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PMID: 1599416 Published · ppublish English Journal Article Research Support, Non-U.S. Gov't

Exocytosis from permeabilized bovine adrenal chromaffin cells is differently modulated by guanosine 5'-[gamma-thio]triphosphate and guanosine 5'-[beta gamma-imido]triphosphate. Evidence for the involvement of various guanine nucleotide-binding proteins.

The Biochemical journal ·Vol. 284 ( Pt 2) ·1992-06-01 ·Pages 321-6

Ahnert-Hilger G, Wegenhorst U, Stecher B, Spicher K, Rosenthal W, Gratz M

Abstract

1. In bovine adrenal chromaffin cells made permeable either to molecules less than or equal to 3 kDa with alphatoxin or to proteins less than or equal to 150 kDa with streptolysin O, the GTP analogues guanosine 5'-[beta gamma-imido]triphosphate (p[NH]ppG) and guanosine 5'-[gamma-thio]triphosphate (GTP[S]) differently modulated Ca(2+)-stimulated exocytosis. 2. In alphatoxin-permeabilized cells, p[NH]ppG up to 20 microM activated Ca(2+)-stimulated exocytosis. Higher concentrations had little or no effect. At a free Ca2+ concentration of 5 microM, 7 microM-p[NH]ppG stimulated exocytosis 6-fold. Increasing the free Ca2+ concentration reduced the effect of p[NH]ppG. Pretreatment of the cells with pertussis toxin prevented the activation of the Ca(2+)-stimulated exocytosis by p[NH]ppG. 3. In streptolysin O-permeabilized cells, p[NH]ppG did not activate, but rather inhibited Ca(2+)-dependent catecholamine release under all conditions studied. In the soluble cytoplasmic material that escaped during permeabilization with streptolysin O, different G-protein alpha-subunits were detected using an appropriate antibody. Around 15% of the cellular alpha-subunits were detected in the supernatant of permeabilized control cells. p[NH]ppG or GTP[S] stimulated the release of alpha-subunits 2-fold, causing a loss of about 30% of the cellular G-protein alpha-subunits under these conditions. Two of the alpha-subunits in the supernatant belonged to the G(o) type, as revealed by an antibody specific for G(o) alpha. 4. GTP[S], when present alone during stimulation with Ca2+, activated exocytosis in a similar manner to p[NH]ppG. Upon prolonged incubation, GTP[S], in contrast to p[NH]ppG, inhibited Ca(2+)-induced exocytosis from cells permeabilized by either of the pore-forming toxins. This effect was resistant to pertussin toxin. 5. The p[NH]ppG-induced activation of Ca(2+)-stimulated release from alphatoxin-permeabilized chromaffin cells may be attributed to one of the heterotrimeric G-proteins lost during permeabilization with streptolysin O. The inhibitory effect of GTP[S] on exocytosis is apparently not mediated by G-protein alpha-subunits, but by another GTP-dependent process still occurring after permeabilization with streptolysin O.

