Roles of C-terminal Src Kinase in the Initiation and the Termination of the High Affinity IgE Receptor-mediated Signaling*
- Zen-ichiro Honda‡,
- Takeshi Suzuki,
- Naoto Hirose,
- Makoto Aihara§,
- Takao Shimizu§,
- Shigeyuki Nada¶,
- Masato Okada¶,
- Chisei Ra‖,
- Yutaka Morita and
- Koji Ito
- From the Departments of Internal Medicine and Physical Therapy and of §Biochemistry, Faculty of Medicine, University of Tokyo, 7-3-1, Hongo, Bunkyo-ku, Tokyo 113, Japan, the ¶Division of Protein Metabolism, Institute of Protein Research, Osaka University, Suita, Osaka 565, Japan, and the ‖Department of Immunology, Juntendo University, School of Medicine, 2-1-1, Hongo Bunkyo-ku Tokyo 113, Japan
Abstract
As an attempt to analyze the roles of C-terminal Src kinase (Csk) in the high affinity IgE receptor (FcεRI)-mediated signaling, we overexpressed Csk, a membrane-targeted form of Csk (mCsk), and a kinase-defective, membrane-targeted form of Csk (mCsk(−)) in rat basophil leukemia (RBL) 2H3 cells. Specific activity of Lyn at the basal state was decreased in Csk-expressing cells, and further decreased in mCsk-expressing cells. In mCsk(−)-expressing cells, basal specific activity of Lyn was increased, thereby indicating that mCsk(−) functioned as a dominant negative molecule. The onset of FcεRI-mediated Lyn activation was delayed in Csk-expressing cells, and further delayed in mCsk-expressing cells. In mCsk(−)-expressing cells, Lyn activation was rapid and quite long lasting. These findings indicate (i) Csk negatively regulates rapid FcεRI/Lyn coupling, and (ii) Csk activity is potentially required for its termination. The onsets of the series of events including tyrosyl phosphorylation of Syk, mitogen-activated protein (MAP) kinase activation, elevation of intracellular calcium concentration ([Ca2+]i), and histamine release were all stepwisely delayed in Csk-expressing cells and in mCsk-expressing cells. The durations of Syk phosphorylation and MAP kinase activation also closely correlated with those of Lyn activation, but [Ca2+]i elevation and histamine release followed different temporal patterns: the delayed responses in Csk-expressing cells and in mCsk-expressing cells led to sustained [Ca2+]i oscillation and histamine release, while the prompt responses in parent cells and mCsk(−)-expressing cells rapidly subsided. These findings provide further evidence that the initiations of the FcεRI-mediated signals are upstreamly regulated by Src family protein tyrosine kinases and revealed that their terminations are regulated by Lyn-dependent (Syk and MAP kinase) and -independent ([Ca2+]i elevation and histamine release) mechanisms.
Footnotes
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↵* This work was supported in part by Grants-in-aid from the Ministry of Education, Science, Sports and Culture and by grants from the Ono Medical Research Foundation, the Manabe Research Foundation, and the Uehara Memorial Foundation.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.
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↵‡ To whom correspondence should be addressed. Tel.: 81-3-3815-5411; Fax: 81-3-3815-5954; E-mail: honda-phy{at}h.u-tokyo.ac.jp.
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↵1 The abbreviations used are: FcεRI, the high affinity IgE receptor; ITAM, immunoreceptor tyrosine-based activation motif; PTK, protein tyrosine kinase; Csk, C-terminal Src kinase; mCsk, membrane-targeted Csk; mCsk(−), membrane-targeted, kinase-defective Csk; RBL, rat basophil leukemia; MAP kinase, mitogen-activated protein kinase; TCR and BCR, T and B cell receptor, respectively; SH, Src homology; anti-DNP IgE, anti-dinitrophenyl IgE; DNP-BSA, 2,4-dinitrophenylated bovine serum albumin; DMEM, Dulbecco’s modified Eagle’s medium; WT, wild-type.
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↵2 Z. Honda, T. Suzuki, and T. Yamamoto, unpublished observation.
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- Received January 30, 1997.
- Revision received July 8, 1997.
