Abstract
Protein kinase-B (PKB) and its target, the forkhead transcription factor like 1 (FKHRL1)/FoxO3a, have been suggested as regulators of neurotrophin-mediated cell survival in neuronal cells. We analyzed human neuroblastoma cells and found that FKHRL1 was phosphorylated, suggesting its inactivation. To study FKHRL1 function, we infected SH-EP and NB15 cells with a 4OH-tamoxifen-regulated FKHRL1(A3)ER(tm) transgene. Activation of FKHRL1 promoted cytochrome-c release and caspase-dependent apoptosis. FKHRL1 induced TRAIL and the BH3-only proteins Noxa and Bim, implicating both extrinsic and intrinsic death pathways. However, expression of dnFADD did not inhibit FKHRL1-induced cell death, whereas Bcl2 protected against apoptosis. This excluded the death-receptor pathway and suggested that cell death decision is regulated by Bcl2-rheostat. Importantly, RNAi knockdown of Noxa or Bim decreased apoptosis, indicating that Noxa and Bim cooperate to mediate FKHRL1-induced cell death. We conclude that Noxa and Bim establish a connection between FKHRL1 and mitochondria, and that both BH3-only proteins are critically involved in FKHRL1-induced apoptosis in neuroblastoma.
Publication types
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Research Support, Non-U.S. Gov't
MeSH terms
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3-Phosphoinositide-Dependent Protein Kinases
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Apoptosis
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Apoptosis Regulatory Proteins / genetics
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Apoptosis Regulatory Proteins / metabolism*
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Bcl-2-Like Protein 11
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Caspases / metabolism
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Cell Death
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Fas-Associated Death Domain Protein / metabolism
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Fas-Associated Death Domain Protein / physiology
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Forkhead Box Protein O3
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Forkhead Transcription Factors / genetics
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Forkhead Transcription Factors / metabolism*
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Gene Expression Regulation, Neoplastic*
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Humans
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Membrane Proteins / genetics
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Membrane Proteins / metabolism*
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Mitochondria / metabolism*
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Models, Biological
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Neuroblastoma / metabolism*
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Peptide Fragments / metabolism
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Phosphatidylinositol 3-Kinases / metabolism
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Protein Serine-Threonine Kinases / metabolism
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Proto-Oncogene Proteins / genetics
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Proto-Oncogene Proteins / metabolism*
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Proto-Oncogene Proteins c-bcl-2 / genetics
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Proto-Oncogene Proteins c-bcl-2 / metabolism*
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Receptors, TNF-Related Apoptosis-Inducing Ligand / metabolism
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Reverse Transcriptase Polymerase Chain Reaction
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Signal Transduction
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TNF-Related Apoptosis-Inducing Ligand / metabolism
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Tamoxifen / analysis
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Tamoxifen / pharmacology
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Transduction, Genetic
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fas Receptor / metabolism
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fas Receptor / physiology
Substances
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Apoptosis Regulatory Proteins
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BCL2L11 protein, human
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Bax protein (53-86)
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Bcl-2-Like Protein 11
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FADD protein, human
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FAS protein, human
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FOXO3 protein, human
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Fas-Associated Death Domain Protein
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Forkhead Box Protein O3
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Forkhead Transcription Factors
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Membrane Proteins
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PMAIP1 protein, human
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Peptide Fragments
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Proto-Oncogene Proteins
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Proto-Oncogene Proteins c-bcl-2
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Receptors, TNF-Related Apoptosis-Inducing Ligand
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TNF-Related Apoptosis-Inducing Ligand
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fas Receptor
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Tamoxifen
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3-Phosphoinositide-Dependent Protein Kinases
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Protein Serine-Threonine Kinases
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Caspases