MeSH Terms
Adrenal Glands/cytology,drug effects,metabolism Animals Bacterial Toxins/pharmacology Calcium/metabolism Cattle Cell Membrane Permeability/drug effects Cells, Cultured Chromaffin Granules/drug effects,metabolism Electrophoresis, Polyacrylamide Gel Exocytosis GTP-Binding Proteins/metabolism Guanosine 5'-O-(3-Thiotriphosphate)/pharmacology Guanylyl Imidodiphosphate/analogs & derivatives,pharmacology Hemolysin Proteins Streptolysins/pharmacology
Chemicals
Bacterial Toxins Hemolysin Proteins Streptolysins staphylococcal alpha-toxin Guanylyl Imidodiphosphate Guanosine 5'-O-(3-Thiotriphosphate) GTP-Binding Proteins Calcium
Authors & Affiliations
6 authors, click to expand affiliations / ORCID
Ahnert-Hilger G
Institut für Neuropsychopharmacologie, Freien Universität Berlin, Germany.
Wegenhorst U
Stecher B
Spicher K
Rosenthal W
Gratz M
References (42)
42 references, click to expand
  1. A pertussis-toxin-sensitive protein controls exocytosis in chromaffin cells at a step distal to the generation of second messengers.
    Biochem J. 1991 Mar 1;274 ( Pt 2):339-47 PMID: 1848752
  2. The phorbol ester TPA increases the affinity of exocytosis for calcium in 'leaky' adrenal medullary cells.
    FEBS Lett. 1983 Aug 22;160(1-2):98-100 PMID: 6884518
  3. Essential synergy between Ca2+ and guanine nucleotides in exocytotic secretion from permeabilized rat mast cells.
    J Cell Biol. 1987 Jul;105(1):191-7 PMID: 2440894
  4. Detection of low molecular mass GTP-binding proteins in chromaffin granules and other subcellular fractions of chromaffin cells.
    FEBS Lett. 1989 Apr 10;247(1):127-31 PMID: 2495990
  5. Galanin inhibits insulin secretion by direct interference with exocytosis.
    FEBS Lett. 1989 Apr 24;247(2):401-4 PMID: 2469602
  6. Pertussis toxin induces structural changes in G alpha proteins independently of ADP-ribosylation.
    Proc Natl Acad Sci U S A. 1989 Apr;86(8):2577-81 PMID: 2523074
  7. Low molecular weight GTP-binding proteins are associated with neuronal organelles involved in rapid axonal transport and exocytosis.
    J Biol Chem. 1989 Nov 5;264(31):18363-7 PMID: 2478544
  8. A role for GTP-binding proteins in vesicular transport through the Golgi complex.
    Soc Gen Physiol Ser. 1989;44:175-88 PMID: 2506649
  9. A T cell receptor-associated GTP-binding protein triggers T cell receptor-mediated granule exocytosis in cytotoxic T lymphocytes.
    J Immunol. 1988 Dec 1;141(11):3785-90 PMID: 3141505
  10. Low molecular mass GTP-binding proteins of adrenal chromaffin cells are present on the secretory granule.
    FEBS Lett. 1989 Mar 13;245(1-2):122-6 PMID: 2494070
  11. Presence of three pertussis toxin substrates and Go alpha immunoreactivity in both plasma and granule membranes of chromaffin cells.
    FEBS Lett. 1987 May 11;215(2):339-44 PMID: 3108031
  12. Nucleotide and deduced amino acid sequences of a GTP-binding protein family with molecular weights of 25,000 from bovine brain.
    J Biol Chem. 1988 Aug 15;263(23):11071-4 PMID: 3136152
  13. A GTP-binding protein required for secretion rapidly associates with secretory vesicles and the plasma membrane in yeast.
    Cell. 1988 Jun 3;53(5):753-68 PMID: 3131018
  14. Do GTPases direct membrane traffic in secretion?
    Cell. 1988 Jun 3;53(5):669-71 PMID: 2836065
  15. Inactivation of a T cell receptor-associated GTP-binding protein by antibody-induced modulation of the T cell receptor/CD3 complex.
    J Exp Med. 1988 Aug 1;168(2):817-22 PMID: 2970522
  16. Adipocyte plasma membranes contain two Gi subtypes but are devoid of Go.
    FEBS Lett. 1988 Sep 26;238(1):191-6 PMID: 3139455
  17. Guanine nucleotides and Ca-dependent exocytosis. Studies on two adrenal cell preparations.
    FEBS Lett. 1985 Sep 23;189(2):345-9 PMID: 3930291
  18. Identification and characterization of the 35-kDa beta subunit of guanine-nucleotide-binding proteins by an antiserum raised against transducin.
    Eur J Biochem. 1986 Jul 15;158(2):255-63 PMID: 3089781
  19. Conformations of the alpha 39, alpha 41, and beta.gamma components of brain guanine nucleotide-binding proteins. Analysis by limited proteolysis.
    J Biol Chem. 1986 Jun 5;261(16):7571-9 PMID: 2423523
  20. Guanine nucleotides induce Ca2+-independent insulin secretion from permeabilized RINm5F cells.
