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J Biol Chem, Vol. 275, Issue 17, 12572-12580, April 28, 2000

Simultaneous Suppression of cdc2 and cdk2 Activities Induces Neuronal Differentiation of PC12 Cells*

Yoh DobashiDagger §, Mitsuhiko ShojiDagger , Masatoshi Kitagawa||, Takashi NoguchiDagger , and Toru KameyaDagger

From the Dagger  Department of Pathology, Kitasato University School of Medicine, 1-15-1, Kitasato, Sagamihara, Kanagawa 228-8555, Japan, the  Department of Molecular and Cellular Biology, Medical Institute for Bioregulation, Kyushu University, Fukuoka 812-8582, Japan, and the || CREST, Japan Science and Technology Corporation (JST), Kawaguchi, Saitama 332-0012, Japan

The involvement of cdc2 and cdk2 during neuronal differentiation in rat pheochromocytoma PC12 cells was examined. When PC12 cells were cultured with nerve growth factor (NGF), expression of cdc2 decreased significantly after day 5, while expression of cdk2 decreased gradually after day 7. Cells overexpressing cdc2 or cdk2 were resistant to NGF-induced differentiation and growth suppression, and maintained high cdc2 or cdk2 kinase activity, respectively, during NGF treatment. In contrast, the NGF-treated parental cells showed a marked decline in these kinase activities after day 3. When PC12 cells were treated with specific inhibitors of cdc2/cdk2 (butyrolactone-I, olomoucin), they showed marked neurite extension and up-regulation of microtubule-associated protein 2 expression. In addition, treatment with mixtures of antisense oligonucleotides for cdc2 and cdk2 resulted in down-regulation of both cdc2 and cdk2 kinase activities as well as significant neurite outgrowth and up-regulation of microtubule-associated protein 2 expression. However, neurite outgrowth was not observed in cells treated with either single antisense oligonucleotide, or antisense cdc2 + cdk4 or cdk2 + cdk4 oligonucleotide mixtures. These results suggest that simultaneous down-regulation of cdc2 and cdk2 activity is sufficient and necessary for neuronal differentiation in PC12 cells.

* This work was supported by Grant-in-aid for Scientific Research 11670191 from the Ministry of Education, Science and Culture in Japan, a Mitsui Life Social Welfare Foundation Research Grant, and Kitasato University Medical Science Research Project 01A-1999.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 should be addressed: Dept. of Pathology, Kitasato University School of Medicine, 1-15-1, Sagamihara, Kanagawa 228-8555, Japan. Tel.: 81-42-778-9020; Fax: 81-42-778-8441; E-mail: ydobashi@med.kitasato-u.ac.jp.

Copyright © 2000 by The American Society for Biochemistry and Molecular Biology, Inc.
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