Skip to main content
PMC home page
The Journal of Cell Biology logoLink to The Journal of Cell Biology
. 1985 Dec 1;101(6):2345–2354. doi: 10.1083/jcb.101.6.2345

Cell surface tubulin in leukemic cells: molecular structure, surface binding, turnover, cell cycle expression, and origin

PMCID: PMC2114015  PMID: 4066762

Abstract

We report here new characteristics of cell surface tubulin from a human leukemia cell line. These cells (CEM cells) possess tubulin that is readily iodinated on the surface of living cells, turns over at a rate identical to that of other surface proteins, and is present throughout the cell cycle. When removed with trypsin, it rapidly returns to the surface. Peptide mapping of iodinated surface tubulin indicates that it possesses a similar, but not identical, primary structure to total CEM and rat brain tubulin. Living CEM cells are able to bind specifically a subfraction of CEM tubulin from metabolically labeled high speed supernatants of lysed CEM cells. Surface tubulin is more basic than the total tubulin pool. The binding, which is saturable, is inhibited by unlabeled CEM high speed supernatants but not by excess thrice-cycled rat or bovine brain tubulin. Surface tubulin is also shown to bind to living nontransformed normal rat kidney cells but not to normal, circulating, mononuclear white cells. Activated lymphocytes produce a tubulin that binds to CEM cells. Since CEM tubulin was detected in the media of 6-h cultures of CEM cells, we must conclude that at least some of the surface tubulin comes from the media. We further conclude that these leukemic cells produce an unusual tubulin that may bind specifically to any membrane. The presence of iodinatable surface tubulin, however, appears to require both the production of a unique tubulin and the presence of a "receptor-like" surface binding component.

Full Text

The Full Text of this article is available as a PDF (1.3 MB).

Selected References

These references are in PubMed. This may not be the complete list of references from this article.

