title: The ECS(SPSB) E3 ubiquitin ligase is the master regulator of the lifetime of inducible nitric-oxide synthase.
abstract: The ubiquitin-proteasome pathway is an important regulatory system for the lifetime of inducible nitric-oxide synthase (iNOS), a high-output isoform compared to neuronal NOS (nNOS) and endothelial NOS (eNOS), to prevent overproduction of NO that could trigger detrimental effects such as cytotoxicity. Two E3 ubiquitin ligases, Elongin B/C-Cullin-5-SPRY domain- and SOCS box-containing protein [ECS(SPSB)] and the C-terminus of Hsp70-interacting protein (CHIP), recently have been reported to target iNOS for proteasomal degradation. However, the significance of each E3 ubiquitin ligase for the proteasomal degradation of iNOS remains to be determined. Here, we show that ECS(SPSB) specifically interacted with iNOS, but not nNOS and eNOS, and induced the subcellular redistribution of iNOS from dense regions to diffused expression as well as the ubiquitination and proteasomal degradation of iNOS, whereas CHIP neither interacted with iNOS nor had any effects on the subcellular localization, ubiquitination, and proteasomal degradation of iNOS. These results differ from previous reports. Furthermore, the lifetime of the iNOS(N27A) mutant, a form of iNOS that does not bind to ECS(SPSB), was substantially extended in macrophages. These results demonstrate that ECS(SPSB), but not CHIP, is the master regulator of the iNOS lifetime.
authors.affiliation: Department of Cellular Pharmacology, Hokkaido University Graduate School of Medicine, Sapporo, Japan.
authors.forename: Kazuma
authors.initials: K
authors.lastname: Matsumoto
authors.forename: Tadashi
authors.initials: T
authors.lastname: Nishiya
authors.forename: Satoshi
authors.initials: S
authors.lastname: Maekawa
authors.forename: Takahiro
authors.initials: T
authors.lastname: Horinouchi
authors.forename: Kouetsu
authors.initials: K
authors.lastname: Ogasawara
authors.forename: Takashi
authors.initials: T
authors.lastname: Uehara
authors.forename: Soichi
authors.initials: S
authors.lastname: Miwa
mesh_terms: D000595:Amino Acid Sequence; D057809:HEK293 Cells; D006801:Humans; D008264:Macrophages / Q000201:enzymology; D008969:Molecular Sequence Data; D052247:Nitric Oxide Synthase Type II / Q000235:genetics / Q000378:metabolism*; D052250:Nitric Oxide Synthase Type III / Q000235:genetics / Q000378:metabolism; D046988:Proteasome Endopeptidase Complex / Q000378:metabolism; D011485:Protein Binding; D050826:Suppressor of Cytokine Signaling Proteins / Q000378:metabolism*; D044767:Ubiquitin-Protein Ligases / Q000378:metabolism*
chemical_list: D050826:Suppressor of Cytokine Signaling Proteins; C496319:NOS2 protein, human; D052247:Nitric Oxide Synthase Type II; D052250:Nitric Oxide Synthase Type III; C467728:STUB1 protein, human; D044767:Ubiquitin-Protein Ligases; D046988:Proteasome Endopeptidase Complex