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.

Selective inactivation of guanine-nucleotide-binding regulatory protein (G-protein) α and βγ subunits by urea

2001 ◽  
Vol 354 (2) ◽  
pp. 337-344 ◽  
Author(s):  
William K. LIM ◽  
Richard R. NEUBIG
Keyword(s):  
G Protein ◽  
G Proteins ◽  
Regulatory Protein ◽  
Significant Loss ◽  
Membrane Extraction ◽  
Α Subunit ◽  
Mammalian Cell Lines ◽  
Membrane Bound ◽  
Guanine Nucleotide Binding ◽  
G Protein Coupled

G-protein-coupled receptors activate signal-transducing G-proteins, which consist of an α subunit and a βγ dimer. Membrane extraction with 5–7M urea has been used to uncouple receptors from endogenous G-proteins to permit reconstitution with purified G-proteins. We show that αi subunits are inactivated with 5M urea whereas the βγ dimer requires at least 7M urea for its inactivation. There is no significant loss of receptors. Surprisingly, Western-blot analysis indicates that the urea-denatured αi subunit remains mostly membrane-bound and that β is only partially removed. After 7M urea treatment, both αi1 and βγ subunits are required to restore high-affinity agonist binding and receptor-catalysed guanosine 5′-[γ-thio]triphosphate binding. We demonstrate the generality of this approach for four Gi-coupled receptors (α2A-adrenergic, adenosine A1, 5-hydroxytryptamine1A and µ-opioid) expressed in insect cells and two mammalian cell lines. Thus a selectivity of urea for G-protein α versus βγ subunits is established in both concentration and mechanism.

Time–resolved capacitance measurements: monitoring exocytosis in single cells

1991 ◽  
Vol 24 (1) ◽  
pp. 75-101 ◽  
Author(s):  
Manfred Lindau
Keyword(s):  
Plasma Membrane ◽  
Mast Cells ◽  
Endocrine Cells ◽  
Secretory Granules ◽  
Single Cells ◽  
Time Resolved ◽  
Capacitance Measurements ◽  
Fusion Pores

Many cells release preformed material contained in secretory granules by exocytosis. Exocytosis is a specialized means of secretion in which the granules fuse with the plasma membrane and thereby discharge their contents through the fusion pores. This mechanism mediates, for example, the formation of the fertilization envelope in eggs, the release of neurotransmitters and neuropeptides by neurons, the release of a variety of enzymes and mediators by mast cells and granulocytes or the secretion of hormones by endocrine cells. Classical methods for investigating exocytosis usually measure release of secreted material.

Functional Organization of the Porosome Complex and Associated Structures Facilitating Cellular Secretion

Physiology ◽  
2009 ◽  
Vol 24 (6) ◽  
pp. 367-376 ◽  
Author(s):  
Bhanu P. Jena
Keyword(s):  
Plasma Membrane ◽  
Functional Organization ◽  
Secretory Activity ◽  
Human Platelets ◽  
Frog Neuromuscular Junction ◽  
Cell Secretion ◽  
Cell Plasma Membrane ◽  
Cell Plasma ◽  
Membrane Bound ◽  
Cellular Secretion

Porosomes, the universal secretory machinery at the cell plasma membrane, are cup-shaped supramolecular lipoprotein structures, where membrane-bound vesicles transiently dock and fuse to release intravesicular contents during cell secretion. In this review, the discovery of the porosome and its structure, dynamics, composition, and functional reconstitution are outlined. Furthermore, the architecture of porosome-like structures such as the “canaliculi system” in human platelets and various associated structures such as the T-bars at the Drosophila synapse or the “beams,” “ribs,” and “pegs” at the frog neuromuscular junction, each organized to facilitate a certain specialized secretory activity, are briefly discussed.

