Summary
Isolated bovine thyroid plasma membrane preparations were obtained by isopycnic density gradient centrifugution. Cyclic AMP-PDEase (EC 3.1.4.c) activity has been demonstrated by electron microscopic histochemistry on the unit membrane of isolated bovine thyroid cells. 3-isobutyl-1-methyl-xanthine (IBMX) produced partial inhibiton, while omission of the substrate revealed no reaction product deposition. These observations correlated well with biochemical studies that showed 0.4% of the total cAMP-PDEase activity to be present in the plasma membrane preparations. Kinetic analysis of cAMP hydrolysis yielded two apparent Michaelis constants for the homogenate and the plasma membrane-rich fraction. Dose-response curves for IBMX inhibition showed cAMP-PDEase of the homogenate to be more sensitive to inhibition than that of the plasma membrane-bound enzyme. Furthermore, wash experiments indicate that the plasma membrane-associated enzyme is tightly bound. This investigation strengthens our previous study and suggests that bovine thyroid cell plasma membranes contain a cAMP-PDEase that may be involved in interactions between the cell and the external environment in a manner yet to be determined.
This is a preview of subscription content, log in via an institution to check access.
Similar content being viewed by others
References
Adinolfi AM, Schmidt SY (1974) Cytochemical localization of cyclic nucleotide phosphodiesterase activity at developing synapses. Brain Res 76:21–31
Appleman MM, Terasaki WL (1975) Regulation of cyclic nucleotide phosphodiesterase. Adv Cyclic Nucl Res 5:153–162
Beavo JA, Hardman JG, Sutherland EW (1971) Stimulation of adenosine 3′,5′-monophosphate hydrolysis by guanosine 3′,5′-monophosphate. J Biol Chem 246:3841–3846
Brown BL, Albano JDM, Ekins RP, Sgherzi AM, Tampion W (1971) A simple and sensitive saturation assay method for the measurement of adenosine 3′,5′-monophosphate. Biochem J 121:561–562
Butcher RW, Sutherland EW (1962) Adenosine 3′,5′-phosphate in biological materials. I. Purification and properties of cyclic 3′,5′-nucleotide phosphodiesterase and use of this enzyme to characterize adenosine 3′,5′-phosphate in human urine. J Biol Chem 237:1244–1250
Coleman R, Finean JB (1966) Preparation and properties of isolated plasma membranes from guinea-pig tissues. Biochim Biophys Acta 125:197–206
DePierre JW, Karnovsky ML (1974) Ecto-enzymes of the guinea pig polymorphonuclear leukocyte. Science 183:1096–1098
Drummond GI, Perrot-Yee S (1961) Enzymatic hydrolysis of adenosine 3′,5′-phosphoric acid. J Biol Chem 236:1126–1129
Fain JN, Shepherd RE (1975) Free fatty acids as feedback regulators of adenylate cyclase and cyclic 3′:5′-AMP accumulation in rat fat cells. J Biol Chem 250:6586–6592
Farham CJM (1975) Cytochemical localization of adenylate cyclase and 3′,5′-nucleotide phosphodiesterase in Dictyostelium. Exp Cell Res 91:36–46
Filburn CR, Liang CT, Sacktor B (1977) Cyclic nucleotide phosphodiesterases of the renal cortex. J Membrane Biol 37:29–43
Fiske C, Subbarow Y (1925) The colorimetric determination of phosphorus. J Biol Chem 66:375–400
Florendo NT, Barrnett RJ, Greengard P (1971) Cyclic 3′,5′-nucleotide phosphodiesterases: Cytochemical localization in cerebral cortex. Science 173:745–747
Hatefi Y, Rieske JSI (1967) The preparation and properties of DPNH-cytochrome c reductase (complex I–III of the respiratory chain). In: Estabrook RW, Pullman ME (eds) Methods in enzymology, Vol 10. Academic Press, New York and London, pp 225–231
Indiveri F, Barabino A, Fontana R, Polleri A (1974) Thyrotropin effect on cyclic 3′,5′-nucleotide phosphodiesterase intracellular distribution. Hormone Res 5:65–75
Kalderon AE, Greenberg M, Dobbs JW (1978) Demonstration of cyclic AMP phosphodiesterase activity in isolated bovine thyroid cell membranes. In: George WJ, Ignarro LJ (eds) Advances in cyclic nucleotide research, Vol 9. Raven Press, New York, p 756
