Research in Experimental Medicine

, Volume 193, Issue 1, pp 323–335 | Cite as

Effects of guanine nucleotides on bombesin-stimulated signal transduction in rat pancreatic acinar cells

  • Albrecht Piiper
  • Danuta Stryjek-Kaminska
  • Jürgen Stein
  • Wolfgang F. Caspary
  • Stefan Zeuzem
Original Papers
Received:
Accepted:

Abstract

To study the role of guanine nucleotide binding proteins (G proteins) in bombesin receptor signal transduction, we investigated the effects of guenine nucleotide analogues and of the G protein activator NaF on bombesin-induced amylase release, inositol 1,4,5-trisphosphate (Ins(1,4,5)P 3) production and release of intracellular Ca2+ in rat pancreatic acini. In digitonin-permeabilized acini, guanosine 5′-[γ-thio]triphosphate (GTPγS), a well-known activator of G proteins, potentiated bombesin-induced Ins(1,4,5)P 3 production and increased amylase release at low bombesin concentrations (<10 nM). By contrast, GTPγS decreased bombesin-stimulated amylase release at high bombesin concentrations (>10 nM). Fluoride (10 mM), another G protein activator, had similar effects to GTPγS on amylase release. However, unlike GTPγS it had no effect on Ins(1,4,5)P 3 production and release of intracellular Ca2+ induced by high bombesin concentrations. GDP and its analogues, such as 2′-desoxyguanosine 5′-diphosphate (dGDP) or guanosine 5′-[β-thio]diphosphate (GDPβS), inhibit activation of G proteins. GDP and dGDP both inhibited amylase release and Ins(1,4,5)P 3 production at all bombesin concentrations tested. In contrast, GDPβS mimicked the effects of GTPγS on bombesin-stimulated amylase release and Ins(1,4,5)P 3 accumulation. In conclusion, we suggest that bombesin receptor-mediated signal transduction involves G proteins in pancreatic acini. The correlation between inhibition of maximum-stimulated enzyme secretion and further increase in Ins(1,4,5)P 3 production in response to GTPγS at high bombesin concentrations suggests that overstimulation of phospholipase C inhibits amylase release. The discrepant effects of GDP and of GDPβS on phospholipase C activity and amylase release might be due to the ability of GDPβS, but not of GDP to activate G proteins persistently after phosphorylation by G protein-associated GDP kinases.

