Skip to main content
NCBI home page
The Journal of Cell Biology logoLink to The Journal of Cell Biology
. 1984 May 1;98(5):1763–1776. doi: 10.1083/jcb.98.5.1763

Role of the centrosome in organizing the interphase microtubule array: properties of cytoplasts containing or lacking centrosomes

PMCID: PMC2113175  PMID: 6725398

Abstract

To study the role of the centrosome in microtubule organization in interphase cells, we developed a method for obtaining cytoplasts (cells lacking a nucleus) that did or did not contain centrosomes. After drug- induced microtubule depolymerization, cytoplasts with centrosomes made from sparsely plated cells reconstituted a microtubule array typical of normal cells. Under these conditions cytoplasts without centrosomes formed only a few scattered microtubules. This difference in degree of polymerization suggests that centrosomes affect not only the distribution but the amount of microtubules in cells. To our surprise, the extent of microtubules assembled increased with the cell density of the original culture. At confluent density, cytoplasts without centrosomes had many microtubules, equivalent to cytoplasts with centrosomes. The additional microtubules were arranged peripherally and differed from the centrosomal microtubules in their sensitivity to nocodazole. These and other results suggest that the centrosome stabilizes microtubules in the cell, perhaps by capping one end. Microtubules with greater sensitivity to nocodazole arise by virtue of change in the growth state of the cell and may represent free or uncapped polymers. These experiments suggest that the spatial arrangement of microtubules may change by shifting the total tubulin concentration or the critical concentration for assembly.

Full Text

The Full Text of this article is available as a PDF (1.7 MB).

