Summary
The effect of energy deprivation on the intracellular transport and secretion of thyroglobulin was studied in open follicles isolated from porcine thyroids. Follicles were pulse-labeled with 3H-leucine or 3H-galactose. Labeled thyroglobulin was secreted into the incubation medium where it was isolated by means of immunoprecipitation. Secretion was followed in chase incubations under various experimental conditions using CCCP (carbonyl-cyanide-mchlorophenylhydrazone) or DNP (dinitrophenol), both uncouplers of oxidative phosphorylation, or CN−, which inhibits respiration. CCCP (1 μM) was shown to inhibit exocytosis by about 80%, DNP (0.1–5 mM) by 45–85%, and CN− (0.5–1.1 mM) by 5–55%. By combining CN− with the ionophore monensin, which blocks transport through the Golgi complex but does not essentially interfere with exocytosis, evidence was obtained that CN− also inhibits transport of thyroglobulin from the Golgi cisternae to the exocytic vesicles by 40%. Electron-miroscopic autoradiography of isolated thyroid lobes from the rat also indicated that transport of 3H-leucine label into the follicle lumen is inhibited in the presence of CCCP or CN−. Intracellular ATP content was found to be about 40% of the control level in follicles incubated with CCCP (1 uM) or CN− (0.9 mM). The results show that the transport of thyroglobulin from the Golgi complex to the exocytic vesicles as well as from the exocytic vesicles into the follicle lumen is dependent upon metabolic energy. The transport blocks are probably associated with inhibited membrane fusions and fissions.
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Abbreviations
- CCCP:
-
carbonylcyanide-m-chlorophenylhydrazone
- FCCP:
-
carbonylcyanide-p-trifluoromethoxyphenylhydrazone
- DNP:
-
dinitrophenol
References
Antoine J-C, Gould B, Jouanne C, Maurice M, Feldmann G (1985) Ammonium chloride, methylamine and chloroquine reversibly inhibit antibody secretion by plasma cells. Biol Cell 55:41–54
Billig H, Rosberg S (1986) Gonadotropin depression of adenosine triphosphate levels and interaction with adenosine in rat granulosa cells. Endocrinology 118:645–652
Björkman U, Ekholm R (1982) Effect of tunicamycin on thyroglobulin secretion. Eur J Biochem 125:585–591
Björkman U, Ekholm R (1984) Generation of H2O2 in isolated porcine thyroid follicles. Endocrinology 115:392–398
Datema R, Schwarz RT (1981) Effect of energy depletion on the glycosylation of a viral glycoprotein. J Biol Chem 256:11191–11198
Denef J-F, Björkman U, Ekholm R (1980) Structural and functional characteristics of isolated thyroid follicles. J Ultrastruct Res 71:185–202
Ekholm R (1981) lodination of thyroglobulin — an intracellular or extracellular process? Mol Cell Endocrinol 24:141–163
Ekholm R, Engström G, Ericson LE, Melander A (1975) Exocytosis of protein into the thyroid follicle lumen: An early effect of TSH. Endocrinology 97:337–346
Fries E, Rothman JE (1980) Transport of vesicular stomatitis virus glycoprotein in a cell-free extract. Proc Natl Acad Sci USA 77:3870–3874
Howell SL (1972) Role of ATP in the intracellular translocation of proinsulin and insulin in the rat pancreatic B-cell. Nature 235:85–86
Jamieson JD, Palade GE (1968) Intracellular transport of secretory proteins in the pancreatic exocrine cell. IV. Metabolic requirements. J Cell Biol 39:589–603
Jamieson JD, Palade GE (1971) Condensing vacuole conversion and zymogen granule discharge in pancreatic exocrine cells: Metabolic studies. J Cell Biol 48:503–522
Kruse NJ, Bornstein P (1975) The metabolic requirements for transcellular movement and secretion of collagen. J Biol Chem 250:4841–4847
Kääriäinen L, Hashimoto K, Saraste J, Virtanen I, Penttinen K (1980) Monensin and FCCP inhibit the intracellular transport of alphavirus membrane glycoproteins. J Cell Biol 87:783–791
Lowry OH, Rosebrough NJ, Farr AL, Randall RJ (1951) Protein measurement with Folin phenol reagent. J Biol Chem 193:265–275
Lyman GE, DeVincenzo JP (1967) Determination of picogram amounts or ATP using the luciferin-luciferase enzyme system. Anal Biochem 21:435–443
