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. 1975 Dec;72(12):4981–4985. doi: 10.1073/pnas.72.12.4981

Cytoplasmic microtubules in normal and transformed cells in culture: analysis by tubulin antibody immunofluorescence.

B R Brinkley, E M Fuller, D P Highfield
PMCID: PMC388858  PMID: 174086

Abstract

Monospecific antibody directed against bovine brain tubulin was used as an immunofluorescent probe to evaluate the distribution of microtubules in normal and transformed cells grown in tissue culture. The fluorescent staining pattern of transformed and nontransformed cells is significantly different and may be used in conjunction with other morphological features to identify transformants in mixed cell populations. Normal cells are flattened, elongated, and fibroblastic; they display numerous Colcemid-sensitive fluorescent cytoplasmic filaments, presumably microtubules. Transformed cells, however, are smaller, more polygonal in shape, and contain very few cytoplasmic tubules. During mistosis the cytoplasmic microtubule complex of normal cells completely disappears, but reappears after cell division. Treatment of transformed cells with dibutyry-adenosine 3':5'-cyclic monophosphate plus testosterone or theophylline restores the normal fibroblastic appearance of the cells and stimulates the assembly of numerous cytoplasmic microtubules. This study provides further evidence for two separate microtubule entities in cycling nontransformed cells: a cytoplasmic microtubule complex and the microtubules of the mitotic spindle. Although an interchange of tubulin dimers seems to exist between microtubules in the two systems, control of tubule assembly may be under separate constraints. Stimulation of cytoplasmic microtuble assembly in transformed cells by derivatives of adenosine 3':5'-cycle monophosphate suggests that impairment of the cytoplasmic microtubule complex in these cells may be due to suboptimal levels of adenosine 3':5'-cyclic monophosphate.

