Death-associated Protein (DAP) Kinase Plays a Central Role in Ceramide-induced Apoptosis in Cultured Hippocampal Neurons

Dori Pelled,Tal Raveh,Christian Riebeling,Mati Fridkin,Hanna Berissi,Anthony H Futerman,Adi Kimchi

doi:10.1074/jbc.m104677200

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https://doi.org/10.1074/jbc.m104677200

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Abstract

Treatment of cultured hippocampal neurons with high concentrations of short-chain acyl ceramide derivatives, such as N-hexanoyl-D-sphingosine (C(6)-Cer), results in apoptotic cell death. We now show that death-associated protein (DAP) kinase plays an important role in mediating this effect. Upon incubation with C(6)-Cer, DAP kinase levels are elevated as early as 1 h after treatment, reaching levels 2-3-fold higher than untreated cells after 4 h. Neurons cultured from DAP kinase-deficient mice were significantly less sensitive to apoptosis induced by C(6)-Cer or by ceramide generated by high concentrations of nerve growth factor. A peptide corresponding to the 17 amino acids at the C terminus of DAP kinase protected wild type neurons from C(6)-Cer-induced death and from death induced by the addition of exogenous bacterial neutral sphingomyelinase, whereas a scrambled peptide had no protective effect, implying that the DAP kinase C-terminal tail inhibits the function of DAP kinase. Together, these data demonstrate that DAP kinase plays a central role in ceramide-induced cell death in neurons, but the pathway in which DAP kinase is involved is not the only one via which ceramide can induce apoptosis.

Highlights

Death-associated protein (DAP)1 kinase is a Ca2ϩ/calmodulin-regulated serine/threonine kinase that acts as a positive mediator of apoptosis induced by interferon-␥, Fas, tumor necrosis factor ␣, detachment from the extracellular matrix, or by deregulated oncogenes such as c-Myc or E2F-1 (1– 4)
High concentrations of ceramide, ceramide produced by the activity of neutral sphingomyelinase, or ceramide produced by the binding of nerve growth factor (NGF) to the p75 neurotrophin receptor,2 induces apoptotic cell death at different stages of neuronal development (14 –16)
deathassociated protein (DAP) Kinase Is Up-regulated by C6-Cer—We previously demonstrated that high concentrations of C6-Cer induce cell death in cultured hippocampal neurons via an apoptotic mechanism (14), inasmuch as nuclear condensation and annexin binding were observed, followed by retraction of axons and dendrites and detachment of neurons from the coverslips

Summary

Introduction

Death-associated protein (DAP)1 kinase is a Ca2ϩ/calmodulin-regulated serine/threonine kinase that acts as a positive mediator of apoptosis induced by interferon-␥, Fas, tumor necrosis factor ␣, detachment from the extracellular matrix, or by deregulated oncogenes such as c-Myc or E2F-1 (1– 4). Neurons cultured from DAP kinase-deficient mice were significantly less sensitive to apoptosis induced by C6-Cer or by ceramide generated by high concentrations of nerve growth factor.

Results

Conclusion