  1. Bachvaroff R. J., Miller F., Rapaport F. T. Appearance of cytoskeletal components on the surface of leukemia cells and of lymphocytes transformed by mitogens and Epstein-Barr virus. Proc Natl Acad Sci U S A. 1980 Aug;77(8):4979–4983. doi: 10.1073/pnas.77.8.4979. [DOI] [PMC free article] [PubMed] [Google Scholar]
  2. Bernier-Valentin F., Aunis D., Rousset B. Evidence for tubulin-binding sites on cellular membranes: plasma membranes, mitochondrial membranes, and secretory granule membranes. J Cell Biol. 1983 Jul;97(1):209–216. doi: 10.1083/jcb.97.1.209. [DOI] [PMC free article] [PubMed] [Google Scholar]
  3. Bhattacharyya B., Volff J. Membrane-bound tubulin in brain and thyroid tissue. J Biol Chem. 1975 Oct 10;250(19):7639–7646. [PubMed] [Google Scholar]
  4. Blitz A. L., Fine R. E. Muscle-like contractile proteins and tubulin in synaptosomes. Proc Natl Acad Sci U S A. 1974 Nov;71(11):4472–4476. doi: 10.1073/pnas.71.11.4472. [DOI] [PMC free article] [PubMed] [Google Scholar]
  5. Bonner W. M., Laskey R. A. A film detection method for tritium-labelled proteins and nucleic acids in polyacrylamide gels. Eur J Biochem. 1974 Jul 1;46(1):83–88. doi: 10.1111/j.1432-1033.1974.tb03599.x. [DOI] [PubMed] [Google Scholar]
  6. Carraway C. A., Jung G., Craik J. R., Rubin R. W., Carraway K. L. Identification of a cytoskeleton-associated glycoprotein from isolated microvilli of a mammary ascites tumor. Exp Cell Res. 1983 Feb;143(2):303–308. doi: 10.1016/0014-4827(83)90055-1. [DOI] [PubMed] [Google Scholar]
  7. Cleveland D. W., Fischer S. G., Kirschner M. W., Laemmli U. K. Peptide mapping by limited proteolysis in sodium dodecyl sulfate and analysis by gel electrophoresis. J Biol Chem. 1977 Feb 10;252(3):1102–1106. [PubMed] [Google Scholar]
  8. Corcoran J. J., Lattanzio F. A., Jr, Rubin R. W., Pressman B. C. Subcellular fractions of the adrenal medulla. Comparison by two-dimensional polyacrylamide gel electrophoresis. Biochim Biophys Acta. 1982 Oct 5;707(2):226–235. doi: 10.1016/0167-4838(82)90355-7. [DOI] [PubMed] [Google Scholar]
  9. FOLEY G. E., LAZARUS H., FARBER S., UZMAN B. G., BOONE B. A., MCCARTHY R. E. CONTINUOUS CULTURE OF HUMAN LYMPHOBLASTS FROM PERIPHERAL BLOOD OF A CHILD WITH ACUTE LEUKEMIA. Cancer. 1965 Apr;18:522–529. doi: 10.1002/1097-0142(196504)18:4<522::aid-cncr2820180418>3.0.co;2-j. [DOI] [PubMed] [Google Scholar]
  10. Foley G. E., Lazarus H. The response in vitro, of continuous cultures of human lymphoblasts (CCRF-CEM cells) to chemotherapeutic agents. Biochem Pharmacol. 1967 Apr;16(4):659–674. doi: 10.1016/0006-2952(67)90078-0. [DOI] [PubMed] [Google Scholar]
  11. Gozes I., Littauer U. Z. The alpha-subunit of tubulin is preferentially associated with brain presynaptic membrnae. FEBS Lett. 1979 Mar 1;99(1):86–90. doi: 10.1016/0014-5793(79)80255-0. [DOI] [PubMed] [Google Scholar]
  12. Grefrath S. P., Reynolds J. A. Polypeptide components of an excitable plasma membrane. J Biol Chem. 1973 Sep 10;248(17):6091–6094. [PubMed] [Google Scholar]
  13. Gurd J. W., Jones L. R., Mahler H. R., Moore W. J. Isolation and partial characterization of rat brain synaptic plasma membranes. J Neurochem. 1974 Feb;22(2):281–290. doi: 10.1111/j.1471-4159.1974.tb11591.x. [DOI] [PubMed] [Google Scholar]
  14. Hubbard A. L., Cohn Z. A. The enzymatic iodination of the red cell membrane. J Cell Biol. 1972 Nov;55(2):390–405. doi: 10.1083/jcb.55.2.390. [DOI] [PMC free article] [PubMed] [Google Scholar]
  15. Kelly W. G., Passaniti A., Woods J. W., Daiss J. L., Roth T. F. Tubulin as a molecular component of coated vesicles. J Cell Biol. 1983 Oct;97(4):1191–1199. doi: 10.1083/jcb.97.4.1191. [DOI] [PMC free article] [PubMed] [Google Scholar]
  16. Krishan A., Dutt K., Israel M., Ganapathi R. Comparative effects of adriamycin and N-trifluoroacetyladriamycin-14-valerate on cell kinetics, chromosomal damage, and macromolecular synthesis in vitro. Cancer Res. 1981 Jul;41(7):2745–2750. [PubMed] [Google Scholar]
