Raf-1 is a necessary component of the mitogenic response of the human megakaryoblastic leukemia cell line MO7 to human stem cell factor, granulocyte-macrophage colony-stimulating factor, interleukin 3, and interleukin 9.
We have examined the role of Raf-1 in the mitogenic response of the factor-deprived human megakaryoblastic leukemia cell line MO7 to recombinant human granulocyte-macrophage colony-stimulating factor, interleukin 3, interleukin 9, and stem cell factor by using c-raf antisense oligodeoxyribonucleotides. Uptake of oligodeoxyribonucleotides by MO7 cells was maximal at 5-10 h in culture, and oligomers remained stable in these cells for at least 24 h. Treatment of MO7 cells with the antisense oligomer resulted in intracellular oligomer/mRNA duplex formation followed by efficient translation blockade of c-raf-1. In contrast, sense and non-sense oligodeoxyribonucleotides failed to form intracellular duplexes and did not interfere with translation of c-raf-1, suggesting specific elimination of c-raf-1 by the antisense oligodeoxyribonucleotide. Furthermore, exposure of MO7 cells to c-raf-1 antisense prevented factor-induced nuclear translocation of Raf-1. Most importantly, proliferation of MO7 cells ([3H]thymidine incorporation) enabled by these growth factors was significantly reduced when the c-raf-1 antisense oligodeoxyribonucleotide was added to cultures, whereas the mitogenic response to these factors remained almost unaffected in the presence of sense and non-sense oligodeoxyribonucleotides.
Brennscheidt
U
U
Department of Medical Oncology and Applied Molecular Biology, Free University of Berlin, Germany.
Riedel
D
D
Kölch
W
W
Bonifer
R
R
Brach
M A
MA
Ahlers
A
A
Mertelsmann
R H
RH
Herrmann
F
F
D001483:Base Sequence; D002455:Cell Division / Q000187:drug effects; D003062:Codon / Q000235:genetics; D016178:Granulocyte-Macrophage Colony-Stimulating Factor / Q000494:pharmacology; D006133:Growth Substances / Q000494:pharmacology*; D016298:Hematopoietic Cell Growth Factors / Q000494:pharmacology; D006801:Humans; D007377:Interleukin-3 / Q000494:pharmacology; D016906:Interleukin-9 / Q000494:pharmacology; D007947:Leukemia, Megakaryoblastic, Acute / Q000473:pathology / Q000503:physiopathology*; D008934:Mitogens / Q000494:pharmacology*; D008969:Molecular Sequence Data; D016376:Oligonucleotides, Antisense; D017346:Protein Serine-Threonine Kinases / Q000502:physiology*; D011518:Proto-Oncogene Proteins / Q000502:physiology*; D019908:Proto-Oncogene Proteins c-raf; D011994:Recombinant Proteins / Q000494:pharmacology; D019089:Stem Cell Factor; D014407:Tumor Cells, Cultured
D003062:Codon; D006133:Growth Substances; D016298:Hematopoietic Cell Growth Factors; D007377:Interleukin-3; D016906:Interleukin-9; D008934:Mitogens; D016376:Oligonucleotides, Antisense; D011518:Proto-Oncogene Proteins; D011994:Recombinant Proteins; D019089:Stem Cell Factor; D016178:Granulocyte-Macrophage Colony-Stimulating Factor; D017346:Protein Serine-Threonine Kinases; D019908:Proto-Oncogene Proteins c-raf