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T Cell Receptor-induced Activation of Phospholipase C-1 Depends on a Sequence-independent Function of the P-I Region of SLP-76*
Ronnie Gonen,
Dvora Beach,
Carmit Ainey, , and
Deborah Yablonski
Rappaport Faculty of Medicine, Technion-Israel Institute of Technology, P. O. Box 9649, Bat Galim, Haifa 31096, Israel
Abstract:
SLP-76 forms part of a hematopoietic-specific adaptor proteincomplex, and is absolutely required for T cell development andactivation. T cell receptor (TCR)-induced activation of phospholipaseC-1 (PLC-1) depends on three features of SLP-76: the N-terminaltyrosine phosphorylation sites, the Gads-binding site, and anintervening sequence, denoted the P-I region, which binds tothe SH3 domain of PLC-1 (SH3PLC) via a low affinity interaction.Despite extensive research, the mechanism whereby SLP-76 regulatesPLC-1 remains uncertain. In this study, we uncover and explorean apparent paradox: whereas the P-I region as a whole is essentialfor TCR-induced activation of PLC-1 and nuclear factor of activatedT cells (NFAT), no particular part of this region is absolutelyrequired. To better understand the contribution of the P-I regionto PLC-1 activation, we mapped the PLC-1-binding site withinthe region, and created a SLP-76 mutant that fails to bind SH3PLC,but is fully functional, mediating TCR-induced phosphorylationof PLC-1 at tyrosine 783, calcium flux, and nuclear factor ofactivated T cells activation. Unexpectedly, full functionalityof this mutant was maintained even under less than optimal stimulationconditions, such as a low concentration of the anti-TCR antibody.Another SLP-76 mutant, in which the P-I region was scrambledto abolish any sequence-dependent protein-binding motifs, alsoretained significant functionality. Our results demonstratethat SLP-76 need not interact with SH3PLC to activate PLC-1,and further suggest that the P-I region of SLP-76 serves a structuralrole that is sequence-independent and is not directly relatedto protein-protein interactions.
Received for publication August 17, 2004.
Revision received December 13, 2004.
* This work was supported by German-Israeli Foundation for ScientificResearch and Development Grant I-2022-1045.11/2000, the MatildaBarnett Revocable Trust, the Technion V.P.R.-Charles Crown ResearchFund, and the Fund for the Promotion of Research at the Technion.The costs of publication of this article were defrayed in partby the payment of page charges. This article must thereforebe hereby marked "advertisement" in accordance with 18 U.S.C.Section 1734 solely to indicate this fact.
To whom correspondence should be addressed: Rappaport Faculty of Medicine, Technion-Israel Institute of Technology, P. O. Box 9649, Bat Galim, Haifa 31096, Israel. Tel.: 972-4-829-5393; Fax: 972-4-829-5255; E-mail: debya{at}tx.technion.ac.il.
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1 Depends on a Sequence-independent Function of the P-I Region of SLP-76
1 (PLC-
To whom correspondence should be addressed: Rappaport Faculty of Medicine, Technion-Israel Institute of Technology, P. O. Box 9649, Bat Galim, Haifa 31096, Israel. Tel.: 972-4-829-5393; Fax: 972-4-829-5255; E-mail: 