  17. Krishan A., Frei E., 3rd Morphological basis for the cytolytic effect of vinblastine and vincristine on cultured human leukemic lymphoblasts. Cancer Res. 1975 Mar;35(3):497–501. [PubMed] [Google Scholar]
  18. Krishan A. Rapid flow cytofluorometric analysis of mammalian cell cycle by propidium iodide staining. J Cell Biol. 1975 Jul;66(1):188–193. doi: 10.1083/jcb.66.1.188. [DOI] [PMC free article] [PubMed] [Google Scholar]
  19. Laemmli U. K. Cleavage of structural proteins during the assembly of the head of bacteriophage T4. Nature. 1970 Aug 15;227(5259):680–685. doi: 10.1038/227680a0. [DOI] [PubMed] [Google Scholar]
  20. Mariash C. N., Seelig S., Oppenheimer J. H. A rapid, inexpensive, quantitative technique for the analysis of two-dimensional electrophoretograms. Anal Biochem. 1982 Apr;121(2):388–394. doi: 10.1016/0003-2697(82)90498-5. [DOI] [PubMed] [Google Scholar]
  21. Nath J., Flavin M. A structural difference between cytoplasmic and membrane-bound tubulin of brain. FEBS Lett. 1978 Nov 15;95(2):335–338. doi: 10.1016/0014-5793(78)81024-2. [DOI] [PubMed] [Google Scholar]
  22. Rousset B., Wolff J. Lactoperoxidase-tubulin interactions. J Biol Chem. 1980 Mar 25;255(6):2514–2523. [PubMed] [Google Scholar]
  23. Rousset B., Wolff J. Purification of brain tubulin by affinity chromatography on immobilized lactoperoxidase. J Biol Chem. 1980 Dec 25;255(24):11677–11681. [PubMed] [Google Scholar]
  24. Rubin R. W., Leonardi C. L. Two-dimensional polyacrylamide gel electrophoresis of membrane proteins. Methods Enzymol. 1983;96:184–192. doi: 10.1016/s0076-6879(83)96016-0. [DOI] [PubMed] [Google Scholar]
  25. Rubin R. W., Quillen M., Corcoran J. J., Ganapathi R., Krishan A. Tubulin as a major cell surface protein in human lymphoid cells of leukemic origin. Cancer Res. 1982 Apr;42(4):1384–1389. [PubMed] [Google Scholar]
  26. Shelanski M. L., Gaskin F., Cantor C. R. Microtubule assembly in the absence of added nucleotides. Proc Natl Acad Sci U S A. 1973 Mar;70(3):765–768. doi: 10.1073/pnas.70.3.765. [DOI] [PMC free article] [PubMed] [Google Scholar]
  27. Soifer D., Czosnek H. Association of newly synthesized tubulin with brain microsomal membranes. J Neurochem. 1980 Nov;35(5):1128–1136. doi: 10.1111/j.1471-4159.1980.tb07868.x. [DOI] [PubMed] [Google Scholar]
  28. Steiner M. Membrane-bound tubulin in human platelets. Biochim Biophys Acta. 1983 Mar 23;729(1):17–22. doi: 10.1016/0005-2736(83)90450-9. [DOI] [PubMed] [Google Scholar]
  29. Stephens R. E. Chemical differences distinguish ciliary membrane and axonemal tubulins. Biochemistry. 1981 Aug 4;20(16):4716–4723. doi: 10.1021/bi00519a030. [DOI] [PubMed] [Google Scholar]
  30. Stephens R. E. Reconstitution of ciliary membranes containing tubulin. J Cell Biol. 1983 Jan;96(1):68–75. doi: 10.1083/jcb.96.1.68. [DOI] [PMC free article] [PubMed] [Google Scholar]
  31. Strocchi P., Brown B. A., Young J. D., Bonventre J. A., Gilbert J. M. The characterization of tubulin in CNS membrane fractions. J Neurochem. 1981 Nov;37(5):1295–1307. doi: 10.1111/j.1471-4159.1981.tb04681.x. [DOI] [PubMed] [Google Scholar]
  32. Swanstrom R., Shank P. R. X-Ray Intensifying Screens Greatly Enhance the Detection by Autoradiography of the Radioactive Isotopes 32P and 125I. Anal Biochem. 1978 May;86(1):184–192. doi: 10.1016/0003-2697(78)90333-0. [DOI] [PubMed] [Google Scholar]
  33. Valeriote F., van Putten L. Proliferation-dependent cytotoxicity of anticancer agents: a review. Cancer Res. 1975 Oct;35(10):2619–2630. [PubMed] [Google Scholar]
  34. Zenner H. P., Pfeuffer T. Microtubular proteins in pigeon erythrocyte membranes. Eur J Biochem. 1976 Dec;71(1):177–184. doi: 10.1111/j.1432-1033.1976.tb11104.x. [DOI] [PubMed] [Google Scholar]
  35. Zisapel N., Levi M., Gozes I. Tubulin: an integral protein of mammalian synaptic vesicle membranes. J Neurochem. 1980 Jan;34(1):26–32. doi: 10.1111/j.1471-4159.1980.tb04617.x. [DOI] [PubMed] [Google Scholar]

Articles from The Journal of Cell Biology are provided here courtesy of The Rockefeller University Press

RESOURCES