McIntire FC, Sproull MF (1957) A simple method for determination of desoxypentosenucleic acid in tissue cultures. Proc Soc Exp Biol Med 95:458–461
Miller DS, Horowitz SB (1986) Intracellular compartmentalization of adenosine triphosphate. J Biol Chem 261:13911–13915
Muir EM, Bowyer DE (1983) Dependence of fluid-phase pinocytosis in arterial smooth-muscle cells on temperature, cellular ATP concentration and the cytoskeletal system. Biochem J 216:467–473
Novick P, Ferro S, Scheleman R (1981) Order of events in the yeast secretory pathway. Cell 25:461–469
Öfverholm T, Ericson LE (1984) Rate of protein transport in thyroid follicle cells of normal, thyroxine-treated and TSH-injected rats. Eur J Cell Biol 35:171–179
Orci L, Ravazzola M, Amherdt M, Madsen O, Vassalli J-D, Perrelet A (1985) Direct identification of prohormone conversion site in insulin-secreting cells. Cell 42:671–681
Peters BP, Krzesicki RF, Perini F, Ruddon RW (1986) The effect of carbonyl cyanide trifluoromethoxyphenylhydrazone and methylamine on the processing and secretion of the glycoprotein hormone chorionic gonadotropin by human carcinoma cells. Endocrinology 119:416–428
Pollard HB, Pazoles CJ, Creutz CE, Scott JH, Zinder O, Hotchkiss A (1984) An osmotic mechanism for exocytosis from dissociated chromaffin cells. J Biol Chem 259:1114–1121
Reyes J, Benos DJ (1984) Changes in interfacial potentials induced by carbonylcyanide phenylhydrazone uncouplers: Possible role of mitochondrial oxygen consumption and other transport processes. Membr Biochem 5:243–268
Ring P, Björkman U, Ekholm R (1987a) Effect of cooling on intracellular transport and secretion. Cell Tissue Res 247:505–513
Ring P, Björkman U, Johanson V, Ekholm R (1987b) The effect of monensin on thyroglobulin secretion. Cell Tissue Res 248:153–160
Ring P, Björkman U, Ekholm R (1987c) Localization of the incorporation of 3H-galactose and 3H-sialic acid into thyroglobulin in relation to the intracellular transport block induced by monensin. Cell Tissue Res (in press)
Schramm M (1967) Secretion of enzymes and other macromolecules. Ann Rev Biochem 36:307–320
Spiro RG, Spiro MJ, Bhoyroo VD (1983) Studies on the regulation of the biosynthesis of glucose-containing oligosaccharide-lipids. J Biol Chem 258:9469–9476
Tartakoff AM (1983) Perturbation of the structure and function of the Golgi complex by monovalent carboxylic ionophores. Methods Enzymol 98:47–59
Tartakoff AM (1986) Temperature and energy dependence of secretory protein transportin the exocrine pancreas. EMBO J 5:1477–1482
Tartakoff AM, Vassalli P (1977) Plasma cell immunoglobulin secretion. Arrest is accompanied by alterations of the Golgi complex. J Exp Med 146:1332–1345
Tartakoff AM, Vassalli P (1978) Comparative studies of intracellular transport of secretory proteins. J Cell Biol 79:694–707
Thorens B, Schulz M-F, Vassalli P (1985) Bone marrow pre-B lymphocytes synthesize immunoglobulin μ chains of membrane type with different properties and intracellular pathways. EMBO J 4:361–368
Van Sande J, Dumont JE (1975) Determination of cyclic 3′, 5′-AMP accumulation in resting and stimulated thyroid slices: Heterogeneity of the ATP precursor pool. Mol Cell Endocrinol 2:289–301
Van Sande J, Ketelbant-Balasse P, Grenier G, Dumont JE (1978) Uncoupling by cooling of secretion and adenosine 3′, 5′-monophosphate accumulation in stimulated dog thyroid slices. Endocrinology 102:576–581
Vilmart-Seuwen J, Kersken H, Sterzl R, Plattner H (1986) ATP keeps exocytosis sites in a primed state but is not required for membrane fusion: An analysis with Paramecium cells invivo and in vitro. J Cell Biol 103:1279–1288
Yates ME, Berry RW (1984) Subcellular sites of processing of precursors to neurosecretory peptides in the bag cells of Aplysia: Inferences from the effects of monensin, FCCP, and chloroquine. J Neurobiol 15:141–155
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Ring, P., Björkman, U. & Ekholm, R. Effect of energy deprivation on intracellular transport and secretion of thyroglobulin. Cell Tissue Res. 250, 701–708 (1987). https://doi.org/10.1007/BF00218966
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DOI: https://doi.org/10.1007/BF00218966