In vitro activation and nuclear translocation of NF-kappa B catalyzed by cyclic AMP-dependent protein kinase and protein kinase C.
We have examined whether a precursor form of NF-kappa B, a DNA-binding protein that plays a role in the transcriptional control of several genes, including kappa immunoglobulin light chain and interleukin-2 receptor alpha subunit, could be activated in vitro by protein kinases. DNA-binding activity of NF-kappa B was induced in the cytosolic fraction of unstimulated 70Z/3 murine pre-B cells by incubation with the catalytic subunit of cyclic AMP-dependent protein kinase (PKA) or protein kinase C (PKC). In contrast, PKA and PKC did not activate NF-kappa B in nuclear extracts from unstimulated cells. Identical results were obtained with the human natural killer-like cell line YT, which can be induced to express the interleukin-2 receptor alpha subunit in response to interleukin-1, cyclic AMP, or phorbol 12-myristate 13-acetate. Furthermore, when nuclei from unstimulated cells were incubated with PKA- or PKC-treated cytosolic fraction for 30 min at 30 degrees C, NF-kappa B was translocated into the nuclei. This translocation did not occur at 4 degrees C and was inhibited by wheat germ agglutinin but not by concanavalin A. Our findings support the conclusion that NF-kappa B exists in the cytoplasm of unstimulated cells in an inactive form that can be converted by exposure to PKA or PKC to an active DNA-binding form that can translocate to the nucleus.
Shirakawa
F
F
Department of Microbiology and Immunology, Wake Forest University Medical Center, Winston-Salem, North Carolina 27103.
Mizel
S B
SB
D000595:Amino Acid Sequence; D000818:Animals; D001692:Biological Transport; D002384:Catalysis; D002467:Cell Nucleus / Q000378:metabolism; D002478:Cells, Cultured; D000242:Cyclic AMP / Q000096:biosynthesis / Q000235:genetics; D003600:Cytosol / Q000378:metabolism; D004268:DNA-Binding Proteins / Q000378:metabolism*; D006801:Humans; D007375:Interleukin-1 / Q000502:physiology; D007546:Isoquinolines / Q000494:pharmacology; D008969:Molecular Sequence Data; D016328:NF-kappa B; D010703:Phorbol Esters / Q000494:pharmacology; D010766:Phosphorylation; D011493:Protein Kinase C / Q000037:antagonists & inhibitors / Q000378:metabolism*; D047428:Protein Kinase Inhibitors; D011494:Protein Kinases / Q000378:metabolism*; D011498:Protein Precursors / Q000378:metabolism*; D014157:Transcription Factors / Q000378:metabolism*; D014162:Transfection
D004268:DNA-Binding Proteins; D007375:Interleukin-1; D007546:Isoquinolines; D016328:NF-kappa B; D010703:Phorbol Esters; D047428:Protein Kinase Inhibitors; D011498:Protein Precursors; D014157:Transcription Factors; C053177:N-(2-(methylamino)ethyl)-5-isoquinolinesulfonamide; D000242:Cyclic AMP; D011494:Protein Kinases; D011493:Protein Kinase C