掲載種別：研究論文（学術雑誌）   国際・国内誌：国際誌  

DOI： 10.3390/antiox12020350

PubMed

担当区分：最終著者,　責任著者   掲載種別：研究論文（学術雑誌）   国際・国内誌：国際誌  

DOI： 10.3389/fmolb.2023.1089213

PubMed

掲載種別：研究論文（学術雑誌）  

DOI： 10.1186/s40478-022-01486-6

PubMed

掲載種別：研究論文（学術雑誌）  

DOI： 10.3892/ol.2022.13514

PubMed

掲載種別：研究論文（学術雑誌）  

掲載種別：研究論文（学術雑誌）  

掲載種別：研究論文（学術雑誌）  

掲載種別：研究論文（学術雑誌）  

健康人の唾液蛋白質(SP)がアンジオテンシン変換酵素2(ACE2)に結合する能力を評価し、新型コロナウイルス(SARS-CoV-2)スパイク蛋白質S1(S1)受容体結合ドメインとACE2の間の結合に及ぼすSPの影響を測定した。SPはACE2に結合し、S1のACE2への結合を妨害し、S1-ACE2相互作用を阻害するカチオン性ヒストンH2Aと好中球エラスターゼを含む4種類のACE2結合性SPを同定した。ウシ胸腺ヒストン(CTH)もH2Aと同じく効果的に結合を阻害した。CTHはACE2発現宿主細胞へのSARS-CoV-2偽ウイルス性侵入を抑制した。ε-ポリ-L-リジンなどの合成ポリペプチドもS1-ACE2結合を妨害し、ACE2結合におけるSPの重要性が示された。以上より、正荷電のSPはACE2の負荷電表面をクローキングしてSARS-CoV-2の侵入に対する障壁になり、カチオン性ポリペプチドは新型コロナウイルス感染症に対する予防的・治療的手法になり得ると考えられた。

掲載種別：研究論文（学術雑誌）   国際・国内誌：国際誌  

DOI： 10.1093/jb/mvac054

PubMed

掲載種別：研究論文（学術雑誌）  

DOI： 10.1038/s41419-022-05145-5

PubMed

その他URL： https://www.nature.com/articles/s41419-022-05145-5

掲載種別：研究論文（学術雑誌）  

DOI： 10.3390/cells11152398

PubMed

掲載種別：研究論文（学術雑誌）   国際・国内誌：国際誌  

DOI： 10.1073/pnas.2202125119.

