Roles of C-terminal Src kinase in the initiation and the termination of the high affinity IgE receptor-mediated signaling.
As an attempt to analyze the roles of C-terminal Src kinase (Csk) in the high affinity IgE receptor (FcepsilonRI)-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 FcepsilonRI-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 FcepsilonRI/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 FcepsilonRI-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.
Honda
Z i
Zi
Department of Internal Medicine, Faculty of Medicine, University of Tokyo, 7-3-1, Hongo, Bunkyo-ku, Tokyo 113, Japan. honda-phy@h.u-tokyo.ac.jp
Suzuki
T
T
Hirose
N
N
Aihara
M
M
Shimizu
T
T
Nada
S
S
Okada
M
M
Ra
C
C
Morita
Y
Y
Ito
K
K
D000595:Amino Acid Sequence; D000818:Animals; D000081247:CSK Tyrosine-Protein Kinase; D002118:Calcium / Q000378:metabolism; D002460:Cell Line; D004789:Enzyme Activation; D007700:Kinetics; D019950:Mitogen-Activated Protein Kinase 1 / Q000378:metabolism; D008969:Molecular Sequence Data; D011505:Protein-Tyrosine Kinases / Q000378:metabolism*; D051381:Rats; D017455:Receptors, IgE / Q000378:metabolism*; D015398:Signal Transduction; D019061:src-Family Kinases / Q000378:metabolism
D017455:Receptors, IgE; D011505:Protein-Tyrosine Kinases; D000081247:CSK Tyrosine-Protein Kinase; C060902:lyn protein-tyrosine kinase; D019061:src-Family Kinases; D019950:Mitogen-Activated Protein Kinase 1; D002118:Calcium