The mitogenic response of T cells to interleukin-2 requires Raf-1.
The product of the c-raf-1 proto-oncogene, Raf-1, is known to encode a 74-kDa ubiquitously expressed cytoplasmic serine/threonine kinase. Various growth factors such as epidermal growth factor, acidic fibroblast growth factor, platelet-derived growth factor, insulin, granulocyte-macrophage colony-stimulating factor, interleukin (IL)-2, IL-3 and erythropoietin have been shown to induce phosphorylation of Raf-1, thereby activating Raf-1 kinase. Raf-1 is, thus, believed to play a role in coupling growth factor receptors to proliferation. We have examined the role of Raf-1 in the mitogenic response of human peripheral blood-derived IL-2 receptor expressing T cells to human recombinant IL-2 employing c-raf antisense (AS) oligodeoxyribonucleotide. Uptake studies of oligonucleotides indicated that incorporation of oligomers was maximal at 4 h and oligodeoxynucleotides remained stable in these cells for up to 24 h. Treatment of T cells with the AS oligodeoxyribonucleotide in intracellular duplex formation followed by efficient translation blockade of c-raf-1. In contrast, sense (S) and nonsense (NS) oligodeoxynucleotides failed to form intracellular duplexes and did not interfere with translation of c-raf-1, suggesting specific elimination of c-raf-1 by the AS oligomer. Proliferation of T cells ([3H]thymidine incorporation) following exposure to IL-2 was substantially reduced when the c-raf-1 AS oligodeoxyribonucleotide was added to cultures, while the mitogenic response to this factor remained almost unaffected in the presence of S and NS oligodeoxyribonucleotides.
Riedel
D
D
Department of Medical Oncology and Applied Molecular Biology, Freie Universität Berlin, FRG.
Brennscheidt
U
U
Kiehntopf
M
M
Brach
M
M
Herrmann
F
F
D001483:Base Sequence; D006801:Humans; D007376:Interleukin-2 / Q000494:pharmacology*; D008213:Lymphocyte Activation / Q000187:drug effects*; D008934:Mitogens / Q000494:pharmacology*; D008969:Molecular Sequence Data; D016376:Oligonucleotides, Antisense / Q000494:pharmacology; D017346:Protein Serine-Threonine Kinases / Q000502:physiology*; D000090063:Proto-Oncogene Mas; D011518:Proto-Oncogene Proteins / Q000502:physiology*; D019908:Proto-Oncogene Proteins c-raf; D011994:Recombinant Proteins / Q000494:pharmacology; D013601:T-Lymphocytes / Q000187:drug effects*
D007376:Interleukin-2; C000715470:MAS1 protein, human; D008934:Mitogens; D016376:Oligonucleotides, Antisense; D000090063:Proto-Oncogene Mas; D011518:Proto-Oncogene Proteins; D011994:Recombinant Proteins; D017346:Protein Serine-Threonine Kinases; D019908:Proto-Oncogene Proteins c-raf