掲載種別：研究論文（学術雑誌）  

DOI： 10.1016/j.redox.2022.102286

PubMed

掲載種別：研究論文（学術雑誌）   国際・国内誌：国際誌  

DOI： 10.1038/s41389-022-00389-4

PubMed

掲載種別：研究論文（学術雑誌）   国際・国内誌：国際誌  

DOI： 10.1042/BST20211078

PubMed

掲載種別：研究論文（学術雑誌）  

DOI： 10.1038/s41598-021-02522-6

PubMed

掲載種別：研究論文（学術雑誌）   国際・国内誌：国際誌  

DOI： 10.1016/j.celrep.2021.109988

PubMed

掲載種別：研究論文（学術雑誌）  

掲載種別：研究論文（学術雑誌）  

掲載種別：研究論文（学術雑誌）   国際・国内誌：国際誌  

DOI： 10.1002/hep.31752

PubMed

掲載種別：研究論文（学術雑誌）   国際・国内誌：国際誌  

DOI： 10.1007/s00011-021-01458-x

PubMed

掲載種別：研究論文（学術雑誌）  

DOI： 10.1007/s00011-021-01458-x

PubMed

掲載種別：研究論文（学術雑誌）   国際・国内誌：国際誌  

DOI： 10.1016/j.jaad.2020.05.051

PubMed

掲載種別：研究論文（学術雑誌）  

BACKGROUND & AIMS: Anti-fibrotic therapy remains an unmet medical need in human chronic liver disease. We report the anti-fibrotic properties of cytoglobin (CYGB), a respiratory protein expressed in hepatic stellate cells (HSCs), the main cell type involved in liver fibrosis. APPROACH & RESULTS: Cygb-deficient mice which had bile duct ligation (BDL)-induced liver cholestasis or choline-deficient L-amino acid-defined (CDAA) diet-induced steatohepatitis significantly exacerbated liver damage, fibrosis and reactive oxygen species (ROS) formation. All these manifestations were attenuated in Cygb-overexpressing mice. We produced 6His-tagged recombinant human CYGB (His-CYGB), traced its bio-distribution and assessed its function in HSCs or in mice with advanced liver cirrhosis using thioacetamide (TAA) or 3,5-diethoxycarbonyl-1,4-dihydrocollidine (DDC). In cultured HSCs, extracellular His-CYGB was endocytosed and accumulated in endosomes via clathrin-mediated pathway. His-CYGB significantly impeded ROS formation spontaneously or in the presence of ROS inducers in HSCs, thus leading to the attenuation of collagen type I alpha 1 production and alpha-smooth muscle actin expression. Replacement the iron centre of the heme group with cobalt nullified the effect of His-CYGB. In addition, His-CYGB induced interferon-β secretion by HSCs which partly contributed to its anti-fibrotic function. Momelotinib incompletely reversed the effect of His-CYGB. Intravenously injected His-CYGB markedly suppressed liver inflammation, fibrosis and oxidative cell damage in TAA- or DDC-administered mice without adverse effects. RNA-seq analysis revealed the downregulation of inflammation and fibrosis-related genes and the upregulation of antioxidant genes in both cell culture and liver tissues. The injected His-CYGB predominantly localised to HSCs but not to macrophages, suggesting specific targeting effects. His-CYGB exhibited no toxicity in humanised liver chimeric PXB mice. CONCLUSIONS: His-CYGB could have anti-fibrotic clinical applications for human chronic liver diseases.

DOI： 10.1002/hep.31752

PubMed

掲載種別：研究論文（学術雑誌）  

Mind bomb 2 (MIB2) is an E3 ligase involved in Notch signalling and attenuates TNF-induced apoptosis through ubiquitylation of receptor-interacting protein kinase 1 (RIPK1) and cylindromatosis. Here we show that MIB2 bound and conjugated K48- and K63-linked polyubiquitin chains to a long-form of cellular FLICE-inhibitory protein (cFLIPL), a catalytically inactive homologue of caspase 8. Deletion of MIB2 did not impair the TNF-induced complex I formation that mediates NF-κB activation but significantly enhanced formation of cytosolic death-inducing signalling complex II. TNF-induced RIPK1 Ser166 phosphorylation, a hallmark of RIPK1 death-inducing activity, was enhanced in MIB2 knockout cells, as was RIPK1 kinase activity-dependent and -independent apoptosis. Moreover, RIPK1 kinase activity-independent apoptosis was induced in cells expressing cFLIPL mutants lacking MIB2-dependent ubiquitylation. Together, these results suggest that MIB2 suppresses both RIPK1 kinase activity-dependent and -independent apoptosis, through suppression of RIPK1 kinase activity and ubiquitylation of cFLIPL, respectively.

DOI： 10.1038/s42003-020-01603-y

PubMed

掲載種別：研究論文（学術雑誌）   国際・国内誌：国際誌  

DOI： 10.1038/s42003-020-01603-y

PubMed

掲載種別：研究論文（学術雑誌）  

DOI： 10.1016/j.bbrc.2020.12.045

PubMed

掲載種別：研究論文（学術雑誌）  

DOI： 10.1016/j.bbrc.2020.12.045

PubMed

掲載種別：研究論文（学術雑誌）  

DOI： 10.3389/fimmu.2021.635475

PubMed

掲載種別：研究論文（学術雑誌）  

Histamine, which is mainly produced by mast cells and basophils, participates in various allergic symptoms, and some studies have reported that macrophages also produce histamine. Moreover, recent studies have revealed that macrophages, especially alternatively activated macrophages (M2) induced by T helper 2 (Th2) cytokines, such as interleukin (IL)-4 and IL-13, participate in the pathogenesis of allergic diseases. The major source of Th2 cytokines is antigen-specific Th2 cells. To elucidate the relationship between histamine, macrophages, and Th2 cells in allergic inflammation, we established a macrophage-Th2 cell co-culture model in vitro and an antigen-specific Th2 cell transfer mouse model of rhinitis. In vitro analyses indicated that macrophages produce histamine by interacting with antigen-specific Th2 cells through the antigen. Furthermore, Th2 cells and macrophages cooperatively elicited rhinitis in the mouse model. We determined that histamine induces Th2- and macrophage-elicited sneezing responses through H1 receptor signaling, whereas it induces nasal eosinophil infiltrations through H4 receptor signaling. Collectively, these results indicate a novel histamine production mechanism by macrophages, in which Th2 cells and macrophages cooperatively induce nasal allergic inflammation through histamine signaling.

