scholarly journals The immunophilin Zonda controls regulated exocytosis in endocrine and exocrine tissues

2020 ◽  
Author(s):  
Rocío de la Riva Carrasco ◽  
Sebastián Perez Pandolfo ◽  
Sofía Suarez Freire ◽  
Nuria M. Romero ◽  
Zambarlal Bhujabal ◽  
...  
Keyword(s):  
Plasma Membrane ◽  
Salivary Gland ◽  
Secretory Granules ◽  
Model System ◽  
Genetic Screen ◽  
Small Gtpase ◽  
Extracellular Milieu ◽  
Insulin Producing Cells ◽  
Ring Gland ◽  
Fundamental Process

ABSTRACTExocytosis is a fundamental process in physiology, communication between cells, organs and even organisms. Hormones, neuropeptides and antibodies, among other cargoes are packed in exocytic vesicles that need to reach and fuse with the plasma membrane to release their content to the extracellular milieu. Hundreds of proteins participate in this process and several others in its regulation. We report here a novel component of the exocytic machinery, the Drosophila transmembrane immunophilin Zonda (Zda), previously found to participate in autophagy. Zda is highly expressed in secretory tissues, and regulates exocytosis in at least three of them: the ring gland, insulin-producing cells and the salivary gland. Using the salivary gland as a model system, we found that Zda is required at final steps of the exocytic process for fusion of secretory granules to the plasma membrane. In a genetic screen we identified the small GTPase RalA as a crucial regulator of secretory granule exocytosis that operates upstream of Zda in the process.

scholarly journals The Rab27a effector exophilin7 promotes fusion of secretory granules that have not been docked to the plasma membrane

2013 ◽  
Vol 24 (3) ◽  
pp. 319-330 ◽  
Author(s):  
Hao Wang ◽  
Ray Ishizaki ◽  
Jun Xu ◽  
Kazuo Kasai ◽  
Eri Kobayashi ◽  
...  
Keyword(s):  
Plasma Membrane ◽  
Knockout Mice ◽  
Secretory Granules ◽  
Small Gtpase ◽  
Membrane Phospholipids ◽  
Β Cells ◽  
Β Cell ◽  
Pancreatic Β Cells ◽  
Insulin Granules ◽  
Insulin Exocytosis

Granuphilin, an effector of the small GTPase Rab27a, mediates the stable attachment (docking) of insulin granules to the plasma membrane and inhibits subsequent fusion of docked granules, possibly through interaction with a fusion-inhibitory Munc18-1/syntaxin complex. However, phenotypes of insulin exocytosis differ considerably between Rab27a- and granuphilin-deficient pancreatic β cells, suggesting that other Rab27a effectors function in those cells. We found that one of the putative Rab27a effector family proteins, exophilin7/JFC1/Slp1, is expressed in β cells; however, unlike granuphilin, exophilin7 overexpressed in the β-cell line MIN6 failed to show granule-docking or fusion-inhibitory activity. Furthermore, exophilin7 has no affinities to either Munc18-1 or Munc18-1–interacting syntaxin-1a, in contrast to granuphilin. Although β cells of exophilin7-knockout mice show no apparent abnormalities in intracellular distribution or in ordinary glucose-induced exocytosis of insulin granules, they do show impaired fusion in response to some stronger stimuli, specifically from granules that have not been docked to the plasma membrane. Exophilin7 appears to mediate the fusion of undocked granules through the affinity of its C2A domain toward the plasma membrane phospholipids. These findings indicate that the two Rab27a effectors, granuphilin and exophilin7, differentially regulate the exocytosis of either stably or minimally docked granules, respectively.

scholarly journals Exophilin8 transiently clusters insulin granules at the actin-rich cell cortex prior to exocytosis

2011 ◽  
Vol 22 (10) ◽  
pp. 1716-1726 ◽  
Author(s):  
Kouichi Mizuno ◽  
José S. Ramalho ◽  
Tetsuro Izumi
Keyword(s):  
Plasma Membrane ◽  
Secretory Granules ◽  
Small Gtpase ◽  
Cell Cortex ◽  
Cortical Actin ◽  
Insulin Granules ◽  
Myosin Va ◽  
Exogenous Expression ◽  
Mutation Analyses

