Volume 271, Number 43,
Issue of October 25, 1996
pp. 26989-26994
©1996 by The American Society for Biochemistry and Molecular Biology, Inc.
Synthesis and Accumulation of 
B Crystallin in C6 Glioma Cells
Is Induced by Agents That Promote the Disassembly of Microtubules
(Received for publication, December 26, 1995, and in revised form, July 1, 1996)
Kanefusa
Kato
,
Hidenori
Ito
,
Yutaka
Inaguma
,
Keiko
Okamoto
and
Shinsuke
Saga
¶
From the Departments of 
Biochemistry and
¶ Morphology, Institute for Developmental Research, Aichi Human
Service Center, Kasugai, Aichi 480-03, Japan
When C6 cells in culture were exposed at 37 °C
to 1 µM colchicine or to 1 µM colcemid, a
tubulin-binding antimitotic alkaloid, levels of 
B crystallin in
cells began to increase after about 10 h, reaching a maximum of
more than 1 µg/mg protein after 24 h. The level of B
crystallin returned to near the control level within two subsequent
days of culture in the normal medium. Northern blot analysis showed
that the accumulation of B crystallin was preceded by an increase in
the level of the mRNA for B crystallin. Nuclear run-off
transcription assays showed that colchicine induced new synthesis of
mRNA for B crystallin. Immunofluorescence staining revealed that
B crystallin accumulated in the peripheral areas of cells, as did
the depolymerized tubulin, after several hours of treatment with
colcemid, and then it gradually became more conspicuous in the
cytoplasm. Vinblastine and nocodazole, which also promote the
disassembly of microtubules by binding to tubulins, also induced the
synthesis of B crystallin. Furthermore, induction of B crystallin
by these drugs was observed in quiescent cells that had been cultured
in serum-free medium. However, taxol, a microtubule-stabilizing
antimitotic agent, did not stimulate the synthesis of B crystallin,
but rather, it suppressed the induction of synthesis of B crystallin
by the microtubule-disrupting drugs. Induction of B crystallin by
colchicine or by other drugs that promote the disassembly of
microtubules was sensitive to staurosporine, an inhibitor of protein
kinases, and the induction was completely suppressed in the presence of
10 nM staurosporine. These results suggest that the
expression of B crystallin is stimulated, via phosphorylation
reactions that are sensitive to staurosporine, when the
depolymerization of microtubules is enhanced.
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