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Determination of phosphotyrosine phosphatase (PTPase) activity in normal and transformed cells

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Abstract

Through the development of a new colorimetric and non-radioactive method it is possible to measure PTPase activity in cell extracts. Applying this method we were able to demonstrate that intracellular PTPase activity in chicken embryo fibroblasts increases significantly after RSV induced transformation of these cells. PTPase activity in normal and transformed cells is inhibited by μM concentrations of vanadate, molybdate and zinc ions, but is not affected by mM concentrations of calcium, magnesium and sodium fluoride. The transformation specific activation of the PTPase seems to represent an early parameter during the events of cellular transformation.

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References

  1. Krebs EG, Beavo JA: Phosphorylation and dephosphorylation of enzymes. Annu Rev Biochem 48: 923–959, 1979

    Google Scholar 

  2. Reynolds FH Jr, Todaro GJ, Fryling C, Stephenson JR: Human transforming growth factors induce tyrosine phosphorylation of EGF receptors. Nature 292: 259–262, 1981

    Google Scholar 

  3. Hunter T, Sefton BM: The transforming gene product of Rous sarcoma virus phosphorylates tyrosine. Proc Natl Acad Sci USA 77: 1311–1315, 1980

    Google Scholar 

  4. Sefton BM, Hunter T, Beemon K, Eckhart N: Evidence that the phosphorylation of tyrosine is essential for cellular transformation by Rous sarcoma virus. Cell 20: 207, 1980

    Google Scholar 

  5. Nelson RL, Branton PE: Identification, purification and characterization of phosphotyrosine-specific protein phosphatases from cultured chicken embryo fibroblasts. Mol Cell Biol 4: 1003–1012, 1984

    Google Scholar 

  6. Woodford-Thomas TA, Rhodes JD, Dixon JE: Expression of a protein tyrosine phosphatase in normal and v-src transformed mouse 3T3 fibroblasts. J Cell Biol 117: 401–414, 1992

    Google Scholar 

  7. Foulkes JG, Howard RF, Ziemiecki A: Detection of a novel mammalian protein phosphatase with activity for phosphotyrosine. FEBS Letters 130: 197–204, 1981

    Google Scholar 

  8. Tonks NK, Diltz CD, Fischer EH: Purification of the major protein-tyrosine-phosphatase of human placenta. J Biol Chem 263: 6722–6730, 1988

    Google Scholar 

  9. Swarup C, Cohen S, Garbers DL: Inhibition of membrane phosphotyrosyl-protein phosphatase activity by vanadate. Biochem Biophys Res Commun 107: 1104–1109, 1982

    Google Scholar 

  10. Klarlund JK: Transformation of cells by an inhibitor of phosphatases acting on phosphotyrosine in proteins. Cell 41: 707–717, 1985

    Google Scholar 

  11. Branton PE, Landry-Maglan J: Plasma membrane phosphoproteins in normal and Rous sarcoma virus transformed chick embryo fibroblasts: characterization byin vitro phosphorylation. J Cell Physiol 100: 159–168, 1979

    Google Scholar 

  12. Weber MJ, Friis RR: Dissociation of transformation parameters using temperature-conditional mutants of Rous sarcoma virus. Cell 16: 25–32, 1979

    Google Scholar 

  13. Ziemiecki A, Friis RR: Simultaneous injection of newborn rabbits with the Schmidt-Ruppin and Prague strains of Rous sarcoma virus induces antibodies which recognize the pp60src of both strains. J Gen Virol 50: 211–216, 1980

    Google Scholar 

  14. Brugge JS, Erikson RL: Identification of a transformation specific antigen induced by avian sarcoma virus. Nature 269: 346–348, 1977

    Google Scholar 

  15. Barnekow A, Bauer H: The differential expression of the cellularsrc-gene product pp60src and its phosphokinase activity in normal chicken cells and tissues. Biochim Biophys Acta 782: 94–102, 1984

    Google Scholar 

  16. Lowry OH, Rosebrough NJ, Farr AL, Randall R: Protein measurement with the folin phenol reagent. J Biol Chem 193: 265–275, 1951

    Google Scholar 

  17. Kessler WS: Rapid isolation of antigens from cells with a Staphyllococcal protein A-antibody adsorbent: parameters of the interaction of the antibody antigen complexes with protein A. J Immunol 115: 1617, 1975

    Google Scholar 

  18. Baykov AA, Evtushenko OA, Avaeva SM: A malachite green procedure for orthophosphate determination and its use in alkaline phosphatase-based enzyme immunoassay. Anal Biochem 171: 266–270, 1988

    Google Scholar 

  19. Boschek BC, Jockusch BM, Friis RR, Back R, Grundmann E, Bauer H: Early changes in the distribution and organization of microfilament proteins during cell transformation. Cell 24: 175–184, 1981

    Google Scholar 

  20. Tonks NK, Yang Q, Guida P Jr: Structure, regulation and function of protein tyrosine phosphatases. Cold Spring Harbor Symposia on Quantitative Biology, Vol. LVI, Cold Spring Harbor Laboratory Press, pp 265–273, 1991

    Google Scholar 

  21. Imes SS, Kaplan NO, Knowles AF: Plasma membrane-associated phosphatase activities hydrolyzing [32P]phosphotyrosyl histones and [32P]phosphatidyl-inositolphosphate. Anal Biochem 161:316–322, 1987

    Google Scholar 

  22. Lau K-HW, Farley JR, Baylink DJ: Phosphotyrosyl protein phosphatase activity of a bovine skeletal acid phosphatase isoenzyme. J Biol Chem 260: 4653–4660, 1985

    Google Scholar 

  23. Chen J, Martin BL, Brautigan DL: Regulation of protein serinethreonine phosphatase Type-2A by tyrosine phosphorylation. Science 257:1261–1264, 1992

    Google Scholar 

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Authors and Affiliations

  1. Dept. Exp. Tumorbiology, University Muenster, Badestr. 9, W-48149, Muenster, Germany

    Joachim Kremerskothen & Angelika Barnekow

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  1. Joachim Kremerskothen
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  2. Angelika Barnekow
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Kremerskothen, J., Barnekow, A. Determination of phosphotyrosine phosphatase (PTPase) activity in normal and transformed cells. Mol Cell Biochem 125, 1–9 (1993). https://doi.org/10.1007/BF00926828

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  • DOI: https://doi.org/10.1007/BF00926828

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