Exophilin8/MyRIP/Slac2-c is an effector protein of the small GTPase Rab27a and is specifically localized on retinal melanosomes and secretory granules. We investigated the role of exophilin8 in insulin granule trafficking. Exogenous expression of exophilin8 in pancreatic β cells or their cell line, MIN6, polarized (exophilin8-positive) insulin granules at the cell corners, where both cortical actin and the microtubule plus-end–binding protein, EB1, were present. Mutation analyses indicated that the ability of exophilin8 to act as a linker between Rab27a and myosin Va is essential for its granule-clustering activity. Moreover, exophilin8 and exophilin8-associated insulin granules were markedly stable and immobile. Total internal reflection fluorescence microscopy indicated that exophilin8 restricts the motion of insulin granules at a region deeper than that where another Rab27a effector, granuphilin, accumulates docked granules directly attached to the plasma membrane. However, the exophilin8-induced immobility of insulin granules was eliminated upon secretagogue stimulation and did not inhibit evoked exocytosis. Furthermore, exophilin8 depletion prevents insulin granules from being transported close to the plasma membrane and inhibits their fusion. These findings indicate that exophilin8 transiently traps insulin granules into the cortical actin network close to the microtubule plus-ends and supplies them for release during the stimulation.

Synaptotagmin (Syt) IX is an essential determinant for protein sorting to secretory granules in mast cells

Blood ◽  
2006 ◽  
Vol 109 (8) ◽  
pp. 3385-3392 ◽  
Author(s):  
Yael Haberman ◽  
Idit Ziv ◽  
Yaara Gorzalczany ◽  
Koret Hirschberg ◽  
Leonide Mittleman ◽  
...  
Keyword(s):  
Plasma Membrane ◽  
Mast Cells ◽  
Secretory Granules ◽  
Protein Sorting ◽  
Small Gtpase ◽  
Secretory Cells ◽  
Clathrin Adaptor ◽  
Lysosome Related Organelles ◽  
Golgi Network ◽  
Basophilic Leukemia

Abstract The secretory granules (SGs) of secretory cells of the hematopoietic lineage, such as the mast cells, are lysosome-related organelles whose membrane proteins travel through the plasma membrane and the endocytic system. Therefore, a mechanism must exist to prevent proteins destined to recycling or to the trans-Golgi network (TGN) from reaching the SGs. We now show that synaptotagmin (Syt) IX, a Syt homologue that is required for recycling from the endocytic recycling compartment (ERC) in rat basophilic leukemia (RBL-2H3) cultured mast cells, is involved in segregating recycling proteins from the SGs. By using as a marker the recycling protein TGN38, which cycles between the TGN, plasma membrane, and the ERC, we show that knock-down of Syt IX results in mistargeting of HA-tagged TGN38 to the SGs. We further demonstrate that Syt IX binds directly the small GTPase ARF1 and associates with the clathrin adaptor complex AP-1. These results therefore implicate Syt IX as an essential factor for the correct sorting of SGs proteins. Moreover, they place Syt IX as part of the machinery that is involved in the formation of transport carriers that mediate SGs protein sorting.

scholarly journals Rab3D, a Small GTPase, Is Localized on Mast Cell Secretory Granules and Translocates to the Plasma Membrane upon Exocytosis

1999 ◽  
Vol 20 (1) ◽  
pp. 79-89 ◽  
Author(s):  
Michael J. Tuvim ◽  
Roberto Adachi ◽  
Jose F. Chocano ◽  
Robert H. Moore ◽  
Robert M. Lampert ◽  
...  
Keyword(s):  
Mast Cell ◽  
Plasma Membrane ◽  
Secretory Granules ◽  
Small Gtpase

Rab27 and its effectors in secretory granule exocytosis: a novel docking machinery composed of a Rab27·effector complex

2006 ◽  
Vol 34 (5) ◽  
pp. 691-695 ◽  
Author(s):  
M. Fukuda
Keyword(s):  
Plasma Membrane ◽  
Secretory Granule ◽  
Binding Proteins ◽  
Secretory Granules ◽  
Small Gtpase ◽  
Secretory Cells ◽  
Cell Type ◽  
Tissue Specific ◽  
C2 Domains ◽  
A Cell

