scholarly journals Atg16L1, an essential factor for canonical autophagy, participates in hormone secretion from PC12 cells independently of autophagic activity

2012 ◽  
Vol 23 (16) ◽  
pp. 3193-3202 ◽  
Author(s):  
Koutaro Ishibashi ◽  
Takefumi Uemura ◽  
Satoshi Waguri ◽  
Mitsunori Fukuda
Keyword(s):  
Pc12 Cells ◽  
Secretory Pathway ◽  
Biological Activities ◽  
Hormone Secretion ◽  
Cell Types ◽  
Small Gtpase ◽  
Dense Core ◽  
Dense Core Vesicle ◽  
Dense Core Vesicles ◽  
Autophagic Activity

Autophagy is a bulk degradation system in all eukaryotic cells and regulates a variety of biological activities in higher eukaryotes. Recently involvement of autophagy in the regulation of the secretory pathway has also been reported, but the molecular mechanism linking autophagy with the secretory pathway remains largely unknown. Here we show that Atg16L1, an essential protein for canonical autophagy, is localized on hormone-containing dense-core vesicles in neuroendocrine PC12 cells and that knockdown of Atg16L1 causes a dramatic reduction in the level of hormone secretion independently of autophagic activity. We also find that Atg16L1 interacts with the small GTPase Rab33A and that this interaction is required for the dense-core vesicle localization of Atg16L1 in PC12 cells. Our findings indicate that Atg16L1 regulates not only autophagy in all cell types, but also secretion from dense-core vesicles, presumably by acting as a Rab33A effector, in particular cell types.

Copper modulates the large dense core vesicle secretory pathway in PC12 cells

Metallomics ◽  
2013 ◽  
Vol 5 (6) ◽  
pp. 700 ◽  
Author(s):  
Clare Duncan ◽  
Laura Bica ◽  
Peter J. Crouch ◽  
Aphrodite Caragounis ◽  
Grace E. Lidgerwood ◽  
...  
Keyword(s):  
Pc12 Cells ◽  
Secretory Pathway ◽  
Dense Core ◽  
Dense Core Vesicle ◽  
Large Dense Core Vesicle

scholarly journals The EARP Complex and its Interactor EIPR-1 are Required for Cargo Sorting to Dense-Core Vesicles

2016 ◽  
Author(s):  
Irini Topalidou ◽  
Jérôme Cattin-Ortolá ◽  
Andrea L. Pappas ◽  
Kirsten Cooper ◽  
Gennifer E. Merrihew ◽  
...  
Keyword(s):  
Peptide Hormones ◽  
Genetic Screen ◽  
Small Gtpase ◽  
Neuroendocrine Cells ◽  
Dense Core ◽  
Dense Core Vesicle ◽  
Dense Core Vesicles ◽  
Wide Range ◽  
Insulinoma Cells ◽  
Cargo Sorting

AbstractThe dense-core vesicle is a secretory organelle that mediates the regulated release of peptide hormones, growth factors, and biogenic amines. Dense-core vesicles originate from the trans-Golgi of neurons and neuroendocrine cells, but it is unclear how this specialized organelle is formed and acquires its specific cargos. To identify proteins that act in dense-core vesicle biogenesis, we performed a forward genetic screen in Caenorhabditis elegans for mutants defective in dense-core vesicle function. We previously reported the identification of two conserved proteins that interact with the small GTPase RAB-2 to control normal dense-core vesicle cargo-sorting. Here we identify several additional conserved factors important for dense-core vesicle cargo sorting: the WD40 domain protein EIPR-1 and the endosome-associated recycling protein (EARP) complex. By assaying behavior and the trafficking of dense-core vesicle cargos, we show that mutants that lack EIPR-1 or EARP have defects in dense-core vesicle cargo-sorting similar to those of mutants in the RAB-2 pathway. Genetic epistasis data indicate that RAB-2, EIPR-1 and EARP function in a common pathway. In addition, using a proteomic approach in rat insulinoma cells, we show that EIPR-1 physically interacts with the EARP complex. Our data suggest that EIPR-1 is a new component of the EARP complex and that dense-core vesicle cargo sorting depends on the EARP-dependent retrieval of cargo from an endosomal sorting compartment.Author SummaryAnimal cells package and store many important signaling molecules in specialized compartments called dense-core vesicles. Molecules stored in dense-core vesicles include peptide hormones like insulin and small molecule neurotransmitters like dopamine. Defects in the release of these compounds can lead to a wide range of metabolic and mental disorders in humans, including diabetes, depression, and drug addiction. However, it is not well understood how dense-core vesicles are formed in cells and package the appropriate molecules. Here we use a genetic screen in the microscopic worm C. elegans to identify proteins that are important for early steps in the generation of dense-core vesicles, such as packaging the correct molecular cargos in the vesicles. We identify several factors that are conserved between worms and humans and point to a new role for a protein complex that had previously been shown to be important for controlling trafficking in other cellular compartments. The identification of this complex suggests new cellular trafficking events that may be important for the generation of dense-core vesicles.