Key words

Amylase secretion Calcium Fluoride G protein Inositol 1,4,5-trisphosphate 

Abbreviations

[Ca2]i

intracellular free Ca2+ concentration

CCK

cholecystokinin

CCK-8

cholecytokinin-octapeptide

G protein

trimeric guanine-nucleotide-binding protein

Gs and Gi

stimulatory and inhibitory G proteins of adenyl cyclase, respectively

Gp

Gprotein of phospholipase C

GDPβS

guanosine 5′-[β-thio]diphosphate

dGDP

2′-desoxyguanosine 5′-diphosphate

GTPγS

guanosine 5′-[γ-thio]triphosphate

GppNHp

guanosine 5′-[β, γ-imdo]diphosphate

Ins(1,4,5)P3

inositol 1,4,5-triphosphate

PtdInsP2

phosphatidylinositiol 4,5-bisphosphate

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References

  1. 1.
    Battery JF, Way JM, Corjay MH, Shapira H, Kusano K, Harkins R, Wu JM, Slattery T, Mann E, Feldman RI (1991) Molecular cloning of the bombesin/gastrin-releasing peptide receptor from Swiss 3T3 cells. Proc Natl Acad Sci USA 88: 395–399CrossRefGoogle Scholar
  2. 2.
    Bigay J, Deterre P, Pfister C, Chabre M (1985) Fluoroaluminates activate transducin-GDP by mimicking the gamma phosphate of GTP in its binding site. FEBS Lett 191: 181–185PubMedCrossRefGoogle Scholar
  3. 3.
    Birnbaumer L, Abramowitz J, Brown AM (1990) Receptor-effector coupling by G-proteins. Biochim Biophys Acta 1031: 163–224PubMedGoogle Scholar
  4. 4.
    Bizzarri C, DiGirolamo M, D'Oracio MC, Corda D (1990) Evidence that a guanine nucleotide-binding protein linked to a muscarinic receptor inhibits directly phospholipase C. Proc Natl Acad Sci USA 87: 4889–4893PubMedCrossRefGoogle Scholar
  5. 5.
    Bradford MM (1976) A rapid and sensitive method for the quantitation of microgram quantities of protein utilizing the principle of protein dye binding. Anal Biochem 72: 248–254PubMedCrossRefGoogle Scholar
  6. 6.
    Corjay MJ, Dobrzanski JD, Way JM, Viallet J, Shapira H, Worland P, Sausville EA, Battey JF (1991) Two distinct bombesin receptor subtypes are expressed and functional in human lung carcinoma cells. J Biol Chem 266: 18771–18779PubMedGoogle Scholar
  7. 7.
    Fischer JB, Schonbrunn A (1988) The bombesin receptor is coupled to a guanine nucleotide-binding protein protein which is insensitive to pertussis and cholera toxin. J Biol Chem 263: 2808–2816PubMedGoogle Scholar
  8. 8.
    Gaisano HY, Miller LJ (1992) Complex role of protein kinase C in mediating the supramaximal inhibition of pancreatic secretion observed with cholecystokinin. Biochem Biophys Res Commun 187: 498–506PubMedCrossRefGoogle Scholar
  9. 9.
    Galas MC, Lignon MF, Rodriguez M, Mendre C, Fulcrand P, Laur J, Martinez J (1988) Structure-activity relationship studies on cholecystokinin-analogues with partial agonist activity. Am J Physiol 254: G176-G182PubMedGoogle Scholar
  10. 10.
    Garcia JGN, Dominguez J, English D (1991) Sodium fluoride induces phosphoinositide hydrolysis, Ca2+ mobilization, and prostacyclin synthesis in cultured human endothelium: further evidence for regulation by a pertussis toxin-insensitive guanine nucleotide-binding protein. Am J Respir Cell Mol Biol 5: 113–124PubMedGoogle Scholar
  11. 11.
    Gardner JD, Jackson MJ (1977) Regulation of amylase release from dispersed pancreatic acinar cells. J Physiol (Lond) 270: 439–454Google Scholar
  12. 12.
    Gilman AG (1987) G-proteins: signal transducers of receptor-generated signals. Annu Rev Biochem 56: 615–649PubMedCrossRefGoogle Scholar
  13. 13.
    Grynkiewicz G, Poenie M, Tsien RY (1985) A new generation of Ca2+ indicators with greatly improved fluorescence properties. J Biol Chem 260: 3440–3450PubMedGoogle Scholar
  14. 14.
    Gutowski S, Smrcka A, Nowak L, Wu D, Simon M, Sternweis PC (1991) Antibodies to the αq subfamily of guanine nucleotide-binding regulatory proteins: α subunits attenuate activation of phosphatidylinositol 4,5-bisphophate hydrolysis by hormones. J Biol Chem 266: 20519–20524PubMedGoogle Scholar
