6-Hydroxydopamine activates the mitochondrial apoptosis pathway through p38 MAPK-mediated, p53-independent activation of Bax and PUMA.
Mitochondrial alterations have been associated with the cytotoxic effect of 6-hydroxydopamine (6-OHDA), a widely used toxin to study Parkinson's disease. In previous work, we have demonstrated that 6-OHDA increases mitochondrial membrane permeability leading to cytochrome c release, but the precise mechanisms involved in this process remain unknown. Herein we studied the mechanism of increased mitochondrial permeability of SH-SY5Y neuroblastoma cells in response to 6-OHDA. Cytochrome c release induced by 6-OHDA occurred, in both SH-SY5Y cells and primary cultures, in the absence of mitochondrial swelling or a decrease in mitochondrial calcein fluorescence, suggesting little involvement of the mitochondrial permeability transition pore in this process. In contrast, 6-OHDA-induced cell death was associated with a significant translocation of the pro-apoptotic Bax protein from the cytosol to mitochondria and with a significant induction of the BH3-only protein PUMA. Experiments in mouse embryonic fibroblasts deficient in Bax or PUMA demonstrated a role for both proteins in 6-OHDA-induced apoptosis. Although 6-OHDA elevated both total and nuclear p53 protein levels, activation of p53 was not essential for subsequent cell death. In contrast, we found that p38 mitogen-activated protein kinase (MAPK) was activated early during 6-OHDA-induced apoptosis, and that treatment with the p38 MAPK inhibitor SKF86002 potently inhibited PUMA induction, green fluorescent protein-Bax redistribution and apoptosis in response to 6-OHDA. These data demonstrate a critical involvement of p38 MAPK, PUMA, and Bax in 6-OHDA-induced apoptosis.
Gomez-Lazaro
Maria
M
Grupo de Neurofarmacología, Departamento de Ciencias Médicas, Facultad de Medicina, Universidad Castilla-La Mancha, Albacete, Spain.
Galindo
Maria F
MF
Concannon
Caoimhín G
CG
Segura
Miguel F
MF
Fernandez-Gomez
Francisco J
FJ
Llecha
Nuria
N
Comella
Joan X
JX
Prehn
Jochen H M
JH
Jordan
Joaquin
J
D000818:Animals; D017209:Apoptosis / Q000187:drug effects / Q000502:physiology*; D051017:Apoptosis Regulatory Proteins / Q000235:genetics / Q000378:metabolism*; D045744:Cell Line, Tumor; D004791:Enzyme Inhibitors / Q000494:pharmacology; D005347:Fibroblasts / Q000166:cytology / Q000187:drug effects / Q000201:enzymology; D049452:Green Fluorescent Proteins / Q000235:genetics; D006801:Humans; D007093:Imidazoles / Q000494:pharmacology; D051379:Mice; D018345:Mice, Knockout; D008928:Mitochondria / Q000187:drug effects / Q000378:metabolism*; D008933:Mitochondrial Swelling / Q000187:drug effects / Q000502:physiology; D009447:Neuroblastoma; D009474:Neurons / Q000166:cytology / Q000187:drug effects / Q000201:enzymology; D016627:Oxidopamine / Q000633:toxicity; D011518:Proto-Oncogene Proteins / Q000235:genetics / Q000378:metabolism*; D013565:Sympatholytics / Q000633:toxicity; D013844:Thiazoles / Q000494:pharmacology; D014162:Transfection; D016159:Tumor Suppressor Protein p53 / Q000235:genetics / Q000378:metabolism*; D051028:bcl-2-Associated X Protein / Q000235:genetics / Q000378:metabolism*; D048051:p38 Mitogen-Activated Protein Kinases / Q000037:antagonists & inhibitors / Q000378:metabolism*
D051017:Apoptosis Regulatory Proteins; C494826:BAX protein, human; C438378:BBC3 protein, human; D004791:Enzyme Inhibitors; D007093:Imidazoles; D011518:Proto-Oncogene Proteins; D013565:Sympatholytics; D013844:Thiazoles; D016159:Tumor Suppressor Protein p53; D051028:bcl-2-Associated X Protein; D049452:Green Fluorescent Proteins; D016627:Oxidopamine; C052026:6-(4-fluorophenyl)-2,3-dihydro-5-(4-pyridinyl)imidazo(2,1-b)thiazole; D048051:p38 Mitogen-Activated Protein Kinases