Caspase activation during phorbol ester-induced apoptosis requires ROCK-dependent myosin-mediated contraction.
Treatment of cells with phorbol ester, phorbol-12-myristate-13-acetate (PMA), triggers differentiation or apoptosis, depending on the cell type. In this study, we used an erythroblastic cell line, TF-1, to investigate the molecular mechanism that determines the cell fate in response to PMA exposure. Upon PMA treatment in the presence of serum or lysophosphatidic acid (LPA), TF-1 cells exhibited contraction followed by apoptosis. By contrast, under serum-free conditions, cells became adherent and survived after PMA treatment. Here, we show that the pathway of Rho kinase (ROCK)/myosin light chain (MLC) phosphorylation/myosin-mediated contraction was activated in PMA-induced apoptotic cells in serum-containing medium, but not in the adherent and survived cells. Pretreatment of cells with a specific ROCK inhibitor, Y27632, not only abrogated MLC phosphorylation and membrane contraction, but also prevented PMA-induced activation of caspase-3 and subsequent cell death, indicating that ROCK-dependent myosin-mediated contraction elicits an upstream signal required for caspase-3 activation in PMA-induced apoptosis. Interestingly, we further found that caspases-8 and -10 are the initiator caspases in PMA-induced apoptosis and a ROCK-dependent enhancement of specific complex formation between the Fas-associated death domain (FADD) and pro-caspase-10 in pro-apoptotic cells. In summary, these results revealed that, following PMA treatment, the upregulation of the RhoA/ROCK pathway contributes to a cellular context that switches-on myosin-mediated contraction, which provides a mechanism for triggering apoptotic induction mediated by caspase-8 and -10.
Lai
Jin-Mei
JM
Graduate Institute of Biochemistry and Molecular Biology, College of Medicine, National Taiwan University, No. 1 section 1 Jen-Ai Road, Taipei 100, Taiwan, Republic of China.
Hsieh
Chia-Ling
CL
Chang
Zee-Fen
ZF
D000577:Amides / Q000494:pharmacology; D017209:Apoptosis / Q000187:drug effects / Q000502:physiology*; D002273:Carcinogens / Q000494:pharmacology; D053455:Caspase 10; D053148:Caspase 3; D053181:Caspase 8; D020169:Caspases / Q000378:metabolism*; D002448:Cell Adhesion / Q000502:physiology; D003001:Cloning, Molecular; D053418:Death Domain Receptor Signaling Adaptor Proteins; D004789:Enzyme Activation; D004791:Enzyme Inhibitors / Q000494:pharmacology; D004900:Erythroblasts / Q000378:metabolism; D006801:Humans; D008246:Lysophospholipids / Q000494:pharmacology; D008856:Microscopy, Fluorescence; D018994:Myosin Light Chains / Q000378:metabolism; D009218:Myosins / Q000378:metabolism; D010703:Phorbol Esters / Q000494:pharmacology*; D010766:Phosphorylation; D017434:Protein Structure, Tertiary / Q000502:physiology; D011725:Pyridines / Q000494:pharmacology; D018124:Receptors, Tumor Necrosis Factor / Q000378:metabolism; D020741:rho GTP-Binding Proteins / Q000037:antagonists & inhibitors / Q000378:metabolism*
D000577:Amides; D002273:Carcinogens; D053418:Death Domain Receptor Signaling Adaptor Proteins; D004791:Enzyme Inhibitors; D008246:Lysophospholipids; D018994:Myosin Light Chains; D010703:Phorbol Esters; D011725:Pyridines; D018124:Receptors, Tumor Necrosis Factor; C108830:Y 27632; C505407:CASP3 protein, human; C505530:CASP8 protein, human; D053455:Caspase 10; D053148:Caspase 3; D053181:Caspase 8; D020169:Caspases; C506438:CASP10 protein, human; D009218:Myosins; D020741:rho GTP-Binding Proteins