  15. 15.
    Jensen RT, Moody T, Pert C, Rivier JE, Garnder JD (1978) Interaction of bombesin and litorin with specific membrane receptors on pancreatic acinar cells. Proc Natl Acad Sci USA 75: 6139–6343PubMedCrossRefGoogle Scholar
  16. 16.
    Kimura T, Imamura K, Eckhardt L, Schulz I (1986) Ca2+, phorbol ester, and cAMP-stimulated enzyme secretion from permeabilized rat pancreatic acini. Am J Physiol 250: G698-G708PubMedGoogle Scholar
  17. 17.
    Matozaki T, Sakamoto C, Nagao M, Nishizaki H, Baba S (1988) G-proteins in stimulation of PI hydrolysis by CCK in isolated rat pancreatic acinar cells. Am J Physiol 255: E652-E659PubMedGoogle Scholar
  18. 18.
    Matozaki T, Göke B, Tsunoda Y, Rodriguez M, Martinez, Williams JA (1990) Two functionally distinct cholecystokinin receptors show different modes of action on Ca++ mobilization and phospholipid hydrolysis in isolated rat pancreatic acini. J Biol Chem 265: 6247–6254PubMedGoogle Scholar
  19. 19.
    Matozaki T, Zhu WY, Tsunoda Y, Göke B, Williams JA (1991) Intracellular mediators of bombesin action on rat pancreatic acinar cells. Am J Physiol 260: G858-G864PubMedGoogle Scholar
  20. 20.
    Menozzi D, Sato S, Jensen RT, Gardner JD (1989) Cyclic GMP does not inhibit protein kinase C-mediated enzyme secretion in rat pancreatic acini. J Biol Chem 264: 995–999PubMedGoogle Scholar
  21. 21.
    Merritt JE, Taylor CW, Rubin RP, Putney JW (1986) Evidence suggesting that a novel guanine nucleotide regulatory binding protein couples receptors to phospholipase C in exocrine pancreas. Biochem J 236: 337–343PubMedGoogle Scholar
  22. 22.
    Mössner J, Secknus R, Spiekermann GM, Sommer C, Biernat M, Bahnsen H, Fischbach W (1991) Prostaglandin E2 inhibits secretagogue-induced enzyme secretion from rat pancreatic acini. Am J Physiol 260: G711-G719PubMedGoogle Scholar
  23. 23.
    Oberdisse E, Lapetina EG (1987) GDPβS enhances the activation of phospholipase C caused by thrombin in human plastelets: evidence for involvement of an inhibitory GTP-caused by thrombin in human platelets: evidence for involvement of an inhibitory GTP-binding protein. Biochem Biophys Res Commun 144: 1188–1196PubMedCrossRefGoogle Scholar
  24. 24.
    Ohtsuki K, Ikeuchi T, Yokoyama M (1986) Characterization of nucleoside diphosphate (NDP)-kinase associated GTP binding proteins from human recombinant interleukin 2 (rIL-2)-treated mouse NK cells. Biochim Biophys Acta 882: 322–330PubMedGoogle Scholar
  25. 25.
    Padfield PJ, Ding TG, Jamieson DJ (1991) Ca2+-dependent amylase secretion from pancreatic acinar cells occurs without activation of phospholipase C linked G-proteins. Biochem Biophys Res Commun 174: 536–541PubMedCrossRefGoogle Scholar
  26. 26.
    Pandol SJ, Schoeffield MS (1986) I,2-Diacylgycerol, protein kinase C and pancreatic enzyme secretion. J Biol Chem 261: 4438–4444PubMedGoogle Scholar
  27. 27.
    Paris S, Pouyssegur J (1990) Guanosine 5′-O-(3-thiotriphosphate) and guanosine 5′-O-2-thiodiphosphate) activate G proteins and potentiate fibroblast growth factor-induced DNA synthesis in hamster fibroblasts. J Biol Chem 265: 11567–11575PubMedGoogle Scholar
  28. 28.
    Piiper A, Plusczyk T, Eckhardt L, Schulz I (1991) Effects of cholecystokinin, cholecystokinin JMV-180 and GTP analogs on enzyme secretion from permeabilized acini and chloride conductance in isolated zymogen granules of the rat pancreas. Eur J Biochem 197: 391–398PubMedCrossRefGoogle Scholar
  29. 29.
    Pröfrock A, Zimmermann P, Schulz I (1992) Bombesin receptors interact with Gi and p21ras proteins in plasma membranes from rat pancreatic acinar cells. Am J Physiol 263: G240-G247PubMedGoogle Scholar
  30. 30.
    Rothman JE, Orci L (1992) Molecular dissection of the secretory pathway. Nature 335: 409–415CrossRefGoogle Scholar
  31. 31.