Ultrastructural Aspects of Golgi Complex Function in Parathyroid Cells of Winter Frogs

1973 ◽  
Vol 31 ◽  
pp. 680-681
Author(s):  
William J. Dougherty
Keyword(s):  
Golgi Complex ◽  
Secretory Granules ◽  
Complex Function ◽  
Secretory Activity ◽  
Secretory Proteins ◽  
Perivascular Spaces ◽  
Pituitary Cells ◽  
Electron Microscopic ◽  
Ca Ions ◽  
Regulation Of Secretion

The regulation of secretion in exocrine and endocrine cells has long been of interest. Electron microscopic and other studies have demonstrated that secretory proteins synthesized on ribosomes are transported by the rough ER to the Golgi complex where they are concentrated into secretory granules. During active secretion, secretory granules fuse with the cell membrane, liberating and discharging their contents into the perivascular spaces. When secretory activity is suppressed in anterior pituitary cells, undischarged secretory granules may be degraded by lysosomes. In the parathyroid gland, evidence indicates that the level of blood Ca ions regulates both the production and release of parathormone. Thus, when serum Ca is low, synthesis and release of parathormone are both stimulated; when serum Ca is elevated, these processes are inhibited.

Analysis of the Anterior Secretory Cells of Onchidohs Muricata (Nudibranchia)

1981 ◽  
Vol 39 ◽  
pp. 614-615
Author(s):  
Roy Skidmore
Keyword(s):  
Endoplasmic Reticulum ◽  
Secretory Granules ◽  
Golgi Body ◽  
The Body ◽  
Secretory Cells ◽  
Secretory Vesicles ◽  
High Magnification ◽  
Large Numbers ◽  
Membrane Bound ◽  
Sole Of The Foot

The long-necked secretory cells in Onchidoris muricata are distributed in the anterior sole of the foot. These cells are interspersed among ciliated columnar and conical cells as well as short-necked secretory gland cells. The long-necked cells contribute a significant amount of mucoid materials to the slime on which the nudibranch travels. The body of these cells is found in the subepidermal tissues. A long process extends across the basal lamina and in between cells of the epidermis to the surface of the foot. The secretory granules travel along the process and their contents are expelled by exocytosis at the foot surface.The contents of the cell body include the nucleus, some endoplasmic reticulum, and an extensive Golgi body with large numbers of secretory vesicles (Fig. 1). The secretory vesicles are membrane bound and contain a fibrillar matrix. At high magnification the similarity of the contents in the Golgi saccules and the secretory vesicles becomes apparent (Fig. 2).

Exterior and Interior Events in Early Stage of Thrombin-induced Platelet Aggregation

1977 ◽  
Vol 38 (03) ◽  
pp. 0630-0639 ◽  
Author(s):  
Shuichi Hashimoto ◽  
Sachiko Shibata ◽  
Bonro Kobayashi
Keyword(s):  
Molecular Weight ◽  
Enzyme Activity ◽  
Plasma Membrane ◽  
Platelet Aggregation ◽  
Early Stage ◽  
Adenyl Cyclase ◽  
Cellular Component ◽  
Primary Event ◽  
Rabbit Platelets ◽  
Membrane Bound

SummaryTreatment of washed rabbit platelets with 1 u/ml of thrombin at 37° C resulted in a disappearance from platelets of a protein with 250,000 dalton molecular weight which was shown to be originated from plasma membrane. Parallel loss of adenyl cyclase was noted, and both reactions were complete within 30 sec. From the patterns of disc electrophoretograms, the importance of quick suppression of thrombin action in demonstrating the primary event was stressed.Thrombin induced an apparent activation of membrane bound phosphodiesterase. This reaction was also complete within 30 sec. The cellular component which contained the enzyme activity was distinct from plasma membrane. Soluble phosphodiesterase was not influenced by thrombin at all.These reactions required intact platelet cells to react with thrombin, and no reaction was detected when subcellular preparations were treated with thrombin.Possibility of collaboration of changes in externally located synthetic enzyme with those in internally located degrading enzyme in the early phase of thrombin action on platelets was suggested.

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