ABSTRACT
Excitotoxic neuronal death contributes to many neurological disorders, and involves calcium influx and stress-activated protein kinases (SAPKs) such as p38alpha. There is indirect evidence that the small Rho-family GTPases Rac and cdc42 are involved in neuronal death subsequent to the withdrawal of nerve growth factor (NGF), whereas Rho is involved in the inhibition of neurite regeneration and the release of the amyloidogenic Abeta(42) peptide. Here we show that Rho is activated in rat neurons by conditions that elevate intracellular calcium and in the mouse cerebral cortex during ischemia. Rho is required for the rapid glutamate-induced activation of p38alpha and ensuing neuronal death. The ability of RhoA to activate p38alpha was not expected, and it was specific to primary neuronal cultures. The expression of active RhoA alone not only activated p38alpha but also induced neuronal death that was sensitive to the anti-apoptotic protein Bcl-2, showing that RhoA was sufficient to induce the excitotoxic pathway. Therefore, Rho is an essential component of the excitotoxic cell death pathway.
Subject(s)
Calcium/metabolism , Mitogen-Activated Protein Kinase 14/metabolism , Neurons/physiology , rhoA GTP-Binding Protein/physiology , ADP Ribose Transferases/metabolism , ADP Ribose Transferases/pharmacology , Animals , Animals, Newborn , Botulinum Toxins/metabolism , Botulinum Toxins/pharmacology , Brain/cytology , Cell Death/drug effects , Cell Death/physiology , Cells, Cultured , Dose-Response Relationship, Drug , Enzyme Activation/drug effects , Enzyme Activation/physiology , Enzyme Inhibitors/pharmacology , Excitatory Amino Acid Agonists/pharmacology , Functional Laterality , Glutamic Acid/pharmacology , Ischemia/metabolism , Ischemia/pathology , Mice , Models, Molecular , N-Methylaspartate/pharmacology , Neurons/drug effects , Transfection/methods , cdc42 GTP-Binding Protein/metabolismABSTRACT
Normal functioning of the nervous system requires precise regulation of dendritic shape and synaptic connectivity. Here, we report a severe impairment of dendritic structures in the cerebellum and motor cortex of c-Jun N-terminal kinase 1 (JNK1)-deficient mice. Using an unbiased screen for candidate mediators, we identify the dendrite-specific high-molecular-weight microtubule-associated protein 2 (MAP2) as a JNK substrate in the brain. We subsequently show that MAP2 is phosphorylated by JNK in intact cells and that MAP2 proline-rich domain phosphorylation is decreased in JNK1-/- brain. We developed compartment-targeted JNK inhibitors to define whether a functional relationship exists between the physiologically active, cytosolic pool of JNK and dendritic architecture. Using these, we demonstrate that cytosolic, but not nuclear, JNK determines dendritic length and arbor complexity in cultured neurons. Moreover, we confirm that MAP2-dependent process elongation is enhanced after activation of JNK. Using JNK1-/- neurons, we reveal a dominant role for JNK1 over ERK in regulating dendritic arborization, whereas ERK only regulates dendrite shape under conditions in which JNK activity is low (JNK1-/- neurons). These results reveal a novel antagonism between JNK and ERK, potentially providing a mechanism for fine-tuning the dendritic arbor. Together, these data suggest that JNK phosphorylation of MAP2 plays an important role in defining dendritic architecture in the brain.
Subject(s)
Cytosol/enzymology , Dendrites/ultrastructure , Microtubule-Associated Proteins/physiology , Mitogen-Activated Protein Kinase 8/physiology , Animals , COS Cells , Cell Differentiation , Cell Nucleus/enzymology , Cells, Cultured/enzymology , Cells, Cultured/ultrastructure , Cerebellar Cortex/cytology , Cerebellar Cortex/enzymology , Cerebellar Cortex/growth & development , Chlorocebus aethiops , Mice , Mice, Knockout , Mitogen-Activated Protein Kinase 1/metabolism , Mitogen-Activated Protein Kinase 3/metabolism , Mitogen-Activated Protein Kinase 8/antagonists & inhibitors , Mitogen-Activated Protein Kinase 8/deficiency , Mitogen-Activated Protein Kinase 8/genetics , Motor Cortex/cytology , Motor Cortex/enzymology , Neurons/ultrastructure , Phosphorylation , Prosencephalon/cytology , Prosencephalon/enzymology , Protein Processing, Post-Translational , Rats , Rats, Sprague-Dawley , Recombinant Fusion Proteins/physiology , TransfectionABSTRACT
High levels of cytokines are risk factors for type 2 diabetes. Therefore, we investigated whether the promoter polymorphisms of the tumor necrosis factor-alpha (TNF-alpha; G-308A) and interleukin 6 (IL-6; C-174G) genes predict the conversion from impaired glucose tolerance (IGT) to type 2 diabetes in the Finnish Diabetes Prevention Study. Altogether, 490 overweight subjects with IGT whose DNA was available were randomly divided into one of the two treatment assignments: the control group and the intensive, individualized diet and exercise intervention group. The -308A allele of the TNF-alpha gene was associated with an approximate twofold higher risk for type 2 diabetes compared with the G-308G genotype (odds ratio 1.80, 95% CI 1.05-3.09; P = 0.034). Subjects with both the A allele of the TNF-alpha gene and the C-174C genotype of the IL-6 gene had a 2.2-fold (CI 1.02-4.85, P = 0.045) higher risk of developing type 2 diabetes than subjects without the risk genotypes. We conclude that the -308A allele of the promoter polymorphism (G-308A) of the TNF-alpha gene is a predictor for the conversion from IGT to type 2 diabetes. Furthermore, this polymorphism seems to have a gene-gene interaction with the C-174C genotype of the IL-6 gene.
