Binding of G-CSF, GM-CSF, Tumor Necrosis Factor-a, and y-Interferon to Cell Surface Receptors on Human Myeloid Leukemia Cells Triggers Rapid Tyrosine and Serine Phosphorylation of a 75-Kd Protein

Granulocyte colony-stimulating factor (G-CSF), granulocytemacrophage colony-stimulating factor (GM-CSF), ?-interferon (?-IFN), or tumor necrosis factor-a (TNF-a) triggered the rapid, stable phosphorylation of a 75-Kd protein ( ~ 7 5 ) when incubated with permeabilized HL60 human myeloid leukemia cells in the presence of [?-32P] ATP. Among several chemical inducers of HL60 cell differentiation. dimethyl sulfoxide also triggered p75 labeling, but retinoic acid or 12-0-tetradecanoylphorbol-I 3-acetate did not elicit this response. Pretreatment of cells with G-CSF or GMCSF for more than 30 seconds before permeabilization rendered the p75 labeling undetectable, suggesting that ligand-stimulated labeling was rapidly completed within this time in intact cells. Phosphorylation of p75 occurred on serine and tyrosine residues. This conclusion was confirmed by direct phosphoamino acid analysis. lmmunoblot analysis of lysates of intact HL60 cells that had been