RT Journal Article SR Electronic T1 UNC-18 and Tomosyn antagonistically control synaptic vesicle priming downstream of UNC-13 in C. elegans JF The Journal of Neuroscience JO J. Neurosci. FD Society for Neuroscience SP 0338-17 DO 10.1523/JNEUROSCI.0338-17.2017 A1 Seungmee Park A1 Na-Ryum Bin A1 Bin Yu A1 Raymond Wong A1 Ewa Sitarska A1 Kyoko Sugita A1 Ke Ma A1 Junjie Xu A1 Chi-Wei Tien A1 Arash Algouneh A1 Ekaterina Turlova A1 Siyan Wang A1 Pranay Siriya A1 Waleed Shahid A1 Lorraine Kalia A1 Zhong-Ping Feng A1 Philippe P. Monnier A1 Hong-Shuo Sun A1 Mei Zhen A1 Shangbang Gao A1 Josep Rizo A1 Shuzo Sugita YR 2017 UL http://www.jneurosci.org/content/early/2017/08/08/JNEUROSCI.0338-17.2017.abstract AB Munc18-1/UNC-18 is believed to prime SNARE-mediated membrane fusion, yet the underlying mechanisms remain enigmatic. Here, we examined how potential gain-of-function mutations of Munc18-1/UNC-18 affect locomotory behavior and synaptic transmission, and how Munc18-1-mediated priming is related to Munc13-1/UNC-13 and Tomosyn/TOM-1, positive and negative SNARE regulators, respectively. We show that a Munc18-1(P335A)/UNC-18(P334A) mutation leads to significantly increased locomotory activity and acetylcholine release in C. elegans, as well as enhanced synaptic neurotransmission in cultured mammalian neurons. Importantly, similar to tom-1 null mutants, unc-18(P334A) mutants partially bypass the requirement of UNC-13. Moreover, unc-18(P334A) and tom-1 null mutations confer a strong synergy in suppressing the phenotypes of unc-13 mutants. Through biochemical experiments, we demonstrate that Munc18-1(P335A) exhibits enhanced activity in SNARE complex formation as well as in binding to the preformed SNARE complex, and partially bypasses the Munc13-1 requirement in liposome fusion assays. Our results indicate that Munc18-1/UNC-18 primes vesicle fusion downstream of Munc13-1/UNC-13 by templating SNARE complex assembly, and acts antagonistically with Tomosyn/TOM-1.SIGNIFICANCE STATEMENTAt presynaptic sites, SNARE-mediated membrane fusion is tightly regulated by several key proteins including Munc18/UNC-18, Munc13/UNC-13 and Tomosyn/TOM-1. However, how these proteins interact with each other to achieve the precise regulation of neurotransmitter release remains largely unclear. Using C. elegans as an in vivo model, we found that a gain-of-function mutant of UNC-18 increases locomotory activity and synaptic acetylcholine release, that it partially bypasses the requirement of UNC-13 for release, and that this bypass is synergistically augmented by the lack of TOM-1. We also elucidated the biochemical basis for the gain-of-function caused by this mutation. Thus, our study provides novel mechanistic insights into how Munc18/UNC-18 primes synaptic vesicle release and how this protein interacts functionally with Munc13/UNC-13 and Tomosyn/TOM-1.