    J Biol Chem. 1987 Apr 15;262(11):5049-56 PMID: 3031036
  21. Two roles for guanine nucleotides in the stimulus-secretion sequence of neutrophils.
    Nature. 1986 Feb 6-12;319(6053):504-7 PMID: 3003581
  22. GTP-dependent inhibition of insulin secretion by epinephrine in permeabilized RINm5F cells. Lack of correlation between insulin secretion and cyclic AMP levels.
    J Biol Chem. 1988 Jun 25;263(18):8615-20 PMID: 2837460
  23. Guanine nucleotide effects on catecholamine secretion from digitonin-permeabilized adrenal chromaffin cells.
    J Biol Chem. 1986 Aug 5;261(22):10182-8 PMID: 3015921
  24. The dynamics of exocytosis in human neutrophils.
    J Cell Biol. 1988 Dec;107(6 Pt 1):2117-23 PMID: 3058717
  25. Dissection of a single round of vesicular transport: sequential intermediates for intercisternal movement in the Golgi stack.
    Cell. 1989 Feb 10;56(3):357-68 PMID: 2536591
  26. A reassessment of guanine nucleotide effects on catecholamine secretion from permeabilized adrenal chromaffin cells.
    J Biol Chem. 1989 Oct 5;264(28):16426-34 PMID: 2674132
  27. Poration by alpha-toxin and streptolysin O: an approach to analyze intracellular processes.
    Methods Cell Biol. 1989;31:63-90 PMID: 2779453
  28. Reductive chain separation of botulinum A toxin--a prerequisite to its inhibitory action on exocytosis in chromaffin cells.
    FEBS Lett. 1989 May 8;248(1-2):23-7 PMID: 2721679
  29. Introduction of macromolecules into bovine adrenal medullary chromaffin cells and rat pheochromocytoma cells (PC12) by permeabilization with streptolysin O: inhibitory effect of tetanus toxin on catecholamine secretion.
    J Neurochem. 1989 Jun;52(6):1751-8 PMID: 2723634
  30. Adrenal medullary chromaffin cells in vitro.
    Physiol Rev. 1984 Oct;64(4):1103-61 PMID: 6208567
  31. Ca2+-stimulated catecholamine release from alpha-toxin-permeabilized PC12 cells: biochemical evidence for exocytosis and its modulation by protein kinase C and G proteins.
    Biochemistry. 1987 Dec 1;26(24):7842-8 PMID: 3322408
  32. The dual effector system for exocytosis in mast cells: obligatory requirement for both Ca2+ and GTP.
    Biosci Rep. 1987 May;7(5):369-81 PMID: 3315030
  33. Damage to mammalian cells by proteins that form transmembrane pores.
    Rev Physiol Biochem Pharmacol. 1987;107:147-223 PMID: 3303271
  34. Capacitance measurements reveal stepwise fusion events in degranulating mast cells.
    Nature. 1984 Nov 29-Dec 5;312(5993):453-5 PMID: 6504157
  35. Isolation and identification of two hemolytic forms of streptolysin-O.
    Infect Immun. 1984 Nov;46(2):394-400 PMID: 6500696
  36. Catecholamine secretion from digitonin-treated PC12 cells. Effects of Ca2+, ATP, and protein kinase C activators.
    J Biol Chem. 1986 Nov 5;261(31):14665-9 PMID: 3490477
  37. Purification of alpha-toxin from Staphylococcus aureus and application to cell permeabilization.
    Anal Biochem. 1987 Jul;164(1):84-9 PMID: 3674374
  38. Intracellular neutral carrier-based Ca2+ microelectrode with subnanomolar detection limit.
    Pflugers Arch. 1987 Jul;409(3):223-8 PMID: 3627942
  39. Hydrolysis-resistant GTP analogs stimulate catecholamine release from digitonin-permeabilized PC12 cells.
    J Neurochem. 1990 Sep;55(3):930-6 PMID: 2117052
  40. Agonist-sensitive binding of a photoreactive GTP analog to a G-protein alpha-subunit in membranes of HL-60 cells.
    FEBS Lett. 1990 Jan 15;260(1):14-8 PMID: 2153575
  41. Stimulation of Ca2(+)-independent catecholamine secretion from digitonin-permeabilized bovine adrenal chromaffin cells by guanine nucleotide analogues. Relationship to arachidonate release.
    Biochem J. 1990 Jul 15;269(2):521-6 PMID: 2117444
  42. A major role for protein kinase C in calcium-activated exocytosis in permeabilised adrenal chromaffin cells.
    FEBS Lett. 1988 Sep 26;238(1):151-5 PMID: 3169247
Article Info
Journal
The Biochemical journal
Abbr.
Biochem J
ISSN
0264-6021
DOI
10.1042/bj2840321
Published
1992-06-01
Pages
321-6
Language
English
Region
England
NLM ID
2984726R
PMCID
PMC1132640
Subset
IM
External Links
PubMed source DOI: 10.1042/bj2840321 PMC Full Text View journal details
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