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Properties of cod metallothionein, its presence in different tissues and effects of Cd and Zn treatment

Fish Physiology and Biochemistry volume 13, Article number: 81 (1994) Cite this article

Abstract

One isoform of the low-molecular-weight metal-binding protein metallothionein (MT) has been isolated from the liver of Atlantic cod by size-exclusion and ion-exchange chromatography. Cod MT contained 33% cysteine, no aromatic amino acids or arginine. As is the case for other piscine MTs, the N-terminus of cod MT lacked the asparagine in position 4 which is present in mammalian MTs. In addition, cod MT differed from all other vertebrate MTs described in that the N-terminal methionine was not acetylated. Antibodies were raised in rabbits against hepatic MT from cod by repeated injections of native protein mixed with adjuvant. Anti-cod MT antisera cross reacted with similarly-sized proteins in liver, brain, anterior kidney, posterior kidney, spleen, intestine, gills and ovaries. The putative MT in cod brain migrated differently to that of the other tissues in native gel electrophoresis. Intraperitoneally injected Cd (1 mg/kg) was nearly entirely associated with the MT-peak in hepatic and renal cytosols, whereas a single injection of Zn (10 mg/kg) resulted in increases in all cytosolic Zn pools of the liver and no apparent change in cytosolic Zn, Cu, Ni or Cd in kidney.

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References cited

  • Arizono, K. Ito, T., Ota, S. and Ariyoshi, T. 1985. Purification and characterization of lead-induced zinc thionein in the liver of rats. Bull. environ. Contam. Toxicol. 35: 143–148.

    PubMed  CAS  Article  Google Scholar 

  • Baer, K.N. and Thomas, P. 1991. Isolation of novel metal-binding proteins distinct from metallothionein from spotted seatrout (Cynoscion nebulosus) and Atlantic croaker (Micropogonias undulatus) ovaries. Mar. Biol. 108: 31–37.

    CAS  Article  Google Scholar 

  • Baksi, S.M., Libbus, N. and Frazier, J.M. 1988. Induction of metal binding proteins in striped bass, Morone saxatilis, following cadmium treatment. Comp. Biochem. Physiol. 91C: 355–363.

    CAS  Article  Google Scholar 

  • Bonham, K., Zafarullah, M. and Gedamu, L. 1987. The rainbow trout metallothioneins: molecular cloning and characterization of two distinct cDNA sequences. DNA 6: 519–528.

    PubMed  CAS  Google Scholar 

  • Bradford, M.M. 1976. A rapid and sensitive method for the quantitation of microgram quantities of protein utilizing the principle of protein-dye binding. Anal. Biochem. 72: 248–254.

    PubMed  CAS  Article  Google Scholar 

  • Bremner, I. 1991. Nutritional and physiologic significance of metallothionein. In Metallobiochemistry, Part B: Metallothionein and Related Molecules, pp. 25–35. Edited by J.F. Riordan and B.L. Vallee. Academic Press, San Diego.

    Google Scholar 

  • Bremner, I. and Davies, N.T. 1975. The induction of metallothionein in rat liver by zinc injection and restriction of food intake. Biochem. J. 149: 733–738.

    PubMed  CAS  Google Scholar 

  • Brown, D.A., Bay, S.M. and Hershelman, G.P. 1990. Exposure of scorpionfish (Scorpaena guttata) to cadmium: effects of acute and chronic exposures on the cytosolic distribution of cadmium, copper and zinc. Aquat. Toxicol. 16: 295–310.

    CAS  Article  Google Scholar 

  • Carpenè, E. and Vasak, M. 1989. Hepatic metallothioneins from goldfish (Carassius auratus L.). Comp. Biochem. Physiol. 92B: 463–468.

    Article  Google Scholar 

  • Cartaña, J., Arola, L. and Mas, A. 1991. Effects of acute nickel toxicity upon plasma and liver metal homeostasis as a function of sex. Toxicol. 69: 133–141.

    Google Scholar 

  • Cartaña, J., Romeu, A. and Arola, L. 1992. Effects of copper, cadmium and nickel on liver and kidney glutathione redox cycle of rats (Rattus sp.). Comp. Biochem. Physiol. 101C: 209–213.

    Google Scholar 

  • Chan, K.M. Davidson, W.S., Hew, C.L. and Fletcher, G.L. 1988. Molecular cloning of metallothionein cDNA and analysis of metallothionein gene expression in winter flounder tissues. Can. J. Zool. 67: 2520–2527.

    Article  Google Scholar 

  • Chatterjee, A. and Maiti, I.B. 1987. Purification and immunological characterization of catfish (Heteropneustes fossilis) metallothionein. Mol. cell. Biochem. 78: 55–64.

