The use of permeabilized cells to investigate secretory granule biogenesis
| Autor: | Anirban Siddhanta, Wai Lam W. Ling, Dennis Shields |
|---|---|
| Rok vydání: | 1998 |
| Předmět: |
Cell Membrane Permeability
Cytological Techniques Golgi Apparatus Biology Cytoplasmic Granules General Biochemistry Genetics and Molecular Biology Exocytosis GTP-Binding Proteins Pituitary Gland Anterior Phospholipase D Phospholipase D activity Animals Secretion Phospholipid Transfer Proteins Molecular Biology ADP-Ribosylation Factors Vesicle Granule (cell biology) Membrane Proteins Secretory Vesicle Cell biology Rats Cytosol Secretory protein Growth Hormone Carrier Proteins Biogenesis |
| Zdroj: | Methods (San Diego, Calif.). 16(2) |
| ISSN: | 1046-2023 |
| Popis: | To investigate the mechanism of secretory granule biogenesis in endocrine cells, our laboratory used rat anterior pituitary GH3 cells which secrete growth hormone and prolactin. Here we describe a simple and rapid procedure for generating permeabilized cells to dissect molecular mechanisms involved in nascent secretory vesicle budding from the trans-Golgi network (TGN). Using this system, we demonstrate that vesicle budding is temperature, energy, and cytosol dependent; in addition, cytosol from a variety of cells, including yeast (Saccharomyces cerevisiae), can support vesicle release. The budding of nascent secretory vesicles from the TGN is stimulated by a phospholipase D activity that is associated with Golgi membranes. Our results suggest that phospholipid metabolism plays an important role in the release of nascent secretory vesicles from the TGN. |
| Databáze: | OpenAIRE |
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