Rapid activation of C-Raf-1 after stimulation of the T-cell receptor or the muscarinic receptor type 1 in resting T cells.
The c-Raf-1 serine/threonine kinase is an important component of signal transduction pathways mediating the effects of a variety of growth factors. In activated T cells, IL-2 has been shown to induce activation of c-Raf-1, but c-Raf-1 has not previously been shown to be activated through the T-cell receptor (TCR) in resting G0 T cells. Using a sensitive immune complex kinase reaction, we show that cross-linking of the stimulatory and costimulatory receptors CD3, CD4, or CD28 induces c-Raf-1 activation in highly purified resting peripheral blood human T cells. In contrast, cross-linking the nonstimulatory receptor CD45 did not induce c-Raf-1. Surprisingly, although earlier studies had shown delayed kinetics in response to Thy-1 stimulation in murine cells, c-Raf-1 activation in response to CD3 cross-linking was one of the earliest measurable events. In spite of its early kinetics, c-Raf-1 activation was found to be downstream of several other early signal transduction events, including activation of a tyrosine kinase and a tyrosine phosphatase. Several lines of evidence suggest that activation of c-Raf-1 in response to TCR stimulation may be PKC-dependent: first, phorbol esters are extremely potent activators of c-Raf-1 in human T cells; second, the kinetics of accumulation of products of phosphatidylinositol hydrolysis coincides with the kinetics of c-Raf-1 activation; and third, physiologic activation of the PLC/PKC pathway through a transfected, G-protein-coupled receptor HM1 induced similar levels of c-Raf-1 activation with a similar time course. We conclude that c-Raf-1 activation is tightly coupled to TCR stimulation and may participate in signal transduction pathways in resting, G0 T cells. The observation that the HM1 receptor can also activate c-Raf-1 suggests that T cells have the capability to utilize both tyrosine kinase-dependent and tyrosine kinase-independent mechanisms of c-Raf-1 activation.
Siegel
J N
JN
Immune Cell Biology Program, Naval Medical Research Institute, Bethesda, MD 20814.
June
C H
CH
Yamada
H
H
Rapp
U R
UR
Samelson
L E
LE
D000818:Animals; D018106:CD28 Antigens / Q000502:physiology; D017252:CD3 Complex / Q000502:physiology; D015704:CD4 Antigens / Q000502:physiology; D004789:Enzyme Activation; D006801:Humans; D051379:Mice; D010716:Phosphatidylinositols / Q000378:metabolism; D017346:Protein Serine-Threonine Kinases / Q000378:metabolism*; D054562:Protein Tyrosine Phosphatase, Non-Receptor Type 1; D017027:Protein Tyrosine Phosphatases / Q000502:physiology; D011505:Protein-Tyrosine Kinases / Q000502:physiology; D011518:Proto-Oncogene Proteins / Q000378:metabolism*; D019908:Proto-Oncogene Proteins c-raf; D011948:Receptors, Antigen, T-Cell / Q000502:physiology*; D011976:Receptors, Muscarinic / Q000502:physiology*; D013601:T-Lymphocytes / Q000378:metabolism*; D013755:Tetradecanoylphorbol Acetate / Q000494:pharmacology
D018106:CD28 Antigens; D017252:CD3 Complex; D015704:CD4 Antigens; D010716:Phosphatidylinositols; D011518:Proto-Oncogene Proteins; D011948:Receptors, Antigen, T-Cell; D011976:Receptors, Muscarinic; D011505:Protein-Tyrosine Kinases; D017346:Protein Serine-Threonine Kinases; D019908:Proto-Oncogene Proteins c-raf; D054562:Protein Tyrosine Phosphatase, Non-Receptor Type 1; D017027:Protein Tyrosine Phosphatases; D013755:Tetradecanoylphorbol Acetate