Biophysical Society Thematic Meeting| Padova 2019

Quantitative Aspects of Membrane Fusion and Fission

Poster Abstracts

27-POS Board 27 REGULATION OF EXOCYTOSIS BY AMISYN, A PI(4,5)P 2 AND SYNTAXIN- BINDING PROTEIN Ilona Kondratiuk 1,2 ; Shrutee Jakhanwal 3 ; Reinhard Jahn 3 ; Ira Milosevic 1,2 ; 1 European Neuroscience Institute (ENI), Göttingen, Niedersachsen, Germany 2 University Medical Center Göttingen (UMG), Göttingen, Niedersachsen, Germany 3 Max Planck Institute for Biophysical Chemistry, Göttingen, Niedersachsen, Germany Higher functions of the brain, for example learning and memory, are mediated by fast and precisely coordinated neurotransmitter release through the process of regulated exocytosis. Intense research in the past three decades has identified numerous proteins involved in exocytosis, including the Sec1/Munc18 (SM) protein family (Munc18, Munc13), synaptotagmins that sense calcium, and the SNARE complex proteins: synaptobrevin-2/VAMP-2, syntaxin-1 and SNAP-25 that mediate membrane fusion. While the key exocytic proteins are highly conserved through evolution, the regulation of exocytosis has advanced and requires more proteins in the higher organisms, such as vertebrates. In addition to the core set of exocytic machinery, exocytosis is regulated by complexin, tomosyn and amisyn (STXBP6), cytosolic proteins that bind the SNARE complex. Amisyn is reported to be an important negative regulator of exocytosis, yet little is known about this brain-enriched protein. We found that, in addition to the C-terminal SNARE motif that interacts with syntaxin-1 and forms ‘fusion-inactive’ SNARE complex, amisyn contains an N-terminal pleckstrin homology (PH) domain. The PH domain of amisyn is phosphatidylinositol-4,5-bisphosphate (PI(4,5)P 2 ) specific, and it mediates its interaction with the plasma membrane. Given that amisyn is a conserved protein present only in vertebrates, it makes it necessary to characterize it better. We have generated amisyn knock-out mice to study amisyn-dependent processes at the vertebrate neurons and neurosecretory cells. We found that amisyn is important for the priming of secretory vesicles and the size of vesicle pools, but not fusion kinetics. Curiously, the inhibition is not due to amisyn’s SNARE motif binding to syntaxin-1, but the full-length protein is needed for the proper control of exocytosis. We are currently further investigating the mechanisms of amisyn-dependent inhibition and its implication to neurotransmission.

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