Kalderon AE, Ravanshenas SF (1974) Localization of 3′,5′-cyclic nucleotide phosphodiesterase activity in isolated thyroid cells and intact thyroid tissue. Histochemistry 39:229–242
King LE, Jr, Florendo NT, Solomon SS, Hashimoto K (1974) Cyclic 3′,5′-nucleotide phosphodiesterase. I. Histochemical localization in rat skin. J Invest Dermatol 62:485–492
Lineweaver H, Burk D (1934) The determination of enzyme dissociation constants. J Amer Chem Soc 56:658–666
Lowry OH, Rosebrough NJ, Farr AL, Randall RJ (1951) Protein measurement with the folin phenol reagents. J Biol Chem 193:265–275
Luft JH (1961) Improvements in epoxy resin embedding methods. J Biophys Biochem Cytol 9:409–414
Malchow D, Nagele B, Schwarz H, Gerisch G (1972) Membrane-bound cyclic AMP phosphodiesterase in chemotactically responding cells of Dictyostelium discoideum. Eur J Biochem 28:136–142
Reynolds ES (1963) The use of lead citrate at high pH as an electron-opaque stain in electron microscopy. J Cell Biol 17:208–212
Robb RM (1974) Histochemical evidence of cyclic nucleotide phosphodiesterase in photoreceptor outer segments. Invest Ophthalmol 13:740–747
Rudel L, Morris M (1973) Determination of cholesterol using o-phthalaldehyde. J Lipid Res 14:364–366
Russell J, Pastan I (1973) Plasma membrane cyclic adenosine 3′:5′-monophosphate phosphodiesterase of cultured cells and its modification after trypsin treatment of intact cells. J Biol Chem 248, 5835–5840
Rutten WJ, Schoot BM, DePont JJHM, Bonting SL (1973) Adenosine 3′,5′-monophosphate phosphodiesterase assay in tissue homogenates. Biochim Biophys Acta 315:384–393
Sakai T, Thompson WJ, Lovis VR, Williams RH (1974) Cyclic nucleotide phosphodiesterase activities from isolated fat cells. Correlation of subcellular distribution with effects of nucleotides and insulin. Arch Biochem Biophys 162:331–339
Stanbury JB, Wicken JV, Lafferty MA (1967) Preparation and Properties of Thyroid Cell Membranes. J Membrane Biol 1:459–467
Thompson WJ, Appleman MM (1971) Multiple cyclic nucleotide phosphodiesterase activities from rat brain. Biochemistry 10:311–316
Thompson WJ, Brooker G, Appleman MM (1974) Assay of cyclic nucleotide phosphodiesterases with radioactive substrates. In: Hardman JG, O'Malley BW (eds) Methods in enzymology. Vol 38. Academic Press, New York and London, pp 205–212
Thompson WS, Ross CP, Pledger RW, Strada SJ, Banner RL, Hersh EM (1976) Cyclic adenosine 3′:5′-monophosphate phosphodiesterase. Distinct forms in human lymphocytes and monocytes. J Biol Chem 251:4922–4929
Van Inwegan RG, Pledger WJ, Strada SJ, Thompson WJ (1976) Characterization of cyclic nucleotide phosphodiesterases with multiple separation techniques. Arch Biochem Biophys 175:700–709
Vorbrodt A, Konwinski M, Solter D (1977) Ultrastructural cytochemistry of membrane-bound phosphatase in preimplantation mouse embryos. Dev Biol 55:117–134
Wharton DC, Tzagoloff A (1967) Cytochrome oxidase from beef heart mitochondria. In: Estabrook RW, Pullman M (eds) Methods in enzymology. Vol 10. Academic Press, New York and London pp 245–250
Wolff J, Jones AB (1971) The purification of bovine thyroid plasma membranes and the properties of membrane-bound adenyl cyclase. J Biol Chem 246:3939–3947
Woo Yin-Tak, Manery JF (1973) Cyclic AMP phosphodiesterase activity at the external surface of intact skeletal muscles and stimulation of the enzyme by insulin. Arch Biochem Biophys 154:510–519
Yamashita K, Field JB (1974) Isolation of the plasma membrane from the thyroid. In: Fleischer S, Packen S (eds) Methods in enzymology, Vol 31. Academic Press, New York and London, pp 144–149
Author information
Authors and Affiliations
Rights and permissions
About this article
Cite this article
Kalderon, A.E., Dobbs, J.W. & Greenberg, M.L. Localization of 3′,5′-cyclic adenosine monophosphate phosphodiesterase (cAMP-PDEase) activity in isolated bovine thyroid plasma membranes. Histochemistry 65, 277–289 (1980). https://doi.org/10.1007/BF00493177
Received:
Issue Date:
DOI: https://doi.org/10.1007/BF00493177