Abstract
The cytoplasmic adaptor protein Disabled-1 (Dab1) is necessary for the regulation of neuronal positioning in the developing brain by the secreted molecule Reelin. Binding of Reelin to the neuronal apolipoprotein E receptors apoER2 and very low density lipoprotein receptor induces tyrosine phosphorylation of Dab1 and the subsequent activation or relocalization of downstream targets like phosphatidylinositol 3 (PI3)-kinase and Nckbeta. Disruption of Reelin signaling leads to the accumulation of Dab1 protein in the brains of genetically modified mice, suggesting that Reelin limits its own action in responsive neurons by down-regulating the levels of Dab1 expression. Here, we use cultured primary embryonic neurons as a model to demonstrate that Reelin treatment targets Dab1 for proteolytic degradation by the ubiquitin-proteasome pathway. We show that tyrosine phosphorylation of Dab1 but not PI3-kinase activation is required for its proteasomal targeting. Genetic deficiency in the Dab1 kinase Fyn prevents Dab1 degradation. The Reelin-induced Dab1 degradation also depends on apoER2 and very low density lipoprotein receptor in a gene-dose dependent manner. Moreover, pharmacological blockade of the proteasome prevents the formation of a proper cortical plate in an in vitro slice culture assay. Our results demonstrate that signaling through neuronal apoE receptors can activate the ubiquitin-proteasome machinery, which might have implications for the role of Reelin during neurodevelopment and in the regulation of synaptic transmission.
Publication types
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Research Support, Non-U.S. Gov't
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Research Support, U.S. Gov't, P.H.S.
MeSH terms
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Amino Acid Sequence
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Animals
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Binding Sites
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Cell Adhesion Molecules, Neuronal / genetics
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Cell Adhesion Molecules, Neuronal / pharmacology*
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Cells, Cultured
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Cerebral Cortex / embryology
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Culture Techniques
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Cysteine Endopeptidases / metabolism*
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Embryo, Mammalian
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Enzyme Inhibitors / pharmacology
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Extracellular Matrix Proteins / genetics
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Extracellular Matrix Proteins / pharmacology*
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Humans
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Immunohistochemistry
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Low Density Lipoprotein Receptor-Related Protein-1 / genetics
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Low Density Lipoprotein Receptor-Related Protein-1 / physiology*
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Mice
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Mice, Inbred C57BL
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Molecular Sequence Data
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Multienzyme Complexes / antagonists & inhibitors
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Multienzyme Complexes / metabolism*
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Mutation
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Nerve Tissue Proteins / chemistry
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Nerve Tissue Proteins / genetics
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Nerve Tissue Proteins / metabolism*
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Neurons / chemistry
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Neurons / drug effects
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Neurons / metabolism*
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Phosphatidylinositol 3-Kinases / metabolism
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Phosphorylation
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Polymerase Chain Reaction
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Proteasome Endopeptidase Complex
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Proto-Oncogene Proteins / genetics
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Proto-Oncogene Proteins / physiology
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Proto-Oncogene Proteins c-fyn
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RNA, Messenger / analysis
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Rats
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Rats, Sprague-Dawley
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Receptors, LDL / genetics
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Receptors, LDL / physiology
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Recombinant Fusion Proteins
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Reelin Protein
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Serine Endopeptidases
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Signal Transduction
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Transfection
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Tyrosine / metabolism
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Ubiquitin / genetics
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Ubiquitin / metabolism
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src-Family Kinases / metabolism
Substances
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Cell Adhesion Molecules, Neuronal
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Dab1 protein, mouse
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Enzyme Inhibitors
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Extracellular Matrix Proteins
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Low Density Lipoprotein Receptor-Related Protein-1
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Multienzyme Complexes
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Nerve Tissue Proteins
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Proto-Oncogene Proteins
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RNA, Messenger
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Receptors, LDL
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Recombinant Fusion Proteins
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Reelin Protein
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Reln protein, rat
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Ubiquitin
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VLDL receptor
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Tyrosine
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FYN protein, human
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Fyn protein, mouse
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Fyn protein, rat
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Proto-Oncogene Proteins c-fyn
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src-Family Kinases
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RELN protein, human
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Reln protein, mouse
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Serine Endopeptidases
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Cysteine Endopeptidases
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Proteasome Endopeptidase Complex