    PubMed  CAS  Google Scholar 

  • Eaton, D.L., Stacey, N.H., Wong, K. and Klaasen, C.D. 1980. Dose-response effects of various metal ions on rat liver metallothionein, glutathione, heme oxygenase and cytochrome P-450. Toxicol. Appl. Pharmacol. 55: 393–402.

    PubMed  CAS  Article  Google Scholar 

  • Fleet, J.C., Golemboski, K.A., Dietert, R.R., Andrews, G.K. and McCormick, C.C. 1990. Induction of hepatic metallothionein by intraperitoneal metal injection: an associated inflammatory response. Am. J. Physiol. 258: G926–G933.

    Google Scholar 

  • Fowler, B.A., Hilderbrand, C.E., Kojima, Y. and Webb, M. 1987. Nomenclature of metallothioneins. Experientia Suppl. 52: 19–22.

    PubMed  CAS  Google Scholar 

  • George, S.G. 1990. Biochemical and cytological assessments of metal toxicity in marine animals. In Heavy Metals in the Marine Environment. pp. 123–142. Edited by R.W. Furness and P.S. Rainbow. CRC Press, Boca Raton.

    Google Scholar 

  • Hidalgo, J., Campmany, L., Borrás, M., Garvey, J.S. and Armario, A. 1988. Metallothionein response to stress in rats: role in free radical scavenging. Am. J. Physiol. 255: E518–E524.

    Google Scholar 

  • Hidalgo, J. and Flos, R. 1986. Dogfish metallothionein —I. Purification and characterization and comparison with rat metallothionein. Comp. Biochem. Physiol. 83C: 99–103.

    CAS  Google Scholar 

  • Hidalgo, J., Giralt, M., Garvey, J.S. and Armario, A. 1987. Sex and restraint stress differences in rat metallothionein and Zn levels. Rev. Esp. Fisiol. 43: 427–432.

    PubMed  CAS  Google Scholar 

  • Hirs, C.H.W. 1967. Determination of cystine as cysteic acid. Meth. Enzymol. 11: 59–65.

    CAS  Google Scholar 

  • Hunziker, P.E. 1991. Metal removal from mammalian metallothioneins. Meth. Enzymol. 205: 451–452.

    PubMed  CAS  Article  Google Scholar 

  • Hyllner, S.J., Andersson, T., Haux, C. and Olsson, P.-E. 1989. Cortisol induction of metallothionein in primary culture of rainbow trout hepatocytes. J. Cell. Physiol. 139: 24–28.

    PubMed  CAS  Article  Google Scholar 

  • Kikuchi, Y., Irie, M., Ikebuchi, H., Sawada, J.-I., Terao, T., Nakyama, S., Iguchi, S. and Okada, U. 1990. Antigenic determinants on rat metallothionein: fine epitope mapping for a murine monoclonal antibody and rabbit polyclonal antisera. J. Biochem. 107: 650–654.

    PubMed  CAS  Google Scholar 

  • Kille, P., Kay, J., Leaver, M. and George, S. 1992. Induction of piscine metallothionein as a primary response to haevy metal pollutants: applicability of new sensitive molecular probes. Aquat. Toxicol. 22: 2792–286.

    Article  Google Scholar 

  • Kille, P., Stephens, P.E. and Kay, J. 1991. Elucidation of cDNA sequences for metallothioneins from rainbow trout, stone loach and pike using the polymerase chain reaction. Biochem. Biophys. Acta 1089: 407–410.

    PubMed  CAS  Google Scholar 

  • Kito, H., Ose, Y., Mizuhira, V., Sato, T., Ishikawa, T. and Tazawa, T. 1982. Separation and purification of (Cd, Cu, Zn)-metallothionein in carp hepato-pancreas. Comp. Biochem. Physiol. 73C: 121–127.

    CAS  Article  Google Scholar 

  • Krezoski, S., Laib, J., Onana, P., Hartmann, T., Chen, P., Shaw, C.F. and Petering, D.H. 1988. Presence of Zn,Cu-binding protein in liver of freswater fishes in the absence of elevated exogenous metal: relevance to toxic metal exposure. Mar. Environ. Res. 24: 147–150.

    CAS  Article  Google Scholar 

  • Kägi, J.H.R. and Schäffer, A. 1988. Biochemistry of metallothionein. Biochem. 27: 8509–8515.

    Article  Google Scholar 

  • Laemmli, U.K. 1970. Cleavage of structural proteins during the assembly of the head of bacteriophage T4. Nature, Lond. 227: 680–685.

    CAS  Article  Google Scholar 

  • Lauder, G.V. and Liem, K.F. 1983. The evolution and interrelationships of the Actinopterygian fishes. Bull. Mus. Comp. Zool., Harvard 150: 1–197.

