MEGAKARYOCYTIC DIFFERENTIATION-INDUCED BY CONSTITUTIVE ACTIVATION OF MITOGEN-ACTIVATED PROTEIN-KINASE KINASE

The K562 erythroleukemia cell line was used to study the molecular mec hanisms regulating lineage commitment of hematopoietic stem cells. Pho rbol esters, which initiate megakaryocyte differentiation in this cell ii ne, caused a rapid increase in extracellular-signal-regulated kina se (ERK), which remained elevated for 2 h and returned to near-basal l evels by 24 h. In the absence of extracellular stimuli, ERK could be a ctivated by expression of constitutively active mutants of mitogen-act ivated protein (MAP) kinase kinase (MKK), resulting in cell adhesion a nd spreading, increased cell size, inhibition of cell growth, and indu ction of the platelet-specific integrin alpha(IIb)beta(3), all hallmar ks of megakaryocytic differentiation. In contrast, expression of wild- type MKK had little effect. In addition, constitutively active MKK sup pressed the expression of an erythroid marker, alpha-globin, indicatin g the ability to suppress cellular responses necessary for alternative cell lineages. The MKK inhibitor PD98059 blocked MKK/ERK activation a nd cellular responses to phorbol ester, demonstrating that activation of MKK is necessary and sufficient to induce a differentiation program along the megakaryocyte lineage. Thus, the MAP kinase cascade, which promotes cell growth and proliferation in many cell types, instead inh ibits cell proliferation and initiates lineage-specific differentiatio n in K562 cells, establishing a model system to investigate the mechan isms by which this signal transduction pathway specifies cell fate and developmental processes.