The goal of this research proposal is to develop a better understanding of the mechanisms underlying neurotransmitter release. Syntaxin-1A is a conserved presynaptic membrane protein that has been shown to be essential for exocytotic release of synaptic vesicles. Syntaxin is thought to mediate its function through its interactions with other proteins involved in neurotransmitter release. In addition to these interactions, phosphorylation of syntaxin may potentially play a critical role in regulating syntaxin's function in neurotransmitter release, as is the case for other synaptic proteins, such as synapsin. This work proposes to perform a mutational analysis of syntaxin 1-A in Drosophila to identify amino acids or regions that are critical for its function in vivo. To perform this analysis, targeted mutagenesis will be performed on syntaxin-1A in Drosophila to generate systematic regional deletions and point mutations that may disrupt syntaxin's interactions with other proteins and disrupt specific phosphorylation sites. The importance and function of these regions and amino acids will be assessed by observation of mutant flies, electrophysiological analysis and in vitro binding assays. The dissection of the function of syntaxin-1A in vivo through electrophysiological and biochemical analysis should yield important insights into the mechanisms of synaptic vesicle release. Understanding this mechanism could provide a framework for addressing issues such as drug action at the synapse disease states associated with changes in neurotransmitter release, and the processes underlying learning and memory.
