A correlation between phorbol diester-induced protein phosphorylation and superoxide anion generation in HL-60 cells during granulocytic maturation.
As HL-60 cells matured along the granulocytic pathway, phorbol diester-induced superoxide anion production was compared to phorbol diester-induced protein phosphorylation using an in vitro phosphorylation technique. Maturation was induced by 0, 2, 4, or 6 days incubation with dimethyl sulfoxide (Me2SO). In 0 day Me2SO HL-60 cells, phorbol 12-myristate 13-acetate induced phosphorylation of protein pp29 (Mr = 28,600) and to a lesser extent protein pp76 (Mr = 76,300). With increased time of Me2SO incubation, phorbol 12-myristate 13-acetate induced phosphorylation of pp212 (Mr = 211,800), pp134 (Mr = 134,200), and pp76, whereas the phosphorylation of pp29 did not change appreciably. In close agreement with this increase in protein phosphorylation was the observed increase in phorbol diester-induced superoxide anion formation. Morphological characterization of cells during Me2SO-induced differentiation reveals that these increases in phorbol diester responses are probably attributable to the proportional rise in metamyelocytes, band, and segmented neutrophils. A variety of phorbol diesters increased superoxide anion generation in HL-60 cells differentiated into granulocyte-like cells by 6-day incubation with Me2SO. The structure-activity relationship of these phorbol diester derivatives for protein phosphorylation was strongly correlated to their ability to increase superoxide anion generation. Thus, we propose that phorbol diester-induced phosphorylation of pp212, pp134, and pp76, but not pp29 may play a role in mediating the functional response of phorbol diester-induced superoxide anion generation in HL-60 cells differentiated into mature granulocyte-like cells.
D002454:Cell Differentiation / Q000187:drug effects*; D002460:Cell Line; D004121:Dimethyl Sulfoxide / Q000494:pharmacology*; D006098:Granulocytes / Q000166:cytology / Q000187:drug effects; D006801:Humans; D007700:Kinetics; D015470:Leukemia, Myeloid, Acute; D008970:Molecular Weight; D009363:Neoplasm Proteins / Q000378:metabolism*; D010750:Phosphoproteins / Q000302:isolation & purification; D010766:Phosphorylation; D011493:Protein Kinase C / Q000378:metabolism*; D013481:Superoxides / Q000378:metabolism*; D013755:Tetradecanoylphorbol Acetate / Q000494:pharmacology*
D009363:Neoplasm Proteins; D010750:Phosphoproteins; D013481:Superoxides; D011493:Protein Kinase C; D013755:Tetradecanoylphorbol Acetate; D004121:Dimethyl Sulfoxide