Neurons release neurotransmitter into the synaptic cleft by exocytosis of synaptic vesicles. The proteins that mediate exocytosis are members of conserved protein families involved in general intracellular fusion events. Critical proteins in this process include the SNARE proteins, syntaxin, SNAP-25 and synaptobrevin, as well as UNC-18 and UNC-13. Although we know that UNC-18 and UNC-13 interact with the essential SNARE protein syntaxin, the precise role and sequential order in which these proteins regulate fusion have yet to be determined. In my laboratory we combine genetic and molecular approaches with a newly developed electrophysiological technique to study exocytosis in the nematode Caenorhabditis elegans. Using these techniques we have demonstrated that exocytosis is arrested at a late stage in unc-13 mutants. We now propose to examine the role of UNC-18 in C. elegans synaptic transmission. UNC-18 binds to a closed conformation of syntaxin, which excludes the formation of the SNARE complex. Several models have been proposed for UNC-18 function: (1) UNC-18 dissociation may promote or maintain syntaxin in an open state, enabling SNARE complex assembly and fusion. (2) UNC-18 may maintain syntaxin in the closed conformation to prevent SNARE complex formation. (3) UNC- 18 and syntaxin may mediate the fusion step directly. To test these models, we propose to: 1) Determine whether UNC-18 promotes exocytosis after vesicles have docked to the plasma membrane. 2) Test whether the UNC-18-syntaxin interaction plays an inhibitory role in exocytosis. 3) Test whether constitutively open syntaxin bypasses the requirement for UNC-18. 4) Identify other proteins that act downstream or in parallel with UNC-18 by a) mapping and identifying a recently isolated unc-18 mutant suppressor and b) conducting a genetic screen to identify other unc-18 suppressors. These experiments may contribute to our understanding of Alzheimer's disease, stroke and vesicle trafficking disorders.

Agency
National Institute of Health (NIH)
Institute
National Institute of Neurological Disorders and Stroke (NINDS)
Type
Research Project (R01)
Project #
5R01NS041477-04
Application #
6717658
Study Section
Special Emphasis Panel (ZRG1-MDCN-1 (01))
Program Officer
Talley, Edmund M
Project Start
2002-03-01
Project End
2007-02-28
Budget Start
2004-03-01
Budget End
2005-02-28
Support Year
4
Fiscal Year
2004
Total Cost
$259,134
Indirect Cost
Name
University of Illinois at Chicago
Department
Biology
Type
Schools of Arts and Sciences
DUNS #
098987217
City
Chicago
State
IL
Country
United States
Zip Code
60612
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Gracheva, Elena O; Hadwiger, Gayla; Nonet, Michael L et al. (2008) Direct interactions between C. elegans RAB-3 and Rim provide a mechanism to target vesicles to the presynaptic density. Neurosci Lett 444:137-42
Eimer, Stefan; Gottschalk, Alexander; Hengartner, Michael et al. (2007) Regulation of nicotinic receptor trafficking by the transmembrane Golgi protein UNC-50. EMBO J 26:4313-23
Gracheva, Elena O; Burdina, Anna O; Touroutine, Denis et al. (2007) Tomosyn negatively regulates CAPS-dependent peptide release at Caenorhabditis elegans synapses. J Neurosci 27:10176-84
Gracheva, Elena O; Burdina, Anna O; Holgado, Andrea M et al. (2006) Tomosyn inhibits synaptic vesicle priming in Caenorhabditis elegans. PLoS Biol 4:e261
Rowland, Aaron M; Richmond, Janet E; Olsen, Jason G et al. (2006) Presynaptic terminals independently regulate synaptic clustering and autophagy of GABAA receptors in Caenorhabditis elegans. J Neurosci 26:1711-20
Weimer, Robby M; Gracheva, Elena O; Meyrignac, Olivier et al. (2006) UNC-13 and UNC-10/rim localize synaptic vesicles to specific membrane domains. J Neurosci 26:8040-7
Wang, Ying; Gracheva, Elena O; Richmond, Janet et al. (2006) The C2H2 zinc-finger protein SYD-9 is a putative posttranscriptional regulator for synaptic transmission. Proc Natl Acad Sci U S A 103:10450-5
Richmond, Janet E (2006) Electrophysiological recordings from the neuromuscular junction of C. elegans. WormBook :1-8
Towers, Paula R; Edwards, Ben; Richmond, Janet E et al. (2005) The Caenorhabditis elegans lev-8 gene encodes a novel type of nicotinic acetylcholine receptor alpha subunit. J Neurochem 93:1-9

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