-SNARE Complex. Structure 24:392-400
Milovanovic, Dragomir; Platen, Mitja; Junius, Meike et al. (2016) Calcium Promotes the Formation of Syntaxin 1 Mesoscale Domains through Phosphatidylinositol 4,5-Bisphosphate. J Biol Chem 291:7868-76
Kreutzberger, Alex J B; Liang, Binyong; Kiessling, Volker et al. (2016) Assembly and Comparison of Plasma Membrane SNARE Acceptor Complexes. Biophys J 110:2147-50
Yang, Sung-Tae; Kreutzberger, Alex J B; Lee, Jinwoo et al. (2016) The role of cholesterol in membrane fusion. Chem Phys Lipids 199:136-43
Park, Yongsoo; Seo, Jong Bae; Fraind, Alicia et al. (2015) Synaptotagmin-1 binds to PIP(2)-containing membrane but not to SNAREs at physiological ionic strength. Nat Struct Mol Biol 22:815-23
Kiessling, Volker; Liang, Binyong; Tamm, Lukas K (2015) Reconstituting SNARE-mediated membrane fusion at the single liposome level. Methods Cell Biol 128:339-63
Kiessling, Volker; Yang, Sung-Tae; Tamm, Lukas K (2015) Supported lipid bilayers as models for studying membrane domains. Curr Top Membr 75:1-23
Kreutzberger, Alex J B; Kiessling, Volker; Tamm, Lukas K (2015) High cholesterol obviates a prolonged hemifusion intermediate in fast SNARE-mediated membrane fusion. Biophys J 109:319-29
Milovanovic, Dragomir; Honigmann, Alf; Koike, Seiichi et al. (2015) Hydrophobic mismatch sorts SNARE proteins into distinct membrane domains. Nat Commun 6:5984

Showing the most recent 10 out of 59 publications