CAPS and Munc13 utilize distinct PIP<sub>2</sub>-linked mechanisms to promote vesicle exocytosis

Kabachinski, Greg; Yamaga, Masaki; Kielar-Grevstad, D. Michelle; Bruinsma, Stephen P.; Martin, Thomas F.J.

doi:10.1091/mbc.e12-11-0829

Cited by 62 publications

(124 citation statements)

References 62 publications

(143 reference statements)

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“…Munc13 binding to the plasma membrane depends on PIP2 and Ca 2 þ (ref. 54) and we noticed rapid depolarization-induced recruitment of munc13 to the membrane, although specific recruitment to the docking site might still depend on accumulation of PIP2 within the clusters 33,34 . Such a mechanism would provide a convenient explanation for the dependence of priming on munc13, PIP2 and Ca 2 þ .…”

Section: Discussionmentioning

confidence: 67%

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

2014

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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.

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Section: Discussionmentioning

confidence: 67%

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

2014

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“…Greater Ca 2+ influx did generate DAG (see below). As anticipated for the lower affinity of the PLCδ 4 -PH domain for PI(4,5)P 2 [15] as compared to a PLCδ 1 -PH domain probe, the PLCδ 4 -PH domain probe exhibited reduced partitioning onto the plasma membrane and only partially inhibited vesicle exocytosis [51]. These studies indicate that vesicle exocytosis can occur at membrane sites highly enriched for PI(4,5)P 2 .…”

Section: Pi(45)p2 Localizes To Sites Of Vesicle Exocytosismentioning

confidence: 60%

“…This was suggested by studies in which the co-localization of PI(4,5)P 2 with vesicles was reduced by briefly evoking exocytosis with Ca 2+ influx [46]. Whether evoked vesicle fusion occurs preferentially at PI(4,5)P 2 -rich plasma membrane sites was addressed in a recent study [51]. Kabachinski et al used a lower affinity PI(4,5)P 2 -binding PH domain probe derived from PLCδ 4 rather than PLCδ 1 to image PI(4,5)P 2 microdomains in live PC12 cells by TIRF microscopy.…”

Section: Pi(45)p2 Localizes To Sites Of Vesicle Exocytosismentioning

confidence: 99%

“…A PI(4)P 5-kinase β isoform co-localized with PI(4,5)P 2 /SM raft domains possibly through associations with RhoA [26]. While it is clear that PI(4,5)P 2 concentrations in microdomains can be modulated by kinases, phosphatases, and PLCs [8, 47, 51], additional studies are needed to identify turnover rates and the basis for enzyme access into high concentration clusters of PI(4,5)P 2 .…”

Section: Basis For Pi(45)p2 Cluster Formationmentioning

confidence: 99%

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PI(4,5)P2-binding effector proteins for vesicle exocytosis

Martin

2015

Biochimica et Biophysica Acta (BBA) - Molecular and Cell Biolog

114

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PI(4,5)P2 participates directly in priming and possibly fusion steps of Ca2+-triggered vesicle exocytosis. High concentration nanodomains of PI(4,5)P2 reside on the plasma membrane of neuroendocrine cells. A subset of vesicles that co-localize with PI(4,5)P2 domains appear to undergo preferential exocytosis in stimulated cells. PI(4,5)P2 directly regulates vesicle exocytosis by recruiting and activating PI(4,5)P2-binding proteins that regulate SNARE protein function including CAPS, Munc13-1/2, synaptotagmin-1, and other C2 domain-containing proteins. These PI(4,5)P2 effector proteins are coincidence detectors that engage in multiple interactions at vesicle exocytic sites. The SNARE protein syntaxin-1 also binds to PI(4,5)P2, which promotes clustering, but an activating role for PI(4,5)P2 in syntaxin-1 function remains to be fully characterized. Similar principles underlie polarized constitutive vesicle fusion mediated in part by the PI(4,5)P2-binding subunits of the exocyst complex (Sec3, Exo70). Overall, focal vesicle exocytosis occurs at sites landmarked by PI(4,5)P2, which serves to recruit and/or activate multifunctional PI(4,5)P2-binding proteins.

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“…Distinct from the consensus view of the cluster organization of syntaxin 1a, the spatial distribution of phosphoinositides in the PM is controversial. PI(4,5)P 2 distribution patterns range from uniform [10][11][12] , large patches 13,14 , to dense clusters [14][15][16][17][18] , depending on cell types and experimental methods used. The spatial organization of PI(4,5)P 2 at higher resolution is also inconsistent.…”

Section: Introductionmentioning

confidence: 99%

Single-molecule Super-resolution Imaging of Phosphatidylinositol 4,5-bisphosphate in the Plasma Membrane with Novel Fluorescent Probes

Chen

Lou

2016

JoVE

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Phosphoinositides in the cell membrane are signaling lipids with multiple cellular functions. Phosphatidylinositol 4,5-bisphosphate (PI(4,5)P 2 ) is a determinant phosphoinositide of the plasma membrane (PM), and it is required to modulate ion channels, actin dynamics, exocytosis, endocytosis, intracellular signaling, and many other cellular processes. However, the spatial organization of PI(4,5)P 2 in the PM is controversial, and its nanoscale distribution is poorly understood due to the technical limitations of research approaches. Here by utilizing single molecule localization microscopy and the Pleckstrin Homology (PH) domain based dual color fluorescent probes, we describe a novel method to visualize the nanoscale distribution of PI(4,5)P 2 in the PM in fixed membrane sheets as well as live cells.

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CAPS and Munc13 utilize distinct PIP₂-linked mechanisms to promote vesicle exocytosis

Cited by 62 publications

References 62 publications

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

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

PI(4,5)P2-binding effector proteins for vesicle exocytosis

Single-molecule Super-resolution Imaging of Phosphatidylinositol 4,5-bisphosphate in the Plasma Membrane with Novel Fluorescent Probes

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CAPS and Munc13 utilize distinct PIP2-linked mechanisms to promote vesicle exocytosis

Cited by 62 publications

References 62 publications

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

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

PI(4,5)P2-binding effector proteins for vesicle exocytosis

Single-molecule Super-resolution Imaging of Phosphatidylinositol 4,5-bisphosphate in the Plasma Membrane with Novel Fluorescent Probes

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CAPS and Munc13 utilize distinct PIP₂-linked mechanisms to promote vesicle exocytosis