Contact-induced clustering of syntaxin and munc18 docks secretory granules at the exocytosis site

Abstract

Docking of secretory vesicles at the plasma membrane is a poorly understood prerequisite for exocytosis. Current models propose raft-like clusters containing syntaxin as docking receptor, but direct evidence for this is lacking. Here we provide quantitative measurements of several exocytosis proteins (syntaxin, SNAP25, munc18, munc13 and rab3) at the insulin granule release site and show that docking coincides with rapid de novo formation of syntaxin1/ munc18 clusters at the nascent docking site. Formation of such clusters prevents undocking and is not observed during failed docking attempts. Overexpression of syntaxins' N-terminal Habc-domain competitively interferes with both cluster formation and successful docking. SNAP25 and munc13 are recruited to the docking site more than a minute later, consistent with munc13's reported role in granule priming rather than docking. We conclude that secretory vesicles dock by inducing syntaxin1/munc18 clustering in the target membrane, and find no evidence for preformed docking receptors.

PIP2Clustering: From model membranes to cells

“…Mutation of basic residues at the cytoplasmic juxtamembrane interface abrogated PIP2 co-clustering with syntaxin 1, supporting an electrostatic mechanism for PIP2 recruitment (van den Bogaart et al, 2011). Association of docked vesicles with syntaxin-PIP2 clusters is required for Ca++-stimulated fusion (Honigmann et al, 2013;Gandasi and Barg, 2014;Martin, 2015). Clustered PIP2 functions to recruit downstream PIP2 binding proteins that regulate SNARE function, including CAPS, Munc13, and synaptotagmin (Honigmann et al, 2013;Martin, 2015).…”

Activity-driven relaxation of the cortical actomyosin II network synchronizes Munc18-1-dependent neurosecretory vesicle docking

“…Recent evidence suggests that SVs can organize their own release sites on reaching the plasma membrane 32,43 . Indeed, syntaxin-1 molecules have been shown to concentrate under docked vesicles before fusion, suggesting that a pool of plasma membrane syntaxin-1 can be recruited by an unknown mechanism under SVs 43 .…”

“…Recent evidence suggests that SVs can organize their own release sites on reaching the plasma membrane 32,43 . Indeed, syntaxin-1 molecules have been shown to concentrate under docked vesicles before fusion, suggesting that a pool of plasma membrane syntaxin-1 can be recruited by an unknown mechanism under SVs 43 . Another possible contributing factor could involve translocated actin exerting an effect on the domain organization of the plasma membrane 44 leading to efficient docking as previously speculated 45 .…”

Endogenous beta-cell CART regulates insulin secretion and transcription of beta-cell genes

“…SNAP25 has earlier been demonstrated to have a fundamental role in beta-cell exocytosis through mechanisms that are independent of the Ca 2þ -influx (Vikman et al, 2006(Vikman et al, , 2009. Syntaxin 1 together with STXBP1 are critical for the formation of granular docking sites (Gandasi and Barg, 2014) and STXBP1 together with SYTL4 are important for docking (Tomas et al, 2008). Thus, the CART KD induced reduction of Stxbp1 expression and increase in Stx1a and Syt4 expression is more complex, but might explain why no effect is observed on granular docking and mobilization (represented by increase in membrane capacitance by the later depolarizations of the train).…”