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J. Biol. Chem., Vol. 278, Issue 15, 13487-13495, April 11, 2003
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From the Axin regulates Wnt signaling through
down-regulation of
Ectopic Expression of Axin Blocks Neuronal Differentiation of
Embryonic Carcinoma P19 Cells*
,
, and**
Department of Ophthalmology, Catholic
University of Korea, Seoul 137-040, Korea, the
§ Department of Genetics and Development, Columbia
University, New York, New York 10032, and the ¶ Department of Life
Science, The University of Seoul, Seoul 130-743, Korea
-catenin. To test the role of Wnt signaling in
neuronal differentiation, embryonal carcinoma P19 cells (P19 EC), which
can be stimulated to differentiate into a neuron-like phenotype in
response to retinoic acid (RA), were used. Reverse transcription-PCR
and Western blot analysis showed that Axin is expressed in
undifferentiated cells, whereas the level is clearly reduced during
RA-induced neuronal differentiation. Interestingly, Axin levels were
not reduced during endodermal differentiation of P19 EC cells and F9 EC
cells by RA, suggesting that the reduction of the Axin level is a
specific property of neuronal differentiation. Western analysis showed that the cytoplasmic level of -catenin increased during neuronal differentiation of P19 EC cells. Indirect immunofluorescence with -catenin antibody showed that the localization of -catenin was changed from membrane in undifferentiated cells to nuclei in neuronal P19 EC cells. Induced expression of Axin during endodermal and early
neuronal differentiation, using the Tet-On system, did not block
normal differentiation. However, maintenance of the Axin level blocked
neuronal differentiation and inhibited expression of a neuron-specific
marker protein, III-tubulin. Also, ectopic induction of a
-catenin signaling inhibitor, ICAT, inhibited expression of
III-tubulin. In contrast, addition of Wnt-3A-conditioned medium
during the neuronal differentiation period enhanced the expression of
III-tubulin. Overall, our data show that Wnt-3a/canonical -catenin signaling through the down-regulation of Axin may play an
important role in neuronal differentiation.
*
This study was supported by the Biomedical Brain Research
Center, Ministry of Health & Welfare, Republic of Korea (Grant
01-PJ8-PG6-01NE01-0003 to E. J.) and the National Institutes of Health
(to F. C.).The costs of publication of this
article were defrayed in part by the
payment of page charges. The article
must therefore be hereby marked
"advertisement" in
accordance with 18 U.S.C. Section
1734 solely to indicate this fact.
To whom correspondence may be addressed. Tel.: 822-2210-2681;
Fax: 822-2210-2888; E-mail: ej70@uos.ac.kr.
**
To whom correspondence may be addressed. Tel.: 822-590-2613; Fax:
822-533-3801; E-mail: ckjoo@catholic.ac.kr.
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