Subject(s)
Induced Pluripotent Stem Cells/cytology , Motor Neurons/cytology , Animals , Cell Differentiation , Cell Shape , Cells, Cultured , Coculture Techniques , Humans , Induced Pluripotent Stem Cells/metabolism , Induced Pluripotent Stem Cells/ultrastructure , Ion Channels/metabolism , Keratinocytes/cytology , Keratinocytes/metabolism , Mice , Mice, Inbred C57BL , Motor Neurons/metabolism , Motor Neurons/ultrastructure , Muscle Fibers, Skeletal/cytology , Muscle Fibers, Skeletal/metabolism , Neuromuscular Junction/metabolism , Neuromuscular Junction/ultrastructure , Synapses/metabolism , Time FactorsABSTRACT
The dynactin p150glued subunit, encoded by the gene DCTN1 is part of the dynein-dynactin motor protein complex responsible for retrograde axonal transport. This subunit is a candidate modifier for neurodegenerative diseases, in particular motoneuron and extrapyramidal diseases. Based on an extensive screening effort of all 32 exons in more than 2,500 ALS/MND patients, patients suffering from Parkinsonian Syndromes and controls, we investigated 24 sequence variants of p150 in cell-based studies. We used both non-neuronal cell lines and primary rodent spinal motoneurons and report on cell biological abnormalities in five of these sequence alterations and also briefly report on the clinical features. Our results suggest the presence of biological changes caused by some p150 mutants pointing to a potential pathogenetic significance as modifier of the phenotype of the human disease.
Subject(s)
Amyotrophic Lateral Sclerosis/genetics , Microtubule-Associated Proteins/genetics , Motor Neurons/metabolism , Parkinsonian Disorders/genetics , Parkinsonian Disorders/pathology , Adaptor Proteins, Signal Transducing , Amyotrophic Lateral Sclerosis/pathology , Animals , Apoptosis/drug effects , Apoptosis/genetics , Autophagy-Related Proteins , Carrier Proteins/metabolism , Cell Cycle Proteins/metabolism , Cells, Cultured , Chlorocebus aethiops , Dynactin Complex , Embryo, Mammalian , Female , Green Fluorescent Proteins/genetics , Humans , Male , Microscopy, Electron, Transmission , Microtubule-Associated Proteins/metabolism , Microtubules/metabolism , Motor Neurons/pathology , Motor Neurons/ultrastructure , Mutation/genetics , Pregnancy , Proteasome Endopeptidase Complex/metabolism , Protein Binding , Rats , Rats, Sprague-Dawley , Retrospective Studies , Spinal Cord/cytology , Time FactorsABSTRACT
The dynactin p150(Glued) subunit, encoded by the gene DCTN1, is part of the dynein-dynactin motor protein complex responsible for retrograde axonal transport in motor neurons. The p150 subunit is a candidate gene for neurodegenerative diseases, in particular motor neuron and extrapyramidal diseases. Tubulin-binding cofactors are believed to be involved in tubulin biogenesis and degradation and therefore to contribute to microtubule functional diversity and regulation. A yeast-two-hybrid screen for putative interacting proteins of dynactin p150(Glued) has revealed tubulin-folding cofactor B (TBCB). We analyzed the interaction of these proteins and investigated the impact of this complex on the microtubule network in cell lines and primary hippocampal neurons in vitro. We especially concentrated on neuronal morphology and synaptogenesis. Overexpression of both proteins or depletion of TBCB alone does not alter the microtubule network and/or neuronal morphology. The demonstration of the interaction of the transport molecule dynactin and the tubulin-regulating factor TBCB is thought to have an impact on several cellular mechanisms. TBCB expression levels have been found to have only a subtle influence on the microtubule network and neuronal morphology. However, overexpression of TBCB leads to the decreased localization of p150 to the microtubule network that might result in a functional modulation of this protein complex.