    Google Scholar 

  • Lerch, K., Johnson, G.F., Grushoff, P.S. and Sternlieb, I. 1985. Canine hepatic lysosomal copper protein; identification as metallothionein, Arch. Biochem. Biophys. 243: 108–114.

    PubMed  CAS  Article  Google Scholar 

  • Ley, H.L., III and Cherry, D.S. 1983. Isolation and characterization of hepatic metallothionein from rainbow trout (Salmo gairdneri). Comp. Biochem. Physiol. 74B: 507–513.

    Google Scholar 

  • Lin, L.-R. and Huang, P.C. 1990. Pigeon metallothionein consists of two species. Biochim. Biophys. Acta 1037: 248–255.

    PubMed  CAS  Google Scholar 

  • Lohrer, H. and Robson, T. 1989. Overexpression of metallothionein in CHO cells and its effect on cell killing by ionizing radiation and alkylating agents. Carcinogenesis 10: 2279–2284.

    PubMed  CAS  Google Scholar 

  • McCormick, C.C. and Dietert, R.R. 1991. Tissue-specific metallothionein induction by non-binding metals: evidence of an associated inflammatory response. In Metallothionein in Biology and Medicine. pp. 209–220. Edited by C.D. Klaasen and K.T. Suzuki. CRC Press, Boca Rotan.

    Google Scholar 

  • McCormick, C.C., Fullmer, C.S. and Garvey, J.S. 1988. Amino acid sequence and comparative antigenicity of chicken metallothionein. Proc. Natl. Acad. Sci. U.S.A. 85: 309–313.

    PubMed  CAS  Article  Google Scholar 

  • Nielson, K.B., Atkin, C.L. and Winge, D.R. 1985. Distinct metal-binding configurations in metallothionein. J. Biol. Chem. 260: 5342–5350.

    PubMed  CAS  Google Scholar 

  • Noël-Lambot, F., Gerday Ch. and Disteche, A. 1978. Distribution of Cd, Zn an Cu in liver and gills of the eel Anguilla anguilla with special reference to metallothionein. Comp. Biochem. Physiol. 61C: 177–187.

    Google Scholar 

  • Os, S.H., Deagen, J.T., Whanger, P.D. and Weswig, P.H. 1978. Biological function of metallothionein. V. Its induction in rats by various stresses. Am. J. Physiol. 234: E282–E285.

    Google Scholar 

  • Olsson, P.-E. and Hogstrand, C. 1987a. Improved separation of perch liver metallothionein by fast liquid chromatography. J. Chromatography 402: 293–299.

    CAS  Article  Google Scholar 

  • Olsson, P.-E. and Hogstrand, C. 1987b. Subcellular distribution and binding of cadmium to metallothionein in tissues of rainbow trout after exposure to 109Cd in water. Environ. Toxicol. Chem. 6: 867–874.

    CAS  Google Scholar 

  • Olsson, P.-E., Zafarullah, M., Foster, R., Hamor, T. and Gedamu, L. 1990. Developmental regulation of metallothionein mRNA, zinc and copper levels in rainbow trout, Salmo gairdneri. Eur. J. Biochem. 193: 229–235.

    PubMed  CAS  Article  Google Scholar 

  • Ou, C.Z. and Ebadi, M. 1992. Pineal and retinal protein kinase C isoenzymes: cooperative activation by calcium and zinc metallothionein. J. Pineal Res. 12: 17–26.

    PubMed  CAS  Google Scholar 

  • Overnell, J., Berger, C. and Wilson, K.J. 1981. Partial amino acid sequence of metallothionein from the plaice (Pleuronectes platessa). Biochem. Soc. Trans. 9: 217–218.

    CAS  Google Scholar 

  • Overnell, J. and Coombs, T.L. 1979. Purification and properties of plaice metallothionein, a cadmium-binding protein from the liver of the plaice (Pleuronectes platessa). Biochem. J. 183: 277–283.

    PubMed  CAS  Google Scholar 

  • Overnell, J., McIntosh, R. and Fletcher, T.C. 1987. The enhanced induction of metallothionein by zinc, its half-life in the marine fish Pleuronectes platessa, and the influences of stress factors on metallothionein levels. Experientia 43: 178–181.

    PubMed  CAS  Article  Google Scholar 

  • Palmiter, R.D., Findley, S.D., Whitmore, T.E. and Durham, D.M. 1992. MT-III, a brain-specific member of the metallothionein gene family. Proc. Natl. Acad. Sci. U.S.A. 89: 6333–6337.

    PubMed  CAS  Article  Google Scholar 

  • Piñuela, C., Baatrup, E. and Geneser, F.A. 1992. Histochemical distribution of zinc in the brain of the rainbow trout, Oncorhynchus mykiss. I. The telencephalon. Anat. Embryol. 185: 379–388.