    Rowley WH, Sato S, Collado-Escobar DM, Huang SC, Martinez J, Gardner JD, Jensen RT (1990) Cholecystokinin-induced formation of inositol phosphates in pancreatic acini. Am J Physiol 259: G655-G665PubMedGoogle Scholar
  32. 32.
    Schnefel S, Pröfrock A, Hinsch KD, Schulz I (1990) Cholecystokin activates Gi1-, Gi2-, Gi3- and several Gs-proteins in rat pancreatic acinar cells. Biochem J 269: 483–488PubMedGoogle Scholar
  33. 33.
    Schulz I (1989) Signaling transduction in hormone and neurotransmitter-induced enzyme secretion from the exocrine pancreas. In: Schulz SG, Forte FG, Rauner BB (eds) Handbook of physiology, vol 3: The gastrointestinal system. American Physiological Society, Bethesda, Md, pp 443–463Google Scholar
  34. 34.
    Seifert R, Rosenthal W, Schultz G, Wieland T, Gierschick P, Jakobs KH (1988) The role of nucleoside-diphosphate kinase in G protein activation of NADPH oxidase by guanine and adenine nucleotides. Eur J Biochem 175: 51–55PubMedCrossRefGoogle Scholar
  35. 35.
    Sekar MC, Uemura N, Coy D, Hirschowitz BI, Dickinson KEJ (1991) Bombesin, neuromedin B and neuromedin C interact with a common rat pancreatic phosphoinositidecoupled receptor, but are differently regulated by guanine nucleotides. Biochem J 280: 163–169PubMedGoogle Scholar
  36. 36.
    Sternweis PC, Gilman G (1982) Aluminium: a requirement for activation of the regulatory component of adenylate cyclase by fluoride. Proc Natl Acad Sci USA 79: 4888–4891PubMedCrossRefGoogle Scholar
  37. 37.
    Taylor CW, Merritt JE, Putney JW Jr, Rubin RP (1986) Effects of Ca2+ on phosphoinositide breakdown in exocrine pancreas. Biochem J 238: 765–772PubMedGoogle Scholar
  38. 38.
    Taylor SJ, Chae HZ, Rhee SG, Exton JH (1991) Activation of the β1 isoenzyme of phospholipase C by α subunits of the Gq class G-proteins. Nature 350: 516–518PubMedCrossRefGoogle Scholar
  39. 39.
    Uhlemann ER, Rottman AJ, Gardner JD (1979) Actions of peptides isolated from amphibian skin on amylase release from dispersed pancreatic acini. Am J Physiol 258: E571-E576Google Scholar
  40. 40.
    Von zur Mühlen F, Eckstein F, Penner R (1991) Guanosine 5′-[β-thio]triphosphate selectively activates calcium signaling in mast cells. Proc Natl Acad Sci USA 88: 926–930CrossRefGoogle Scholar
  41. 41.
    Wakui M, Kase H, Petersen OH (1991) Cytoplasmic Ca2+ signals evoked by activation of cholecystokinin receptors. Ca2+-dependent current recording in internally perfused pancreatic acinar cells. J Membr Biol 124: 179–187PubMedCrossRefGoogle Scholar
  42. 42.
    Wank SA, Harkins R, Jensen RT, Shapira H, De Weerth A, Slattery T (1992) Purification, molecular cloning, and functional expression of the cholecystokinin receptor from rat pancreas. Proc Natl Acad Sci USA 89: 3125–3129PubMedCrossRefGoogle Scholar
  43. 43.
    Watson SP, Stanley AF, Sasaguri T (1988) Does the hydrolysis of inositol phospholipids lead to the opening of voltage operated Ca2+ channels in guinea pig ileum? Studies with fluoride ions and caffeine. Biochem Biophys Res Commun 153: 14–20PubMedCrossRefGoogle Scholar
  44. 44.
    Williams JA, Burnham DB, Hootman SR (1989) Cellular regulation of pancreatic secretion. In: Schulz SG, Forte FG, Rauner BB (eds). Handbook of physiology, vol 3: The gastrointestinal system. American Physiological Society. Bethesda, Md, pp 419–441Google Scholar
  45. 45.
    Yatani A, Brown AM (1991) Mechanism of fluoride activation of G protein-gated muscarinic atrial K+ channels. J Biol Chem 266: 22872–22877PubMedGoogle Scholar

Copyright information

© Springer-Verlag 1993

Authors and Affiliations

  • Albrecht Piiper
    • 1
  • Danuta Stryjek-Kaminska
    • 1
  • Jürgen Stein
    • 1
  • Wolfgang F. Caspary
    • 1
  • Stefan Zeuzem
    • 1
  1. 1.Abteilung für Gastroenterologie, Zentrum der Inneren MedizinUniversitätsklinikumFrankfurt/MainGermany

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