Skip to main content
NCBI home page
The Journal of Cell Biology logoLink to The Journal of Cell Biology
. 1978 Feb 1;76(2):547–555. doi: 10.1083/jcb.76.2.547

Proteins from morphologically differentiated neuroblastoma cells promote tubulin polymerization

N W Seeds 1, R B Maccioni 1
PMCID: PMC2109974  PMID: 10605456

Abstract

Clonal cells (N18) of the mouse neuroblastoma C-1300 can be induced to undergo a morphological differentiation characterized by the outgrowth of very long neurites (> 150 microns) that contain many microtubules. Because the marked increase in the number and length of microtubules is apparently not due to an increase in the concentration of tubulin subunits, the possible role of additional macromolecules in the regulation of tubulin polymerization during neurite formation by N18 cells was examined. Using an in vitro system where the polymerization of low concentrations (< 4 mg/ml) of purified brain tubulin requires microtubule-associated proteins (MAPs), high-speed supernates (250,000 g) from neuroblastoma and glioma cells were assayed for their ability to replace MAPs in the polymerization of brain tubulin. Only the supernates from "differentiated" N18 cells were polymerization competent. Electron microscope observations of these supernates failed to demonstrate the presence of nucleation structures (rings or disks). The active factor(s) sedimented at approximately 7S on sucrose gradient centrifugation and eluted from 4B Sepharose in the region of 170,000 mol wt proteins. Furthermore, the inactive supernates from other cells did not inhibit polymerization when tested in the presence of limiting MAPs. Thus, microtubule formation accompanying neurite outgrowth in neuroblastoma cells appears to be regulated by the presence of additional macromolecular factor(s) that may be functionally equivalent to the MAPs found with brain microtubules.

Full Text

The Full Text of this article is available as a PDF (1.2 MB).

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. Bryan J. B., Nagle B. W., Doenges K. H. Inhibition of tubulin assembly by RNA and other polyanions: evidence for a required protein. Proc Natl Acad Sci U S A. 1975 Sep;72(9):3570–3574. doi: 10.1073/pnas.72.9.3570. [DOI] [PMC free article] [PubMed] [Google Scholar]
  3. Fellous A., Francon J., Lennon A. M., Nunez J., Osty J., Chantoux F. Initiation of neurotubulin polymerisation and rat brain development. FEBS Lett. 1976 May 1;64(2):400–403. doi: 10.1016/0014-5793(76)80336-5. [DOI] [PubMed] [Google Scholar]
  4. Gaskin F., Cantor C. R., Shelanski M. L. Turbidimetric studies of the in vitro assembly and disassembly of porcine neurotubules. J Mol Biol. 1974 Nov 15;89(4):737–755. doi: 10.1016/0022-2836(74)90048-5. [DOI] [PubMed] [Google Scholar]
  5. Keates R. A., Hall R. H. Tubulin requires an accessory protein for self assembly in microtubules. Nature. 1975 Oct 2;257(5525):418–421. doi: 10.1038/257418a0. [DOI] [PubMed] [Google Scholar]
  6. LOWRY O. H., ROSEBROUGH N. J., FARR A. L., RANDALL R. J. Protein measurement with the Folin phenol reagent. J Biol Chem. 1951 Nov;193(1):265–275. [PubMed] [Google Scholar]
  7. Maccioni R., Seeds N. W. Stoichiometry of GTP hydrolysis and tubulin polymerization. Proc Natl Acad Sci U S A. 1977 Feb;74(2):462–466. doi: 10.1073/pnas.74.2.462. [DOI] [PMC free article] [PubMed] [Google Scholar]
  8. Mizel S. B., Bamburg J. R. Studies on the action of nerve growth factor. II. Neurotubule protein levels during neurite outgrowth. Neurobiology. 1975 Oct;5(5):283–290. [PubMed] [Google Scholar]
  9. Morgan J. L., Seeds N. W. Properties of tubulin prepared by affinity chromatography. Ann N Y Acad Sci. 1975 Jun 30;253:260–271. doi: 10.1111/j.1749-6632.1975.tb19205.x. [DOI] [PubMed] [Google Scholar]
  10. Morgan J. L., Seeds N. W. Tubulin constancy during morphological differentiation of mouse neuroblastoma cells. J Cell Biol. 1975 Oct;67(1):136–145. doi: 10.1083/jcb.67.1.136. [DOI] [PMC free article] [PubMed] [Google Scholar]
  11. Olmsted J. B., Carlson K., Klebe R., Ruddle F., Rosenbaum J. Isolation of microtubule protein from cultured mouse neuroblastoma cells. Proc Natl Acad Sci U S A. 1970 Jan;65(1):129–136. doi: 10.1073/pnas.65.1.129. [DOI] [PMC free article] [PubMed] [Google Scholar]
  12. Olmsted J. B., Marcum J. M., Johnson K. A., Allen C., Borisy G. G. Microtuble assembly: some possible regulatory mechanisms. J Supramol Struct. 1974;2(2-4):429–450. doi: 10.1002/jss.400020230. [DOI] [PubMed] [Google Scholar]
  13. Seeds N. W., Gilman A. G., Amano T., Nirenberg M. W. Regulation of axon formation by clonal lines of a neural tumor. Proc Natl Acad Sci U S A. 1970 May;66(1):160–167. doi: 10.1073/pnas.66.1.160. [DOI] [PMC free article] [PubMed] [Google Scholar]
  14. Sloboda R. D., Dentler W. L., Rosenbaum J. L. Microtubule-associated proteins and the stimulation of tubulin assembly in vitro. Biochemistry. 1976 Oct 5;15(20):4497–4505. doi: 10.1021/bi00665a026. [DOI] [PubMed] [Google Scholar]
  15. Snyder J. A., McIntosh J. R. Biochemistry and physiology of microtubules. Annu Rev Biochem. 1976;45:699–720. doi: 10.1146/annurev.bi.45.070176.003411. [DOI] [PubMed] [Google Scholar]
  16. Stöckel K., Solomon F., Paravicini U., Thoenen H. Dissociation between effects of nerve growth factor on tyrosine hydrolase and tubulin synthesis in sympathetic ganglia. Nature. 1974 Jul 12;250(462):150–151. doi: 10.1038/250150a0. [DOI] [PubMed] [Google Scholar]
  17. Weingarten M. D., Lockwood A. H., Hwo S. Y., Kirschner M. W. A protein factor essential for microtubule assembly. Proc Natl Acad Sci U S A. 1975 May;72(5):1858–1862. doi: 10.1073/pnas.72.5.1858. [DOI] [PMC free article] [PubMed] [Google Scholar]
  18. Wiche G., Cole R. D. Reversible in vitro polymerization of tubulin from a cultured cell line (rat glial cell clone C6). Proc Natl Acad Sci U S A. 1976 Apr;73(4):1227–1231. doi: 10.1073/pnas.73.4.1227. [DOI] [PMC free article] [PubMed] [Google Scholar]
  19. Yamada K. M., Wessells N. K. Axon elongation. Effect of nerve growth factor on microtubule protein. Exp Cell Res. 1971 Jun;66(2):346–352. doi: 10.1016/0014-4827(71)90687-2. [DOI] [PubMed] [Google Scholar]

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

RESOURCES