NIDD, a novel DHHC-containing protein, targets neuronal nitric-oxide synthase (nNOS) to the synaptic membrane through a PDZ-dependent interaction and regulates nNOS activity.
Targeting of neuronal nitric-oxide synthase (nNOS) to appropriate sites in a cell is mediated by interactions with its PDZ domain and plays an important role in specifying the sites of reaction of nitric oxide (NO) in the central nervous system. Here we report the identification and characterization of a novel nNOS-interacting DHHC domain-containing protein with dendritic mRNA (NIDD) (GenBank accession number AB098078), which increases nNOS enzyme activity by targeting the nNOS to the synaptic plasma membrane in a PDZ domain-dependent manner. The deduced NIDD protein consisted of 392 amino acid residues and possessed five transmembrane segments, a zinc finger DHHC domain, and a PDZ-binding motif (-EDIV) at its C-terminal tail. In vitro pull-down assays suggested that the C-terminal tail region of NIDD specifically interacted with the PDZ domain of nNOS. The PDZ dependence was confirmed by an experiment using a deletion mutant, and the interaction was further confirmed by co-sedimentation assays using COS-7 cells transfected with NIDD and nNOS. Both NIDD and nNOS were enriched in synaptosome and synaptic plasma membrane fractions and were present in the lipid raft and postsynaptic density fractions in the rat brain. Co-localization of these proteins was also observed by double staining of the proteins in cultured cortical neurons. Thus, NIDD and nNOS were co-localized in the brain, although the colocalizing regions were restricted, as indicated by the distribution of their mRNA expression. Most important, co-transfection of NIDD and nNOS increased NO-producing nNOS activity. These results suggested that NIDD plays an important role in the regulation of the NO signaling pathway at postsynaptic sites through targeting of nNOS to the postsynaptic membrane.
Saitoh
Fuminori
F
Department of Neuroplasticity Institute on Aging and Adaptation, Shinshu University Graduate School of Medicine, 3-1-1 Asahi, Matsumoto 390-8621, Japan.
Tian
Qing Bao
QB
Okano
Akira
A
Sakagami
Hiroyuki
H
Kondo
Hisatake
H
Suzuki
Tatsuo
T
D000595:Amino Acid Sequence; D000818:Animals; D019556:COS Cells; D002352:Carrier Proteins / Q000235:genetics / Q000378:metabolism*; D002458:Cell Fractionation; D002478:Cells, Cultured; D002540:Cerebral Cortex / Q000166:cytology; D002522:Chlorocebus aethiops; D017403:In Situ Hybridization; D008297:Male; D008969:Molecular Sequence Data; D009419:Nerve Tissue Proteins / Q000235:genetics / Q000378:metabolism*; D009474:Neurons / Q000166:cytology / Q000378:metabolism; D019001:Nitric Oxide Synthase / Q000235:genetics / Q000378:metabolism*; D052248:Nitric Oxide Synthase Type I; D017434:Protein Structure, Tertiary; D012333:RNA, Messenger / Q000378:metabolism; D051381:Rats; D017208:Rats, Wistar; D011993:Recombinant Fusion Proteins / Q000235:genetics / Q000378:metabolism; D015398:Signal Transduction / Q000502:physiology; D013570:Synaptic Membranes / Q000737:chemistry / Q000378:metabolism*; D014018:Tissue Distribution
D002352:Carrier Proteins; D009419:Nerve Tissue Proteins; D012333:RNA, Messenger; D011993:Recombinant Fusion Proteins; C488668:Zdhhc23 protein, rat; D019001:Nitric Oxide Synthase; D052248:Nitric Oxide Synthase Type I; C496322:Nos1 protein, rat