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Protein phosphorylation patterns during aestivation in the land snailOtala lactea

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Abstract

Protein phosphorylation patterns were investigated in whole tissues and subcellular fractions of active and aestivatingOtala lactea (Müller) (Pulmonata, Helicidae). Measurement of overall protein phosphorylation showed that incorporation of32P increased until the second day after injection and remained constant for the remaining 4 days of the time course. Comparison of tissues from aestivating and active snails on day 3 showed a decreased protein phosphorylation in aestivating snails (44% of active). No differences in total and protein-associated radioactivity for foot, mantle or haemolymph were observed. Subcellular fractionation of the hepatopancreas localized the changes to plasma membrane, microsomal, and cytosolic fractions: values for aestivating animals were reduced to 71, 37 and 58% of the corresponding active values. Separation of the individual subcellular fractions on isoelectric focusing columns revealed differences in the phosphate incorporation patterns. Plasma membrane from aestivating animal hepatopancreas had a lower overall level of incorporation and fewer radioactive peaks in the pH 7–10 region than did the plasma membrane fraction from active animals. SDS-PAGE analysis of plasma membrane fractions from active and aestivating snails showed a relative decrease in phosphorylation between 60–80 kDa and 30–40 kDa. IEF analysis of cytosolic proteins from aestivating snail hepatopancreas also showed peaks of radioactivity that were apparently shifted by 0.3 pH units toward higher pI values. Increased phosphate incorporation was observed at a peak that corresponded to the pI value for pyruvate kinase in aestivating snails but definite assignment of peaks was not possible. SDS-PAGE analysis of cytosolic proteins showed an aestivation-related decrease in relative protein phosphorylation between 30–35 kDa and 40–45 kDa. A relative increase in phosphorylation during aestivation was observed for proteins between 16–22 kDa. Overall, the data indicate that snails dramatically alter their protein phosphorylation pattern in hepatopancreas during aestivation. (Mol Cell Biochem143: 7–13, 1995)

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Abbreviations

CY:

cytosol

dpm:

radioactive disintegrations per minute

IEF:

isoelectrofocusing

GP:

glycogen phosphorylase

MC:

microsomes

MT:

mitochondria

PAGE:

polyacrylamide gel electrophoresis

PKF:

phosphofructokinase

PK:

pyruvate kinase

PM:

plasma membrane

SDS:

sodium dodecyl sulphate

References

  1. Machin J: Water relationships. In: V Fretter, J Peake (eds) The Pulmonates. Academic Press, New York, pp 105–163, 1975

    Google Scholar 

  2. Livingstone DR, de Zwaan A: Carbohydrate metabolism in gastropods. In: KM Wilbur (ed.) The Mollusca. Academic Press, New York, vol 1, pp 177–242, 1983

    Google Scholar 

  3. Herreid CF: Metabolism of land snails (Otala lactea) during dormancy, arousal and activity. Comp Biochem Physiol A 56: 211–215, 1977

    Google Scholar 

  4. Barnhart MC, McMahon BR: Discontinuous CO2 release and metabolic depression in dormant land snails. J Exp Biol 128: 123–138, 1987

    Google Scholar 

  5. Rees BB, Hand SC: Heat dissipation, gas exchange and acid-base status in the land snailOreohelix during short-term estivation. J Exp Biol 152: 77–92, 1990

    Google Scholar 

  6. Brooks SPJ, Storey KB: Glycolytic enzyme binding and metabolic control in estivation and anoxia in the land snailOtala lactea. J Exp Biol 151: 193–204, 1990

    Google Scholar 

  7. Whitwam RE, Storey KB: Pyruvate kinase from the land snailOtala lactea: regulation by reversible phosphorylation during estivation and anoxia. J Exp Biol 154: 321–337, 1990

    Google Scholar 

  8. Whitwam RE, Storey KB: Regulation of phosphofructokinase during estivation and anoxia in the land snailOtala lactea. Physiol Zool 64: 595–610, 1991

    Google Scholar 

  9. Storey KB: Suspended animation: the molecular basis of metabolic depression. Can J Zool 66: 124–132, 1988

    Google Scholar 

  10. Storey KB, Storey JM: Metabolic rate depression and biochemical adaptation in anaerobiosis, hibernation and estivation. Quart Rev Biol 65: 145–174, 1990

    Google Scholar 

  11. Edelman AM, Blumenthal DK, Krebs EG: Protein serine/threonine kinases. Ann Rev Biochem 56: 567–613, 1987

    Google Scholar 

  12. Blackshear PJ, Nairn AC, Kuo JF: Protein kinases, 1988: a current perspective. FASEB J 2: 2957–2969, 1988

    Google Scholar 

  13. Shenolikar S: Protein phosphorylation: hormones, drugs and bioregulation. FASEB J 2: 2753–2764, 1988

    Google Scholar 

  14. Brooks SPJ, Storey KB: Properties of pyruvate dehydrogenase from the land snail,Otala lactea: control of enzyme activity during estivation. Physiol Zool 65: 620–633, 1992

