Synaptic vesicle exocytosis is a highly specialized vesicle trafficking process in which calcium triggers fusion of synaptic vesicles with the plasma membrane, resulting in neurotransmitter release. SNARE complex assembly between synaptobrevin, SNAP-25 and syntaxin is a critical requirement preceeding this vesicle fusion event. Several SNARE-interacting proteins, have been shown to profoundly influence the strength of synaptic transmssion, through their regulatory effects on the SNARE complex. Recently, a new SNARE binding partner, tomosyn was isolated from rat brain cytosol. Tomosyn has a SNARE binding domain that can compete with synaptobrevin for assembly into a tomosyn SNARE complex with syntaxin and SNAP-25. Based on these biochemical observations as well as tomosyn overexpression data, tomosyn is proposed to regulate vesicle release through an undefined mechanism. There are presently no loss-of-functions mutants available in any organism other than C. elegans. Therefore, we intend to examine the mechanism of tomosyn action at synapses in this powerful genetic model organism.
Aim 1) Characterize the synaptic phenotype of tom-1 deletion mutants. We have obtained two tom-1 deletion mutants that have phenotypes consistent with increased synaptic transmission. We will conduct a detailed characterization of these tom-1 mutants including behavioral, cytoarchitectural, pharmacological, electophysiological and ultrastructural analyses.
Aim 2) Determine which TOM-1 isoforms regulate synaptic transmission. C. elegans tom-1 encodes three isoforms. The isoform expression patterns will be ascertained and mosaic analysis and tissue specific rescue experiments will be performed.
Aim 3) Genetic analysis of TOM-1 function. We hyptheisize that tomosyn regulates the priming step of exocytosis. To test this model we will generate and characterize double mutants between tom-1 and several mutants known to effect the vesicle primed pool (unc-13, unc- 10, open-syntaxin and unc-18).
Aim 4) Identify TOM-1 domains required for the regulation of synaptic transmission. TOM-1 protein domains essential for the regulation of exocytosis will be identified using a genetic screen for mutants that fail to complement the tom-1 mutation. These experiments are likely to further our understanding of neurotransmission, a foundation that may contribute to our understanding of neurological diseases and vesicle trafficking disorders.

Agency
National Institute of Health (NIH)
Institute
National Institute of Mental Health (NIMH)
Type
Research Project (R01)
Project #
5R01MH073156-04
Application #
7579948
Study Section
Special Emphasis Panel (ZRG1-MDCN-L (02))
Program Officer
Asanuma, Chiiko
Project Start
2006-04-01
Project End
2011-01-31
Budget Start
2009-02-01
Budget End
2010-01-31
Support Year
4
Fiscal Year
2009
Total Cost
$312,165
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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