Prolonged activation of the mitogen-activated protein kinase pathway is required for macrophage-like differentiation of a human myeloid leukemic cellline

The role of the mitogen-activated protein kinase (MAPK) signal transduction pathway in the proliferation of mammalian cells has been well established. However, there are relatively few reports concerning cell differentiation being mediated by MAPK. The effect of phorbol 12-myristate 13-acetate (PMA) on cell differentiation and signal transduction in a human myeloid leukemi a cell line, TF-1a, was investigated. When TF-1a cells were treated with 10 (-6), 10(-7), 10(-8) and 10(-9) M PMA for 24 h, they underwent 98, 93, 91, and 51% macrophage-like differentiation, respectively. PMA treatment rapidl y (10 min) induced phosphorylation of MAPK kinase (MEK and p44/42 MAPK), wh ich persisted for at least 24 h, p44/42 MAPK immunoprecipitates from lysate s of PMA-treated cells had increased ability to phosphorylate the transcrip tion factor Elk-1. This is important because phosphorylated Elk-1 can be co nsidered an "end-product" of the MAPK pathway. In contrast, treatment of TF -1a cells with granulocyte/macrophage-colony stimulating factor induced onl y transient activation of MEK and p44/42 MAPK (10-20 min) and an increase ( similar to 50%) in cell proliferation, without any change in cellular diffe rentiation. These results suggest that macrophage-like differentiation may be dependent on prolonged activation of the MAPK pathway. Additional suppor t for this conclusion was obtained from experiments showing that treatment of TF-1a cells with antisense oligonucleotides for MEK1 coding sequences pr ior to adding PMA inhibited macrophage-like differentiation. Furthermore, t ransient transfection with an inactive, dominant-negative MEK mutant also i nhibited PMA-induced differentiation, whereas transient transfection with a plasmid coding for constitutively activated MEK led to macrophage-like dif ferentiation in the absence of PMA.