A small GTPase Rab27 is present on secretory granules in a wide variety of secretory cells and on melanosomes in melanocytes, and it is involved in controlling the trafficking of these organelles through interaction with a cell-type- or tissue-specific Rab27 effector(s). Slps (synaptotagmin-like proteins) and rabphilin contain an N-terminal Rab27-binding domain and C-terminal tandem C2 domains, and some of the Rab27-binding proteins have recently been shown to promote docking of Rab27-bound organelles to the plasma membrane. This mini-review presents a model for how the Rab27·effector complex controls the docking step in the trafficking of Rab27-bound organelles. Our results indicate that Slp2-a, Slp4-a/granuphilin-a and rabphilin are capable of interacting with the plasma membrane directly or indirectly, and thus that these Rab27 effectors form a bridge between Rab27-bound organelles and the plasma membrane.

scholarly journals Redistribution of a rab3-like GTP-binding protein from secretory granules to the Golgi complex in pancreatic acinar cells during regulated exocytosis

1994 ◽  
Vol 124 (1) ◽  
pp. 43-53 ◽  
Author(s):  
BP Jena ◽  
FD Gumkowski ◽  
EM Konieczko ◽  
GF von Mollard ◽  
R Jahn ◽  
...  
Keyword(s):  
Plasma Membrane ◽  
Golgi Complex ◽  
Binding Protein ◽  
Secretory Granules ◽  
Acinar Cells ◽  
Pancreatic Acinar Cells ◽  
Apical Plasma Membrane ◽  
Gtp Binding Protein ◽  
Regulated Exocytosis ◽  
Gtp Binding

Regulated secretion from pancreatic acinar cells occurs by exocytosis of zymogen granules (ZG) at the apical plasmalemma. ZGs originate from the TGN and undergo prolonged maturation and condensation. After exocytosis, the zymogen granule membrane (ZGM) is retrieved from the plasma membrane and ultimately reaches the TGN. In this study, we analyzed the fate of a low M(r) GTP-binding protein during induced exocytosis and membrane retrieval using immunoblots as well as light and electron microscopic immunocytochemistry. This 27-kD protein, identified by a monoclonal antibody that recognizes rab3A and B, may be a novel rab3 isoform. In resting acinar cells, the rab3-like protein was detected primarily on the cytoplasmic face of ZGs, with little labeling of the Golgi complex and no significant labeling of the apical plasmalemma or any other intracellular membranes. Stimulation of pancreatic lobules in vitro by carbamylcholine for 15 min, resulted in massive exocytosis that led to a near doubling of the area of the apical plasma membrane. However, no relocation of the rab3-like protein to the apical plasmalemma was seen. After 3 h of induced exocytosis, during which time approximately 90% of the ZGs is released, the rab3-like protein appeared to translocate to small vesicles and newly forming secretory granules in the TGN. No significant increase of the rab3-like protein was found in the cytosolic fraction at any time during stimulation. Since the protein is not detected on the apical plasmalemma after stimulation, we conclude that recycling may involve a membrane dissociation-association cycle that accompanies regulated exocytosis.

Barrier role of actin filaments in regulated mucin secretion from airway goblet cells

2005 ◽  
Vol 288 (1) ◽  
pp. C46-C56 ◽  
Author(s):  
Camille Ehre ◽  
Andrea H. Rossi ◽  
Lubna H. Abdullah ◽  
Kathleen De Pestel ◽  
Sandra Hill ◽  
...  
Keyword(s):  
Plasma Membrane ◽  
Actin Filaments ◽  
Fluorescent Protein ◽  
Secretory Granules ◽  
Yellow Fluorescent Protein ◽  
Goblet Cells ◽  
Actin Binding ◽  
Secretory Cells ◽  
Cortical Actin ◽  
Mucin Secretion

