The Journal of Cell Biology, Vol 76, Issue 2 547-555, Copyright © 1978 by The Rockefeller University Press
Proteins from morphologically differentiated neuroblastoma cells promote tubulin polymerization
N. W. Seeds and R. B. Maccioni
Department of Biophysics and Genetics, University of Colorado Medical Center, Denver 80262, USA.
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.
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