Sustained activation of the extracellular signal-regulated kinase/mitogen-activated protein kinase pathway is required for megakaryocytic differentiation of K562 cells.
The extracellular signal-regulated kinase (ERK), originally identified as a participant in mitogenic signaling, has recently been implicated in the signaling of cellular differentiation. To examine the role of the ERK/MAP kinase pathway in megakaryocytic differentiation of K562 cells, the effects of 12-O-tetradecanoylphorbol-13-acetate (TPA) and bryostatin on ERK activation were determined. Both TPA and bryostatin are known to activate PKC but paradoxically have opposing effects on megakaryocytic differentiation. TPA, a differentiation inducer, caused sustained activation of ERK (>24 h), whereas bryostatin, a differentiation blocker, only transiently activated ERK ( approximately 6 h) and attenuated the activation of ERK by TPA. To confirm a requirement for sustained ERK activation for megakaryocytic differentiation, PD098059, a synthetic inhibitor of the MAP kinase kinase 1 (MEK1) was employed. Introduction of PD098059 at any time during the first 18 h of TPA treatment completely abrogated megakaryocytic differentiation of K562 cells. After 24 h of TPA treatment, introduction of PD098059 failed to block differentiation. Differentiation blockade by PD098059 occurred via inhibition of MEK because transfection of a constitutively active mutant of MEK2 could override the PD098059 blockade. Experiments with conditioned media suggested that sustained activation of the ERK/MAP kinase pathway promoted the autocrine secretion of megakaryocytic lineage determination factors.
Racke
F K
FK
Institute of Pathology, Case Western Reserve University School of Medicine, Cleveland, Ohio 44106-4943, USA.
Lewandowska
K
K
Goueli
S
S
Goldfarb
A N
AN
D017871:Calcium-Calmodulin-Dependent Protein Kinases / Q000037:antagonists & inhibitors / Q000378:metabolism*; D002454:Cell Differentiation; D002478:Cells, Cultured; D017077:Culture Media, Conditioned; D004789:Enzyme Activation; D004791:Enzyme Inhibitors / Q000494:pharmacology; D005419:Flavonoids / Q000494:pharmacology; D048369:MAP Kinase Kinase 1; D048370:MAP Kinase Kinase 2; D008533:Megakaryocytes / Q000187:drug effects / Q000502:physiology*; D020929:Mitogen-Activated Protein Kinase Kinases; D020928:Mitogen-Activated Protein Kinases / Q000378:metabolism*; D009419:Nerve Tissue Proteins / Q000378:metabolism*; D019039:Platelet Glycoprotein GPIIb-IIIa Complex; D017346:Protein Serine-Threonine Kinases / Q000235:genetics / Q000378:metabolism*; D011505:Protein-Tyrosine Kinases / Q000235:genetics / Q000378:metabolism*; D015398:Signal Transduction; D013755:Tetradecanoylphorbol Acetate
D017077:Culture Media, Conditioned; D004791:Enzyme Inhibitors; D005419:Flavonoids; D009419:Nerve Tissue Proteins; D019039:Platelet Glycoprotein GPIIb-IIIa Complex; D011505:Protein-Tyrosine Kinases; D017346:Protein Serine-Threonine Kinases; D017871:Calcium-Calmodulin-Dependent Protein Kinases; D020928:Mitogen-Activated Protein Kinases; D048369:MAP Kinase Kinase 1; D048370:MAP Kinase Kinase 2; D020929:Mitogen-Activated Protein Kinase Kinases; D013755:Tetradecanoylphorbol Acetate; C093973:2-(2-amino-3-methoxyphenyl)-4H-1-benzopyran-4-one