C-Jun N-terminal kinases/c-Jun and p38 pathways cooperate in ceramide-induced neuronal apoptosis

Neuroscience. 2003;119(2):387-97. doi: 10.1016/s0306-4522(02)00996-x.

Abstract

Understanding the regulation of the apoptotic program in neurons by intracellular pathways is currently a subject of great interest. Recent results suggest that c-Jun N-terminal kinases (JNK), mitogen-activated protein kinases and the transcription factor c-Jun are important regulators of this cell death program in post-mitotic neurons following survival-factor withdrawal. Our study demonstrates that ceramide levels increase upon survival-factor withdrawal in primary cultured cortical neurons. Furthermore, survival-factor withdrawal or addition of exogenous c(2)-ceramide induces JNK pathway activation in these cells. Western blot analyses of JNK and c-Jun using phospho-specific antibodies reveal that JNK and subsequent c-Jun phosphorylation occur hours before the initiation of apoptosis, reflected morphologically by neurite retraction and fragmentation, cell-body shrinkage and chromatin fragmentation. Immunocytochemistry using the same antibodies shows that phospho-JNK are localized in the neurites of control neurons and translocate to the nucleus where phospho-c-Jun concurrently appears upon ceramide-induced apoptosis. To determine if ceramide-induced c-Jun activation is responsible for the induction of the apoptotic program, we performed transient transfections of a dominant negative form of c-Jun, truncated in its transactivation region. Our results show that DNc-Jun partially protects cortical neurons from ceramide-induced apoptosis. Treatment of dominant negative c-Jun-expressing neurons with the pharmacological inhibitor of p38 kinase, SB203580, completely blocked neuronal death. Thus our data show that p38 and JNK/c-Jun pathways cooperate to induce neuronal apoptosis.

Publication types

  • Comparative Study
  • Research Support, Non-U.S. Gov't

MeSH terms

  • Animals
  • Apoptosis*
  • Blotting, Western
  • Cell Survival / drug effects
  • Ceramides / pharmacology*
  • Cerebral Cortex / cytology
  • Cerebral Cortex / drug effects
  • Cerebral Cortex / metabolism
  • Culture Media, Serum-Free / pharmacology
  • Culture Techniques
  • DNA-Binding Proteins / metabolism
  • Early Growth Response Protein 1
  • Embryo, Mammalian
  • Enzyme Inhibitors / pharmacology
  • Female
  • Gene Expression Regulation / drug effects
  • Green Fluorescent Proteins
  • Imidazoles / pharmacology
  • Immediate-Early Proteins*
  • Immunohistochemistry
  • JNK Mitogen-Activated Protein Kinases*
  • Luminescent Proteins / metabolism
  • MAP Kinase Kinase 4
  • Mice
  • Mitogen-Activated Protein Kinase Kinases / metabolism
  • Mitogen-Activated Protein Kinases / metabolism*
  • Neurons / drug effects*
  • Neurons / metabolism
  • Neurons / pathology
  • Phosphorylation / drug effects
  • Pregnancy
  • Proteins / metabolism
  • Proto-Oncogene Proteins c-jun / metabolism*
  • Pyridines / pharmacology
  • Sphingosine / analogs & derivatives*
  • Sphingosine / pharmacology
  • Subcellular Fractions / drug effects
  • Subcellular Fractions / metabolism
  • Time Factors
  • Transcription Factors / metabolism
  • Transfection / methods
  • p38 Mitogen-Activated Protein Kinases

Substances

  • Ceramides
  • Culture Media, Serum-Free
  • DNA-Binding Proteins
  • Early Growth Response Protein 1
  • Egr1 protein, mouse
  • Enzyme Inhibitors
  • Imidazoles
  • Immediate-Early Proteins
  • Luminescent Proteins
  • N-acetylsphingosine
  • Proteins
  • Proto-Oncogene Proteins c-jun
  • Pyridines
  • Transcription Factors
  • Green Fluorescent Proteins
  • JNK Mitogen-Activated Protein Kinases
  • Mitogen-Activated Protein Kinases
  • p38 Mitogen-Activated Protein Kinases
  • MAP Kinase Kinase 4
  • Mitogen-Activated Protein Kinase Kinases
  • Sphingosine
  • SB 203580