Expression of a Constitutively Active Phosphatidylinositol 3-Kinase Induces Process Formation in Rat PC12 Cells
USE OF Cre/loxP RECOMBINATION SYSTEM*
- Michimoto Kobayashi‡,
- Satoshi Nagata‡,
- Yoshihiro Kita‡,
- Noriyuki Nakatsu‡,
- Sayoko Ihara‡,
- Kozo Kaibuchi§,
- Shinya Kuroda§,
- Motoyasu Ui¶,
- Hideo Iba¶,
- Hiroaki Konishi‖,
- Ushio Kikkawa‖,
- Izumu Saitoh** and
- Yasuhisa Fukui‡‡
- From the ‡Laboratory of Biological Chemistry, Department of Applied Biological Chemistry, Faculty of Agricultural and Life Science, University of Tokyo, 1-1-1 Yayoi-cho, Bunkyo-ku, Tokyo 113, Japan, the §Division of Signal Transduction, Nara Institute of Science and Technology, Ikoma 630-01, Nara, Japan, the ‖Biosignal Research Center, Kobe University, 1-1 Rokkodai-cho, Nada-ku, Kobe 657, Japan, and the ¶Department of Gene Regulation and **Laboratory of Molecular Genetics, Institute of Medical Science, University of Tokyo, Tokyo 108, Japan
Abstract
It has been shown that inhibition of phosphatidylinositol (PI) 3-kinase blocks neurite outgrowth of PC12 cells stimulated with nerve growth factor. To further assess the role of PI 3-kinase, the active form of PI 3-kinase was expressed in PC12 cells by the adenovirus mediated introduction of a site-specific recombinase, Cre. After expression of the active PI 3-kinase, elevation of the levels of PI 3,4-diphosphate and PI 3,4,5-trisphosphate as well as formation of neurite-like processes was observed. The process formation was inhibited by wortmannin, a selective inhibitor of PI 3-kinase, which suggests that a high activity of PI 3-kinase was responsible for the formation of these processes. The processes lacked accumulation of F-actin and GAP43 at the growth cone, which suggests that the processes were incomplete compared with neurites. Instead, the bundling of microtubules was enhanced, which suggests that organization of the microtubules might be driving the process of elongation in the cells expressing the active PI 3-kinase. Induction of active PI 3-kinase resulted in activation of Jun N-terminal kinase but not of mitogen-activated protein kinase or protein kinase B/Rac protein kinase/Akt. These results suggest that PI 3-kinase is involved in neurite outgrowth in PC12 cells and that activation of Jun N-terminal kinase cascade may be involved in the cell response.
Footnotes
-
↵* This work was supported by Grants-in-Aid 08275210 and 07456046 (to Y. F.) from the Ministry of Education, Science, Sports, and Culture of Japan.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. Tel.: 81-3-3812-2111, Ext. 5111; Fax: 81-3-3812-0544.
-
↵1 The abbreviations used are: NGF, nerve growth factor; MAP, mitogen-activated protein kinase; PI, phosphatidylinositol; PCR, polymerase chain reaction; PIPES, 1,4-piperazinediethanesulfonic acid; JNK, Jun N-terminal kinase; PKB, protein kinase B.
-
↵2 Y. Kita, K. Kimura, M. Kobayashi, S. Ihara, K. Kaibuchi, S. Kuroda, M. Ui, H. Iba, H. Konishi, U. Kikkawa, S. Nagata, and Y. Fukui, submitted for publication.
-
- Received February 24, 1997.
- Revision received May 8, 1997.