Selected References

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

  1. Albrecht-Buehler G. Autonomous movements of cytoplasmic fragments. Proc Natl Acad Sci U S A. 1980 Nov;77(11):6639–6643. doi: 10.1073/pnas.77.11.6639. [DOI] [PMC free article] [PubMed] [Google Scholar]
  2. Albrecht-Buehler G. Daughter 3T3 cells. Are they mirror images of each other? J Cell Biol. 1977 Mar;72(3):595–603. doi: 10.1083/jcb.72.3.595. [DOI] [PMC free article] [PubMed] [Google Scholar]
  3. Brinkley B. R., Cox S. M., Pepper D. A., Wible L., Brenner S. L., Pardue R. L. Tubulin assembly sites and the organization of cytoplasmic microtubules in cultured mammalian cells. J Cell Biol. 1981 Sep;90(3):554–562. doi: 10.1083/jcb.90.3.554. [DOI] [PMC free article] [PubMed] [Google Scholar]
  4. Brinkley B. R., Fuller E. M., Highfield D. P. Cytoplasmic microtubules in normal and transformed cells in culture: analysis by tubulin antibody immunofluorescence. Proc Natl Acad Sci U S A. 1975 Dec;72(12):4981–4985. doi: 10.1073/pnas.72.12.4981. [DOI] [PMC free article] [PubMed] [Google Scholar]
  5. Brooks R. F., Richmond F. N. Microtubule-organizing centres during the cell cycle of 3T3 cells. J Cell Sci. 1983 May;61:231–245. doi: 10.1242/jcs.61.1.231. [DOI] [PubMed] [Google Scholar]
  6. Calarco-Gillam P. D., Siebert M. C., Hubble R., Mitchison T., Kirschner M. Centrosome development in early mouse embryos as defined by an autoantibody against pericentriolar material. Cell. 1983 Dec;35(3 Pt 2):621–629. doi: 10.1016/0092-8674(83)90094-6. [DOI] [PubMed] [Google Scholar]
  7. De Brabander M. A model for the microtubule organizing activity of the centrosomes and kinetochores in mammalian cells. Cell Biol Int Rep. 1982 Oct;6(10):901–915. doi: 10.1016/0309-1651(82)90001-7. [DOI] [PubMed] [Google Scholar]
  8. De Brabander M., Geuens G., Nuydens R., Willebrords R., De Mey J. Microtubule assembly in living cells after release from nocodazole block: the effects of metabolic inhibitors, taxol and PH. Cell Biol Int Rep. 1981 Sep;5(9):913–920. doi: 10.1016/0309-1651(81)90206-x. [DOI] [PubMed] [Google Scholar]
  9. Frankel F. R. Organization and energy-dependent growth of microtubules in cells. Proc Natl Acad Sci U S A. 1976 Aug;73(8):2798–2802. doi: 10.1073/pnas.73.8.2798. [DOI] [PMC free article] [PubMed] [Google Scholar]
  10. 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]
  11. Giloh H., Sedat J. W. Fluorescence microscopy: reduced photobleaching of rhodamine and fluorescein protein conjugates by n-propyl gallate. Science. 1982 Sep 24;217(4566):1252–1255. doi: 10.1126/science.7112126. [DOI] [PubMed] [Google Scholar]
  12. Goldman R. D., Pollack R., Hopkins N. H. Preservation of normal behavior by enucleated cells in culture. Proc Natl Acad Sci U S A. 1973 Mar;70(3):750–754. doi: 10.1073/pnas.70.3.750. [DOI] [PMC free article] [PubMed] [Google Scholar]
  13. Gounon P., Karsenti E. Involvement of contractile proteins in the changes in consistency of oocyte nucleoplasm of the newt Pleurodeles waltlii. J Cell Biol. 1981 Feb;88(2):410–421. doi: 10.1083/jcb.88.2.410. [DOI] [PMC free article] [PubMed] [Google Scholar]
  14. Hill T. L., Kirschner M. W. Bioenergetics and kinetics of microtubule and actin filament assembly-disassembly. Int Rev Cytol. 1982;78:1–125. [PubMed] [Google Scholar]
  15. Hill T. L., Kirschner M. W. Regulation of microtubule and actin filament assembly--disassembly by associated small and large molecules. Int Rev Cytol. 1983;84:185–234. doi: 10.1016/s0074-7696(08)61018-9. [DOI] [PubMed] [Google Scholar]
  16. Hoebeke J., Van Nijen G., De Brabander M. Interaction of oncodazole (R 17934), a new antitumoral drug, with rat brain tubulin. Biochem Biophys Res Commun. 1976 Mar 22;69(2):319–324. doi: 10.1016/0006-291x(76)90524-6. [DOI] [PubMed] [Google Scholar]
  17. Lucas J. J., Szekely E., Kates J. R. The regeneration and division of mouse L-cell karyoplasts. Cell. 1976 Jan;7(1):115–122. doi: 10.1016/0092-8674(76)90261-0. [DOI] [PubMed] [Google Scholar]
  18. Marchisio P. C., Weber K., Osborn M. Identification of multiple microtubule initiating sites in mouse neuroblastoma cells. Eur J Cell Biol. 1979 Oct;20(1):45–50. [PubMed] [Google Scholar]
