Differential expression of cytosolic activation factors for NADPH oxidase in HL-60 leukemic cells.
Activation of NADPH oxidase in undifferentiated HL-60 leukemic cells and in HL-60 cells differentiated along the myeloid pathway with dibutyryl cyclic AMP (dbcAMP) or dimethyl sulfoxide (Me2SO) was studied. Upon stimulation with a calcium ionophore, a phorbol ester, arachidonic acid or gamma-hexachlorocyclohexane, Me2SO-differentiated HL-60 cells generated superoxide (O2-) at higher rates than dbcAMP-differentiated cells. Undifferentiated cells generated O2- only at low rates upon stimulation with the above agents. In cell-free systems, NADPH oxidase activity was reconstituted by combining membranes of undifferentiated or dbcAMP- or Me2SO-differentiated HL-60 cells, cytosol of Me2SO-differentiated cells and arachidonic acid. This basal O2- formation was enhanced several-fold by guanosine 5'-O-(3-thiotriphosphate) (GTP[gamma S]), a potent activator of guanine nucleotide-binding proteins. In contrast, cytosol of dbcAMP-differentiated cells reconstituted O2- formation only in the presence of GTP[gamma S], and cytosol of undifferentiated cells was inactive. Submaximally stimulatory amounts of cytosolic protein of Me2SO- and dbcAMP-differentiated cells synergistically stimulated O2- formation in the presence but not in the absence of GTP[gamma S]. We conclude that differentiations of HL-60 cells with Me2SO and dbcAMP are not equivalent with respect to activation of NADPH oxidase and that two cytosolic activation factors are involved in the regulation of this effector system.
Seifert
R
R
Institut für Pharmakologie, Freie Universität Berlin, FRG.
Jungblut
P
P
Schultz
G
G
D016718:Arachidonic Acid; D001095:Arachidonic Acids / Q000494:pharmacology; D003994:Bucladesine / Q000494:pharmacology; D000001:Calcimycin / Q000494:pharmacology; D002454:Cell Differentiation / Q000187:drug effects; D002460:Cell Line; D003600:Cytosol / Q000502:physiology; D004121:Dimethyl Sulfoxide / Q000494:pharmacology; D004789:Enzyme Activation; D016244:Guanosine 5'-O-(3-Thiotriphosphate); D006160:Guanosine Triphosphate / Q000031:analogs & derivatives / Q000494:pharmacology; D001556:Hexachlorocyclohexane / Q000494:pharmacology; D006801:Humans; D007700:Kinetics; D015473:Leukemia, Promyelocytic, Acute; D009240:N-Formylmethionine Leucyl-Phenylalanine / Q000494:pharmacology; D009247:NADH, NADPH Oxidoreductases / Q000378:metabolism*; D019255:NADPH Oxidases; D013481:Superoxides / Q000378:metabolism; D013755:Tetradecanoylphorbol Acetate / Q000494:pharmacology; D013873:Thionucleotides / Q000494:pharmacology
D001095:Arachidonic Acids; D013873:Thionucleotides; D013481:Superoxides; D016718:Arachidonic Acid; D016244:Guanosine 5'-O-(3-Thiotriphosphate); D000001:Calcimycin; D009240:N-Formylmethionine Leucyl-Phenylalanine; D001556:Hexachlorocyclohexane; D003994:Bucladesine; D006160:Guanosine Triphosphate; D009247:NADH, NADPH Oxidoreductases; D019255:NADPH Oxidases; D013755:Tetradecanoylphorbol Acetate; D004121:Dimethyl Sulfoxide