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Selected References

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

  1. Borisy G. G., Olmsted J. B. Nucleated assembly of microtubules in porcine brain extracts. Science. 1972 Sep 29;177(4055):1196–1197. doi: 10.1126/science.177.4055.1196. [DOI] [PubMed] [Google Scholar]
  2. Borle A. B. Cyclic AMP stimulation of calcium efflux from kidney, liver and heart mitochondria. J Membr Biol. 1974;16(3):221–236. doi: 10.1007/BF01872416. [DOI] [PubMed] [Google Scholar]
  3. Brinkley B. R., Cartwright J., Jr Cold-labile and cold-stable microtubules in the mitotic spindle of mammalian cells. Ann N Y Acad Sci. 1975 Jun 30;253:428–439. doi: 10.1111/j.1749-6632.1975.tb19218.x. [DOI] [PubMed] [Google Scholar]
  4. Buckley I. K. Three dimensional fine structure of cultured cells: possible implications for subcellular motility. Tissue Cell. 1975;7(1):51–72. doi: 10.1016/s0040-8166(75)80007-3. [DOI] [PubMed] [Google Scholar]
  5. Fuller G. M., Brinkley B. R., Boughter J. M. Immunofluorescence of mitotic spindles by using monospecific antibody against bovine brain tubulin. Science. 1975 Mar 14;187(4180):948–950. doi: 10.1126/science.1096300. [DOI] [PubMed] [Google Scholar]
  6. Hsie A. W., Jones C., Puck T. T. Further changes in differentiation state accompanying the conversion of Chinese hamster cells of fibroblastic form by dibutyryl adenosine cyclic 3':5'-monophosphate and hormones. Proc Natl Acad Sci U S A. 1971 Jul;68(7):1648–1652. doi: 10.1073/pnas.68.7.1648. [DOI] [PMC free article] [PubMed] [Google Scholar]
  7. Hsie A. W., Puck T. T. Morphological transformation of Chinese hamster cells by dibutyryl adenosine cyclic 3':5'-monophosphate and testosterone. Proc Natl Acad Sci U S A. 1971 Feb;68(2):358–361. doi: 10.1073/pnas.68.2.358. [DOI] [PMC free article] [PubMed] [Google Scholar]
  8. Inoué S., Sato H. Cell motility by labile association of molecules. The nature of mitotic spindle fibers and their role in chromosome movement. J Gen Physiol. 1967 Jul;50(6 Suppl):259–292. [PMC free article] [PubMed] [Google Scholar]
  9. Johnson G. S., Friedman R. M., Pastan I. Restoration of several morphological characteristics of normal fibroblasts in sarcoma cells treated with adenosine-3':5'-cyclic monphosphate and its derivatives. Proc Natl Acad Sci U S A. 1971 Feb;68(2):425–429. doi: 10.1073/pnas.68.2.425. [DOI] [PMC free article] [PubMed] [Google Scholar]
  10. Monahan T. M., Fritz R. R., Abell C. W. Levels of cyclic AMP in murine L5178Y lymphoblasts grown in different concentrations of serum. Biochem Biophys Res Commun. 1973 Dec 10;55(3):642–646. doi: 10.1016/0006-291x(73)91192-3. [DOI] [PubMed] [Google Scholar]
  11. Monahan T. M., Marchand N. W., Fritz R. R., Abell C. W. Cyclic adenosine 3':5'-monophosphate levels and activities of related enzymes in normal and leukemic lymphocytes. Cancer Res. 1975 Sep;35(9):2540–2547. [PubMed] [Google Scholar]
  12. Oliver J. M., Zurier R. B., Berlin R. D. Concanavalin a cap formation on polymorphonuclear leukocytes of normal and beige (chediak-higashi) mice. Nature. 1975 Feb 6;253(5491):471–473. doi: 10.1038/253471a0. [DOI] [PubMed] [Google Scholar]
  13. Olmsted J. B., Borisy G. G. Ionic and nucleotide requirements for microtubule polymerization in vitro. Biochemistry. 1975 Jul;14(13):2996–3005. doi: 10.1021/bi00684a032. [DOI] [PubMed] [Google Scholar]
  14. Olmsted J. B., Borisy G. G. Microtubules. Annu Rev Biochem. 1973;42:507–540. doi: 10.1146/annurev.bi.42.070173.002451. [DOI] [PubMed] [Google Scholar]
  15. Porter K. R., Puck T. T., Hsie A. W., Kelley D. An electron microscopy study of the effects on dibutyryl cyclic AMP on Chinese hamster ovary cells. Cell. 1974 Jul;2(3):145–162. doi: 10.1016/0092-8674(74)90089-0. [DOI] [PubMed] [Google Scholar]
  16. Puck T. T., Waldren C. A., Hsie A. W. Membrane dynamics in the action of dibutyryl adenosine 3':5'-cyclic monophosphate and testosterone on mammalian cells. Proc Natl Acad Sci U S A. 1972 Jul;69(7):1943–1947. doi: 10.1073/pnas.69.7.1943. [DOI] [PMC free article] [PubMed] [Google Scholar]
  17. Rosenbaum J. L., Child F. M. Flagellar regeneration in protozoan flagellates. J Cell Biol. 1967 Jul;34(1):345–364. doi: 10.1083/jcb.34.1.345. [DOI] [PMC free article] [PubMed] [Google Scholar]
  18. 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]
  19. Sheppard J. R. Restoration of contact-inhibited growth to transformed cells by dibutyryl adenosine 3':5'-cyclic monophosphate. Proc Natl Acad Sci U S A. 1971 Jun;68(6):1316–1320. doi: 10.1073/pnas.68.6.1316. [DOI] [PMC free article] [PubMed] [Google Scholar]
  20. Soifer D., Laszlo A. H., Scotto J. M. Enzymatic activity in tubulin preparations. I. Intrinsic protein kinase activity in lyophilized preparations of tubulin from porcine brain. Biochim Biophys Acta. 1972 Jun 22;271(1):182–192. [PubMed] [Google Scholar]
  21. Tilney L. G., Porter K. R. Studies on the microtubules in heliozoa. II. The effect of low temperature on these structures in the formation and maintenance of the axopodia. J Cell Biol. 1967 Jul;34(1):327–343. doi: 10.1083/jcb.34.1.327. [DOI] [PMC free article] [PubMed] [Google Scholar]
  22. Weber K., Pollack R., Bibring T. Antibody against tuberlin: the specific visualization of cytoplasmic microtubules in tissue culture cells. Proc Natl Acad Sci U S A. 1975 Feb;72(2):459–463. doi: 10.1073/pnas.72.2.459. [DOI] [PMC free article] [PubMed] [Google Scholar]
  23. Weisenberg R. C. Microtubule formation in vitro in solutions containing low calcium concentrations. Science. 1972 Sep 22;177(4054):1104–1105. doi: 10.1126/science.177.4054.1104. [DOI] [PubMed] [Google Scholar]

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