Abstract
Stimulation of the T cell antigen receptor (TCR) activates a protein tyrosine kinase and leads to the tyrosine phosphorylation of phosphoinositide-specific phospholipase C-gamma 1 (PLC gamma 1). The molecular interactions involved in this phosphorylation are not known. After stimulation of the TCR on Jurkat T cells, tyrosine-phosphorylated proteins of 36, 38, 58, and 63 kD coprecipitate with PLC gamma 1. An identical pattern of proteins precipitate with TrpE fusion proteins that contain the Src homology (SH) 2 domains of PLC gamma 1, indicating that these regions of PLC gamma 1 are responsible for binding. TCR stimulation leads to an association between the SH2 domains of PLC gamma 1 and a protein tyrosine kinase, which, by peptide mapping, is identical to p56lck. These studies establish that p56lck associates with PLC gamma 1 as a result of TCR stimulation of Jurkat cells, suggesting that p56lck plays a central role in coupling the TCR to the activation of PLC gamma 1.
Highlights
From the Department of Medicing University of California, and the VeteransAffairs Medical Center, San Franciscg California 94121; and the *Division of Molecular and Developmental Biology, Samuel Lunenfeld Research Institutg Mount Sinai Hospital, Toronto, Ontarig M5G IX5, Canada
To determine whether the coprecipitation of these rnolecules is a result of their binding to the Src homology 2 (SH2) domains of phospholipase C (PLC)'y1, we used fusion proteins of TrpE and PI.C'y1 SH2 domains as affinity ligands
Comparable levels of all four tyrosine-phosphorylated proteins bind to a fusion prorein that has only the NH2-terminal SH2 domain
Summary
TCR-mediated PI turnover is sensitive to PTK inhibitors, suggesting that tyrosine phosphorylation regulates PLC activity [6, 7] Consistent with this possibility, stimulation of the TCK on Jurkat T cells leads to the tyrosine phosphorylation of PLC~/1 and, subsequently, to an association between PLCv1 and the CD3 components of the TCR (8-t2). Critical to the activation of PLC-/1 by growth factor receptor protein tyrosine kinases, such as the receptors for epidermal growth factor (EGF) and platdetderived growth factor (PDGF), is the direct binding of PLCT1 to the receptor [13,14,15,16,17] This binding requires autophosphorylation of the receptor and appears to be mediated by the Src homology 2 (SH2) domains of PLC3,1 [18,19,20]. We have examined the possibilities that PLC'y1 associates with tyrosine phosphorylated molecules as a result of TCR stimulation of Jurkat cells, and that this association involves the SH2 domains of PLC'yl
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