Airway goblet cells secrete mucin onto mucosal surfaces under the regulation of an apical, phospholipase C/Gq-coupled P2Y2receptor. We tested whether cortical actin filaments negatively regulate exocytosis in goblet cells by forming a barrier between secretory granules and plasma membrane docking sites as postulated for other secretory cells. Immunostaining of human lung tissues and SPOC1 cells (an epithelial, mucin-secreting cell line) revealed an apical distribution of β- and γ-actin in ciliated and goblet cells. In goblet cells, actin appeared as a prominent subplasmalemmal sheet lying between granules and the apical membrane, and it disappeared from SPOC1 cells activated by purinergic agonist. Disruption of actin filaments with latrunculin A stimulated SPOC1 cell mucin secretion under basal and agonist-activated conditions, whereas stabilization with jasplakinolide or overexpression of β- or γ-actin conjugated to yellow fluorescent protein (YFP) inhibited secretion. Myristoylated alanine-rich C kinase substrate, a PKC-activated actin-plasma membrane tethering protein, was phosphorylated after agonist stimulation, suggesting a translocation to the cytosol. Scinderin (or adseverin), a Ca2+-activated actin filament severing and capping protein was cloned from human airway and SPOC1 cells, and synthetic peptides corresponding to its actin-binding domains inhibited mucin secretion. We conclude that actin filaments negatively regulate mucin secretion basally in airway goblet cells and are dynamically remodeled in agonist-stimulated cells to promote exocytosis.

The heterotrimeric Gi(3) protein acts in slow but not in fast exocytosis of rat melanotrophs

1999 ◽  
Vol 112 (22) ◽  
pp. 4143-4150 ◽  
Author(s):  
M. Kreft ◽  
S. Gasman ◽  
S. Chasserot-Golaz ◽  
V. Kuster ◽  
M. Rupnik ◽  
...  
Keyword(s):  
Plasma Membrane ◽  
G Proteins ◽  
Flash Photolysis ◽  
Secretory Granules ◽  
Secretory Activity ◽  
Α Subunit ◽  
Capacitance Measurements ◽  
Regulated Secretion ◽  
Kinetic Component ◽  
Membrane Bound

Besides having a role in signal transduction some trimeric G-proteins may be involved in a late stage of exocytosis. Using immunocytochemistry and confocal microscopy we found that Gi(3)-protein resides mainly in the plasma membrane, whereas Gi(1/2-)protein is preferentially associated with secretory granules. To study the function of trimeric Gi(3)- and Gi(1/2)-proteins, secretory responses in single rat melanotrophs were monitored by patch-clamp membrane capacitance measurements. We report here that mastoparan, an activator of trimeric G-proteins, enhances calcium-induced secretory activity in rat melanotrophs. The introduction of synthetic peptides corresponding to the C-terminal domain of the (α)-subunit of Gi(3)- and Gi(1/2)-proteins indicated that Gi(3)peptide specifically blocked the mastoparan-stimulated secretory activity, which indicates an involvement of a trimeric Gi(3)-protein in mastoparan-stimulated secretory activity. Flash photolysis of caged Ca(2+)-elicited biphasic capacitance increases consisting of a fast and a slower component. Injection of anti-Gi(3) antibodies selectively inhibited the slow but not the fast component of secretory activity in rat melanotrophs. We propose that the plasma membrane-bound Gi(3)-protein may be involved in regulated secretion by specifically controlling the slower kinetic component of exocytosis.

Synaptotagmin-1: A Multi-Functional Protein that Mediates Vesicle Docking, Priming, and Fusion

2021 ◽  
Vol 22 ◽  
Author(s):  
Najeeb Ullah ◽  
Ezzouhra El Maaiden ◽  
Md. Sahab Uddin ◽  
Ghulam Md Ashraf
Keyword(s):  
Plasma Membrane ◽  
Secretory Granules ◽  
Secretory Vesicle ◽  
Neuroendocrine Cells ◽  
Snare Complex ◽  
Secretory Vesicles ◽  
Functional Protein ◽  
Vesicle Docking ◽  
Synaptotagmin 1

: The fusion of secretory vesicles with the plasma membrane depends on the assembly of v-SNAREs (VAMP2/synaptobrevin2) and t-SNAREs (SNAP25/syntaxin1) into the SNARE complex. Vesicles go through several upstream steps, referred to as docking and priming, to gain fusion competence. The vesicular protein synaptotagmin-1 (Syt-1) is the principal Ca2+ sensor for fusion in several central nervous system neurons and neuroendocrine cells and part of the docking complex for secretory granules. Syt-1 binds to the acceptor complex such as synaxin1, SNAP-25 on the plasma membrane to facilitate secretory vesicle docking, and upon Ca2+-influx promotes vesicle fusion. This review assesses the role of the Syt-1 protein involved in the secretory vesicle docking, priming, and fusion.

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