  19. Osborn M., Weber K. Cytoplasmic microtubules in tissue culture cells appear to grow from an organizing structure towards the plasma membrane. Proc Natl Acad Sci U S A. 1976 Mar;73(3):867–871. doi: 10.1073/pnas.73.3.867. [DOI] [PMC free article] [PubMed] [Google Scholar]
  20. Prescott D. M., Kates J., Kirkpatrick J. B. Replication of vaccinia virus DNA in enucleated L-cells. J Mol Biol. 1971 Aug 14;59(3):505–508. doi: 10.1016/0022-2836(71)90313-5. [DOI] [PubMed] [Google Scholar]
  21. ROBBINS E., GONATAS N. K. THE ULTRASTRUCTURE OF A MAMMALIAN CELL DURING THE MITOTIC CYCLE. J Cell Biol. 1964 Jun;21:429–463. doi: 10.1083/jcb.21.3.429. [DOI] [PMC free article] [PubMed] [Google Scholar]
  22. Schiff P. B., Fant J., Horwitz S. B. Promotion of microtubule assembly in vitro by taxol. Nature. 1979 Feb 22;277(5698):665–667. doi: 10.1038/277665a0. [DOI] [PubMed] [Google Scholar]
  23. Schiff P. B., Horwitz S. B. Taxol stabilizes microtubules in mouse fibroblast cells. Proc Natl Acad Sci U S A. 1980 Mar;77(3):1561–1565. doi: 10.1073/pnas.77.3.1561. [DOI] [PMC free article] [PubMed] [Google Scholar]
  24. Schliwa M., Pryzwansky K. B., Euteneuer U. Centrosome splitting in neutrophils: an unusual phenomenon related to cell activation and motility. Cell. 1982 Dec;31(3 Pt 2):705–717. doi: 10.1016/0092-8674(82)90325-7. [DOI] [PubMed] [Google Scholar]
  25. Sharp G. A., Osborn M., Weber K. Ultrastructure of multiple microtubule initiation sites in mouse neuroblastoma cells. J Cell Sci. 1981 Feb;47:1–24. doi: 10.1242/jcs.47.1.1. [DOI] [PubMed] [Google Scholar]
  26. Shay J. W., Gershenbaum M. R., Porter K. R. Enucleation of CHO cells by means of cytochalasin B and centrifugation: the topography of enucleation. Exp Cell Res. 1975 Aug;94(1):47–55. doi: 10.1016/0014-4827(75)90529-7. [DOI] [PubMed] [Google Scholar]
  27. Shay J. W., Porter K. R., Prescott D. M. The surface morphology and fine structure of CHO (Chinese hamster ovary) cells following enucleation. Proc Natl Acad Sci U S A. 1974 Aug;71(8):3059–3063. doi: 10.1073/pnas.71.8.3059. [DOI] [PMC free article] [PubMed] [Google Scholar]
  28. Sherline P., Mascardo R. Epidermal growth factor-induced centrosomal separation: mechanism and relationship to mitogenesis. J Cell Biol. 1982 Oct;95(1):316–322. doi: 10.1083/jcb.95.1.316. [DOI] [PMC free article] [PubMed] [Google Scholar]
  29. Solomon F. Neuroblastoma cells recapitulate their detailed neurite morphologies after reversible microtubule disassembly. Cell. 1980 Sep;21(2):333–338. doi: 10.1016/0092-8674(80)90469-9. [DOI] [PubMed] [Google Scholar]
  30. Spiegelman B. M., Lopata M. A., Kirschner M. W. Aggregation of microtubule initiation sites preceding neurite outgrowth in mouse neuroblastoma cells. Cell. 1979 Feb;16(2):253–263. doi: 10.1016/0092-8674(79)90003-5. [DOI] [PubMed] [Google Scholar]
  31. Spiegelman B. M., Lopata M. A., Kirschner M. W. Multiple sites for the initiation of microtubule assembly in mammalian cells. Cell. 1979 Feb;16(2):239–252. doi: 10.1016/0092-8674(79)90002-3. [DOI] [PubMed] [Google Scholar]
  32. Tuffanelli D. L., McKeon F., Kleinsmith D. M., Burnham T. K., Kirschner M. Anticentromere and anticentriole antibodies in the scleroderma spectrum. Arch Dermatol. 1983 Jul;119(7):560–566. [PubMed] [Google Scholar]
  33. Weber K., Rathke P. C., Osborn M. Cytoplasmic microtubular images in glutaraldehyde-fixed tissue culture cells by electron microscopy and by immunofluorescence microscopy. Proc Natl Acad Sci U S A. 1978 Apr;75(4):1820–1824. doi: 10.1073/pnas.75.4.1820. [DOI] [PMC free article] [PubMed] [Google Scholar]
  34. Witt D. P., Gordon J. A. Most iodinatable fibroblast surface proteins accompany the cytoplast membrane during cytochalasin B-mediated enucleation of chick embryo fibroblasts. J Cell Biol. 1982 Sep;94(3):557–564. doi: 10.1083/jcb.94.3.557. [DOI] [PMC free article] [PubMed] [Google Scholar]
  35. Zorn G. A., Lucas J. J., Kates J. R. Purification and characterization of regenerating mouse L929 karyoplasts. Cell. 1979 Nov;18(3):659–672. doi: 10.1016/0092-8674(79)90121-1. [DOI] [PubMed] [Google Scholar]

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

RESOURCES