The small GTPase Cdc42 modulates the number of exocytosis-competent dense-core vesicles in PC12 cells

2012 ◽  
Vol 420 (2) ◽  
pp. 417-421 ◽  
Author(s):  
Mai Sato ◽  
Tetsuya Kitaguchi ◽  
Rika Numano ◽  
Kazuya Ikematsu ◽  
Masaki Kakeyama ◽  
...  
Keyword(s):  
Pc12 Cells ◽  
Small Gtpase ◽  
Dense Core ◽  
Dense Core Vesicles

scholarly journals Synaptotagmin VII Is Targeted to Dense-core Vesicles and Regulates Their Ca2+-dependent Exocytosis in PC12 Cells

2004 ◽  
Vol 279 (50) ◽  
pp. 52677-52684 ◽  
Author(s):  
Mitsunori Fukuda ◽  
Eiko Kanno ◽  
Megumi Satoh ◽  
Chika Saegusa ◽  
Akitsugu Yamamoto
Keyword(s):  
Plasma Membrane ◽  
Neuropeptide Y ◽  
Cell Line ◽  
Pc12 Cells ◽  
Pc12 Cell ◽  
Small Interfering Rna ◽  
Dense Core ◽  
Dense Core Vesicles ◽  
Pc12 Cell Line ◽  
Interfering Rna

It has recently been proposed that synaptotagmin (Syt) VII functions as a plasma membrane Ca2+sensor for dense-core vesicle exocytosis in PC12 cells based on the results of transient overexpression studies using green fluorescent protein (GFP)-tagged Syt VII; however, the precise subcellular localization of Syt VII is still a matter of controversy (plasma membraneversussecretory granules). In this study we established a PC12 cell line “stably expressing” the Syt VII-GFP molecule and demonstrated by immunocytochemical and immunoelectron microscopic analyses that the Syt VII-GFP protein is localized on dense-core vesicles as well as in other intracellular membranous structures, such as thetrans-Golgi network and lysosomes. Syt VII-GFP forms a complex with endogenous Syts I and IX, but not with Syt IV, and it colocalize well with Syts I and IX in the cellular processes (where dense-core vesicles are accumulated) in the PC12 cell line. We further demonstrated by an N-terminal antibody-uptake experiment that Syt VII-GFP-containing dense-core vesicles undergo Ca2+-dependent exocytosis, the same as endogenous Syt IX-containing vesicles. Moreover, silencing of Syt VII-GFP with specific small interfering RNA dramatically reduced high KCl-dependent neuropeptide Y secretion from the stable PC12 cell line (∼60% of the control cells), whereas the same small interfering RNA had little effect on neuropeptide Y secretion from the wild-type PC12 cells (∼85–90% of the control cells), indicating that the level of endogenous expression of Syt VII molecules must be low. Our results indicate that the targeting of Syt VII-GFP molecules to specific membrane compartment(s) is affected by the transfection method (transient expressionversusstable expression) and suggested that Syt VII molecule on dense-core vesicles functions as a vesicular Ca2+sensor for exocytosis in endocrine cells.

scholarly journals Role of the polybasic sequence in the Doc2α C2B domain in dense-core vesicle exocytosis in PC12 cells

2010 ◽  
pp. no-no ◽  
Author(s):  
Mai Sato ◽  
Yasunori Mori ◽  
Takahide Matsui ◽  
Ryo Aoki ◽  
Manami Oya ◽  
...  
Keyword(s):  
Pc12 Cells ◽  
Dense Core ◽  
Dense Core Vesicle ◽  
Vesicle Exocytosis

scholarly journals Preferential localization of a vesicular monoamine transporter to dense core vesicles in PC12 cells.

1994 ◽  
Vol 127 (5) ◽  
pp. 1419-1433 ◽  
Author(s):  
Y Liu ◽  
E S Schweitzer ◽  
M J Nirenberg ◽  
V M Pickel ◽  
C J Evans ◽  
...  
Keyword(s):  
Pc12 Cells ◽  
Gradient Centrifugation ◽  
Secretory Vesicle ◽  
Dense Core ◽  
Gamma Aminobutyric Acid ◽  
Dense Core Vesicles ◽  
Protein Marker ◽  
Preferential Localization ◽  
Large Dense Core Vesicles ◽  
Pheochromocytoma Pc12

Neurons and endocrine cells have two types of secretory vesicle that undergo regulated exocytosis. Large dense core vesicles (LDCVs) store neural peptides whereas small clear synaptic vesicles store classical neurotransmitters such as acetylcholine, gamma-aminobutyric acid (GABA), glycine, and glutamate. However, monoamines differ from other classical transmitters and have been reported to appear in both LDCVs and smaller vesicles. To localize the transporter that packages monoamines into secretory vesicles, we have raised antibodies to a COOH-terminal sequence from the vesicular amine transporter expressed in the adrenal gland (VMAT1). Like synaptic vesicle proteins, the transporter occurs in endosomes of transfected CHO cells, accounting for the observed vesicular transport activity. In rat pheochromocytoma PC12 cells, the transporter occurs principally in LDCVs by both immunofluorescence and density gradient centrifugation. Synaptic-like microvesicles in PC12 cells contain relatively little VMAT1. The results appear to account for the storage of monoamines by LDCVs in the adrenal medulla and indicate that VMAT1 provides a novel membrane protein marker unique to LDCVs.

Silence of Synaptotagmin VII inhibits release of dense core vesicles in PC12 cells

2009 ◽  
Vol 52 (12) ◽  
pp. 1156-1163 ◽  
Author(s):  
JiangLi Li ◽  
Yang Xiao ◽  
Wei Zhou ◽  
ZhengXing Wu ◽  
RongYing Zhang ◽  
...  
Keyword(s):  
Pc12 Cells ◽  
Dense Core ◽  
Dense Core Vesicles ◽  
Synaptotagmin Vii
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