    PubMed  Article  Google Scholar 

  • Sato, M. 1991. Metallothionein synthesis in rats exposed to radical generating agents. In Metallothionein in Biology and Medicine. pp. 221–235. Edited by C.D. Klaassen and K.T. Suzuki. CRC Press, Boca Raton.

    Google Scholar 

  • Sato, M. and Sasaki, M. 1991. Enhanced lipid peroxidation is not necessary for induction of metallothionein-I by oxidative stress. Chem. -Biol. Interact. 78: 143–154.

    PubMed  CAS  Article  Google Scholar 

  • Scudiero, R., De Prisco, P.P., Camardella, L., D'Avino, R., Di Prisco, G. and Parisi, E. 1992. Apparent deficiency of metallothionein in the liver of the Antarctic icefish Chionodraco hamatus. Identification and isolation of a zinc-containing protein unlike metallothionein. Comp. Biochem. Physiol. 103B: 201–207.

    CAS  Article  Google Scholar 

  • Shears, M.A. and Fletcher, G.L. 1984. The relationship between metallothionein and intestinal zinc absorption in the winter flounder. Can. J. Zool. 62: 2211–2220.

    CAS  Article  Google Scholar 

  • Takeda, H. and Shimizu, C. 1982a. Existence of the metallothionein-like protein in various fish tissues. Bull. Jap. Soc. Sci. Fish. 48: 711–715.

    CAS  Google Scholar 

  • Takeda, H. and Shimizu, C. 1982b. Purification of metallothionein from the liver of skipjack and its properties. Bull. Jap. Soc. Sci. Fish. 48: 717–723.

    CAS  Google Scholar 

  • Thornally, P.J. and Vasak, M. 1985. Possible role for metallothionein in protection against radiation-induced oxidative stress. Kinetics and mechanism of its reaction with superoxide and hydroxyl radicals. Biochim. Biophys. Acta 827: 36–44.

    Google Scholar 

  • Towbin, H., Staehelin, T. and Gordon, J. 1979. Electrophoretic transfer of proteins from polyacrylamide gels to nitrocellulose sheets: procedure and some applications. Proc. Natl. Acad. Sci. U.S.A. 76: 4350–4354.

    PubMed  CAS  Article  Google Scholar 

  • Weber, D.N., Eisch, S., Spieler, R.E. and Petering, D.H. 1992. Metal redistribution in largemouth bass (Micropterus salmoides) in response to restrainment stress and dietary cadmium: role of metallothionein and other metal-binding proteins. Comp. Biochem. Physiol. 101C: 255–262.

    CAS  Google Scholar 

  • Winge, D.R. and Garvey, J.S. 1983. Antigenicity of metallothionein. Proc. Natl. Acad. Sci. U.S.A. 80: 2472–2476.

    PubMed  CAS  Article  Google Scholar 

  • Winge, D.R. and Rajagopalan, K.V. 1972. Purification and some properties of Cd-binding protein from rat liver. Arch. Biochem. Biophys. 70: 153–175.

    Google Scholar 

  • Yamamura, M. and Suzuki, K.T. 1984. Isolation and characterization of metallothionein from the tortoise Clemmys mutica. Comp. Biochem. Physiol. 79C: 63–69.

    CAS  Article  Google Scholar 

  • Zafarullah, M., Olsson, P.-E. and Gedamu, L. 1989. Endogenous and heavy-metal-ion-induced metallothionein gene expression in salmonid tissues and cell lines. Gene 83: 85–93.

    PubMed  CAS  Article  Google Scholar 

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Author notes

  1. K. Hylland

    Present address: Norwegian Institute for Water Research (NIVA), P.O. Box 173, Kjels»s, N-0411, Oslo, Norway

  2. C. Hogstrand

    Present address: Department of Biology, McMaster University, 1280 Main St. West, Hamilton, Ontario, L8S 4K1, Canada

Affiliations

  1. Section of Marine Zoology and Marine Chemistry, Department of Biology, University of Oslo, P.O. Box 1064, N-0316, Oslo, Norway

    K. Hylland

  2. Department of Zoophysiology, University of Göteborg, Medicinaregatan 18, S-41 390, Göteborg, Sweden

    C. Hogstrand

  3. Department of Biochemistry, University of Oslo, P.O. Box 1041, N-0316, Oslo, Norway

    C. Haux & K. Sletten

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  1. K. Hylland
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  2. C. Haux
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  3. C. Hogstrand
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Hylland, K., Haux, C., Hogstrand, C. et al. Properties of cod metallothionein, its presence in different tissues and effects of Cd and Zn treatment. Fish Physiol Biochem 13, 81 (1994). https://doi.org/10.1007/BF00004122

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Keywords

  • metallothionein
  • Gadus morhua
  • liver
  • anterior kidney
  • brain
  • PAGE
  • immunoblotting
  • N-terminal sequence
  • amino acid composition