Binding of G-CSF, GM-CSF, tumor necrosis factor-alpha, and gamma-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), granulocyte-macrophage colony-stimulating factor (GM-CSF), gamma-interferon (gamma-IFN), or tumor necrosis factor-alpha (TNF-alpha) triggered the rapid, stable phosphorylation of a 75-Kd protein (p75) when incubated with permeabilized HL60 human myeloid leukemia cells in the presence of [gamma-32P] ATP. Among several chemical inducers of HL60 cell differentiation, dimethyl sulfoxide also triggered p75 labeling, but retinoic acid or 12-O-tetradecanoylphorbol-13-acetate did not elicit this response. Pretreatment of cells with G-CSF or GM-CSF 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. Immunoblot analysis of lysates of intact HL60 cells that had been incubated with G-CSF, GM-CSF, IFN, or TNF confirmed that tyrosine phosphorylation of a p75 also occurred in response to these cytokines in intact cells. Pretreatment of intact HL60 cells with one biologic agent or dimethyl sulfoxide abolished p75 labeling in response to incubation of permeabilized cells with a second agent, strongly suggesting that the same protein was phosphorylated in response to these treatments. p75 labeling was strictly dependent on expression of the appropriate ligand receptor. Data suggest that activation of a tyrosine kinase system is an early response to the binding of G-CSF, GM-CSF, TNF, or IFN to their respective cell surface receptors, or to the addition of dimethyl sulfoxide, and that the resulting phosphorylation event(s) may play a role in securing common elements in the biologic responses to these agents.
Evans
J P
JP
Department of Haematology, Royal Free Hospital School of Medicine, London, UK.
Mire-Sluis
A R
AR
Hoffbrand
A V
AV
Wickremasinghe
R G
RG
D003115:Colony-Stimulating Factors / Q000378:metabolism*; D016179:Granulocyte Colony-Stimulating Factor; D016178:Granulocyte-Macrophage Colony-Stimulating Factor; D006133:Growth Substances / Q000378:metabolism*; D006801:Humans; D007371:Interferon-gamma / Q000378:metabolism*; D007376:Interleukin-2 / Q000494:pharmacology; D008970:Molecular Weight; D010750:Phosphoproteins / Q000378:metabolism*; D010768:Phosphoserine / Q000378:metabolism; D011494:Protein Kinases / Q000502:physiology*; D011505:Protein-Tyrosine Kinases / Q000502:physiology*; D011956:Receptors, Cell Surface / Q000502:physiology*; D016184:Receptors, Colony-Stimulating Factor; D016188:Receptors, Granulocyte Colony-Stimulating Factor; D011971:Receptors, Immunologic / Q000502:physiology; D017471:Receptors, Interferon; D018124:Receptors, Tumor Necrosis Factor; D013997:Time Factors; D014407:Tumor Cells, Cultured; D014409:Tumor Necrosis Factor-alpha / Q000378:metabolism*
D003115:Colony-Stimulating Factors; D006133:Growth Substances; D007376:Interleukin-2; D010750:Phosphoproteins; D011956:Receptors, Cell Surface; D016184:Receptors, Colony-Stimulating Factor; D016188:Receptors, Granulocyte Colony-Stimulating Factor; D011971:Receptors, Immunologic; D017471:Receptors, Interferon; D018124:Receptors, Tumor Necrosis Factor; D014409:Tumor Necrosis Factor-alpha; D016179:Granulocyte Colony-Stimulating Factor; D010768:Phosphoserine; D007371:Interferon-gamma; D016178:Granulocyte-Macrophage Colony-Stimulating Factor; D011494:Protein Kinases; D011505:Protein-Tyrosine Kinases