    Google Scholar 

  15. Catterall WA: The molecular basis of neuronal excitability. Science 223: 653–661, 1984

    Google Scholar 

  16. Reuter H: Regulation of ion channels by phosphorylation and second messengers. News in Physiol Sci 2: 168–171, 1987

    Google Scholar 

  17. Catterall WA, Seagar MJ, Takahashi M: Molecular properties of dihydropyridine-sensitive calcium channels in skeletal muscle. J Biol Chem 263: 3535–3538, 1988

    Google Scholar 

  18. Brautigan DL: Molecular defects in ion channel regulation in cystic fibrosis predicted from analysis of protein phosphorylation/dephosphorylation. Int J Biochem 8: 745–752, 1988

    Google Scholar 

  19. Birnie GD: Subcellular Components: Preparation and Fractionation. London: Butterworth, 1972

    Google Scholar 

  20. Morre DJ: Isolation of Golgi Apparatus. In: WB Jacoby (ed.) Methods in enzymology. Academic Press, New York, vol XXII, pp 131–144, 1971

    Google Scholar 

  21. Sottocasa GL, Kuylenstierna B, Ernster L, Bergstrand A: An electron transport system associated with the outer membrane of liver mitochondria. A biochemical and morphological study. J Cell Biol 32: 415–438, 1967

    Google Scholar 

  22. Strobel HW, Dignam JD: Purification and properties of NADPH-cytochrome c P-450 reductase. In: S. Fleischer, L Packer (eds) Methods in Enzymology. Academic Press, New York, vol LII, pp 89–96, 1978

    Google Scholar 

  23. Laemmli UK: Cleavage of structural proteins during the assembly of the head of bacteriophage T4. Nature 227: 680–685, 1970

    Google Scholar 

  24. Vesterberg O: Isoelectrofocusing of protein. In: SP Colowick, NO Kaplan (eds) Methods in Enzymology. Academic Press, New York, vol XXII, pp 389–412, 1971

    Google Scholar 

  25. Helmerhorst E, Stokes GB: Microcentrifuge desalting: A rapid, quantitative method for desalting small amounts of protein. Anal Biochem 104: 130–135, 1980

    Google Scholar 

  26. Barnhart MC, McMahon BR: Depression of aerobic metabolism and intracellular pH by hypercapnia in land snails,Otala lactea. J Exp Biol 138: 289–299, 1988

    Google Scholar 

  27. Hemmings HC Jr, Nairn AC, McGuiness TL, Huganir RL, Greengard P: Role of protein phosphorylation in neuronal signal transduction. FASEB J 3: 1583–1592, 1989

    Google Scholar 

  28. De Peyer JE, Cachelin AB, Levitan IB, Reuter H: Ca2+-activated K+ conductance in internally perfused snail neurons is enhanced by protein phosphorylation. Proc Ntl Acad Sci USA 79: 4207–4211, 1982

    Google Scholar 

  29. Levitan IB: Phosphorylation of ion channels. J Membr Biol 87: 177–190, 1985

    Google Scholar 

  30. Mooibroek MJ, Wang JH: Integration of signal-transduction processes. Can J Biochem Cell Biol 66: 557–566, 1988

    Google Scholar 

  31. Hochachka PW: Metabolic, channel-, and pump-coupled functions: constraints and compromises of coadaptation. Can J Zool 66: 1015–1027, 1988

    Google Scholar 

  32. Lutz PL: Interaction between hypometabolism and acid-base balance. Can J Zool 67: 3018–3023, 1988

    Google Scholar 

  33. Raju PH, Reddy GR, Babu GRV, Reddanna P, Chetty CS: Kinetic study of Mg2+ and Ca2+-ATPase in the hepatopancreas of estivating freshwater snail,Pila globosa (Swainson). Biochem Int 18: 1069–1075, 1989

    Google Scholar 

  34. Brooks SPJ, Storey KB: Patterns of protein synthesis and protein phosphorylation during anoxia in the land snailOtala lactea. Can J Zool, in press, 1994

  35. Brooks SPJ, Storey KB: Evidence for estivation specific proteins inOtala lactea. Mol Cell Biochem 143: 15–20, 1995

    Google Scholar 

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Author notes

  1. Stephen P. J. Brooks

    Present address: Nutrition Research Division, Food Directorate, Health Protection Branch, Health Canada, Banting Research Centre, Tunney's Pasture, K1A 0L2, Ottawa, Ontario, Canada

Authors and Affiliations

  1. Department of Biology and Institute of Biochemistry, Carleton University, K1S 5B6, Ottawa, Ontario, Canada

    Stephen P. J. Brooks & Kenneth B. Storey

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  1. Stephen P. J. Brooks
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  2. Kenneth B. Storey
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Brooks, S.P.J., Storey, K.B. Protein phosphorylation patterns during aestivation in the land snailOtala lactea . Mol Cell Biochem 143, 7–13 (1995). https://doi.org/10.1007/BF00925921

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