Abstract
Post-synaptic density-95 (PSD-95) is a neuronal scaffolding protein that associates with N -methyl-D-aspartate (NMDA) receptors and links them to intracellular signalling molecules. In neurons, neuronal nitric oxide synthase (nNOS) binds selectively to the second PDZ domain (PDZ2) of PSD-95, thereby exhibiting physiological activation triggered via NMDA receptors. We have demonstrated previously that Ca(2+)/calmodulin-dependent protein kinase IIalpha (CaM-K IIalpha) directly phosphorylates nNOS at residue Ser(847), and can attenuate the catalytic activity of the enzyme in neuronal cells [Komeima, Hayashi, Naito and Watanabe (2000) J. Biol. Chem. 275, 28139-28143]. In the present study, we examined how CaM-K II participates in the phosphorylation by analysing the functional interaction between nNOS and PSD-95 in cells. The results showed that PSD-95 directly promotes the nNOS phosphorylation at Ser(847) induced by endogenous CaM-K II. In transfected cells, this effect of PSD-95 required its dual palmitoylation and the PDZ2 domain, but did not rely on its guanylate kinase domain. CaM-K Ialpha and CaM-K IV failed to phosphorylate nNOS at Ser(847) in transfected cells. Thus PSD-95 mediates cellular trafficking of nNOS, and may be required for the efficient phosphorylation of nNOS at Ser(847) by CaM-K II in neuronal cells.
Publication types
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Comparative Study
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Research Support, Non-U.S. Gov't
MeSH terms
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Animals
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Brain / enzymology*
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Calcium
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Calcium-Calmodulin-Dependent Protein Kinase Kinase
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Calcium-Calmodulin-Dependent Protein Kinase Type 2
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Calcium-Calmodulin-Dependent Protein Kinases / metabolism*
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Cells, Cultured / cytology
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Cells, Cultured / metabolism
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Disks Large Homolog 4 Protein
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Immunoenzyme Techniques
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Intracellular Signaling Peptides and Proteins
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Membrane Proteins
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Nerve Tissue Proteins / pharmacology*
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Nitric Oxide Synthase / isolation & purification
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Nitric Oxide Synthase / metabolism*
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Nitric Oxide Synthase Type I
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Phosphorylation
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Plasmids
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Precipitin Tests
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Protein Kinases / metabolism
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Protein Serine-Threonine Kinases / metabolism
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Rats
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Receptors, N-Methyl-D-Aspartate / metabolism
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Serine / metabolism*
Substances
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Disks Large Homolog 4 Protein
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Dlg4 protein, rat
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Intracellular Signaling Peptides and Proteins
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Membrane Proteins
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Nerve Tissue Proteins
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Receptors, N-Methyl-D-Aspartate
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postsynaptic density proteins
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Serine
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Nitric Oxide Synthase
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Nitric Oxide Synthase Type I
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Nos1 protein, rat
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Protein Kinases
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(CaM)-dependent protein kinase Ia kinase
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Protein Serine-Threonine Kinases
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Calcium-Calmodulin-Dependent Protein Kinase Kinase
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Calcium-Calmodulin-Dependent Protein Kinase Type 2
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Calcium-Calmodulin-Dependent Protein Kinases
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Camk2a protein, rat
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Calcium