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In response to treatment with phorbol-12-myristate-13-acetate (PMA), the half-population of erythromyeloblast D2 cells, a cytokine-independent variant of TF-1 cells, displayed adhesion and differentiated into a monocyte/macrophage-like morphology, while the other half-population remained in suspension and underwent apoptosis. Expression of the cell cycle inhibitor p21(Cip1/Waf1) was induced after PMA treatment in the adherent cells but not in the proapoptotic cells. We investigated the mechanism responsible for the impairment of p21(Cip1/Waf1) induction in PMA-induced proapoptotic cells. We demonstrated that in PMA-induced adherent cells, upregulation of p21(Cip1/Waf1) requires the activation and nuclear translocation of phosphorylated extracellular signal-regulated kinase (phospho-ERK). Although ERK was phosphorylated to comparable levels in PMA-induced proapoptotic and adherent cells, nuclear distribution of phospho-ERK was seen only in the adherent, not in the proapoptotic cells. We also found that only PMA-induced proapoptotic cells contained the phosphorylated form of myosin light chain, which is dependent on Rho-associated kinase (ROCK) activation, and that expression of a dominant-active form of ROCK suppressed activation of the p21(Cip1/Waf1) promoter during PMA induction. Finally, we demonstrated that inhibition of ROCK restores nuclear distribution of phospho-ERK and activation of p21(Cip1/Waf1) expression. Based on these findings, we propose that a ROCK-mediated signal is involved in interfering with the process of ERK-mediated p21(Cip1/Waf1) induction in PMA-induced proapoptotic TF-1 and D2 cells.

molecular and cellular biology

Cytosolic Retention of Phosphorylated Extracellular Signal-Regulated Kinase and a Rho-Associated Kinase-Mediated Signal Impair Expression of p21<sup>Cip1/Waf1</sup> in Phorbol 12-Myristate-13- Acetate-Induced Apoptotic Cells

Cytosolic Retention of Phosphorylated Extracellular Signal-Regulated Kinase and a Rho-Associated Kinase-Mediated Signal Impair Expression of p21Cip1/Waf1 in Phorbol 12-Myristate-13- Acetate-Induced Apoptotic Cells

Cytosolic Retention of Phosphorylated Extracellular Signal-Regulated Kinase and a Rho-Associated Kinase-Mediated Signal Impair Expression of p21^Cip1/Waf1 in Phorbol 12-Myristate-13- Acetate-Induced Apoptotic Cells