DOI： 10.1371/journal.pone.0248158

PubMed

掲載種別：研究論文（学術雑誌）  

DOI： 10.15036/arerugi.70.1239

CiNii Article

掲載種別：論文集(書籍)内論文  

DOI： 10.1007/978-981-16-4866-3_14

掲載種別：研究論文（学術雑誌）   国際・国内誌：国際誌  

DOI： 10.3389/fimmu.2021.635475

PubMed

掲載種別：研究論文（学術雑誌）  

掲載種別：研究論文（学術雑誌）   国際・国内誌：国際誌  

DOI： 10.1371/journal.pone.0248158

PubMed

掲載種別：研究論文（学術雑誌）  

The LUBAC ubiquitin ligase complex, composed of the HOIP, HOIL-1L, and SHARPIN subunits, stimulates the canonical nuclear factor-κB (NF-κB) activation pathways through its Met1-linked linear ubiquitination activity. Here we performed cellular and mathematical modeling analyses of the LUBAC involvement in the T cell receptor (TCR)-mediated NF-κB activation pathway, using the Jurkat human T cell line. LUBAC is indispensable for TCR-induced NF-κB and T cell activation, and transiently associates with and linearly ubiquitinates the CARMA1-BCL10-MALT1 (CBM) complex, through the catalytic HOIP subunit. In contrast, the linear ubiquitination of NEMO, a substrate of the TNF-α-induced canonical NF-κB activation pathway, was limited during the TCR pathway. Among deubiquitinases, OTULIN, but not CYLD, plays a major role in downregulating LUBAC-mediated TCR signaling. Mathematical modeling indicated that linear ubiquitination of the CBM complex accelerates the activation of IκB kinase (IKK), as compared with the activity induced by linear ubiquitination of NEMO alone. Moreover, simulations of the sequential linear ubiquitination of the CBM complex suggested that the allosteric regulation of linear (de)ubiquitination of CBM subunits is controlled by the ubiquitin-linkage lengths. These results indicated that, unlike the TNF-α-induced NF-κB activation pathway, the TCR-mediated NF-κB activation in T lymphocytes has a characteristic mechanism to induce LUBAC-mediated NF-κB activation.

DOI： 10.3389/fimmu.2020.601926

PubMed

掲載種別：研究論文（学術雑誌）  

掲載種別：研究論文（学術雑誌）  

掲載種別：研究論文（学術雑誌）  

掲載種別：研究論文（学術雑誌）  

DOI： 10.3390/biomedicines8060152

PubMed

掲載種別：研究論文（学術雑誌）  

Protein ubiquitinations play pivotal roles in many cellular processes, including homeostasis, responses to various stimulations, and progression of diseases. Deubiquitinating enzymes (DUBs) remove ubiquitin molecules from ubiquitinated proteins and cleave the polyubiquitin chain, thus negatively regulating numerous ubiquitin-dependent processes. Dysfunctions of many DUBs reportedly cause various diseases; therefore, DUBs are considered as important drug targets, although the biochemical characteristics and cellular functions of many DUBs are still unclear. Here, we established a human DUB protein array to detect the activity and linkage specificity of almost all human DUBs. Using a wheat cell-free protein synthesis system, 88 full-length recombinant human DUB proteins were prepared and termed the DUB array. In vitro DUB assays were performed with all of these recombinant DUBs, using eight linkage types of diubiquitins as substrates. As a result, 80 DUBs in the array showed DUB activities, and their linkage specificities were determined. These 80 DUBs included many biochemically uncharacterized DUBs in the past. In addition, taking advantage of these active DUB proteins, we applied the DUB array to evaluate the selectivities of DUB inhibitors. We successfully developed a high-throughput and semi-quantitative DUB assay based on AlphaScreen technology, and a model study using two commercially available DUB inhibitors revealed individual selectivities to 29 DUBs, as previously reported. In conclusion, the DUB array established here is a powerful tool for biochemical analyses and drug discovery for human DUBs.

DOI： 10.3390/biomedicines8060152

PubMed

掲載種別：研究論文（学術雑誌）  

DOI： 10.1016/j.jaad.2020.05.051

PubMed

掲載種別：研究論文（学術雑誌）   国際・国内誌：国際誌  

DOI： 10.3390/ijms21093381

PubMed

掲載種別：研究論文（学術雑誌）  

DOI： 10.3390/ijms21093381

PubMed

掲載種別：研究論文（学術雑誌）   国際・国内誌：国際誌  

DOI： 10.1038/s42003-020-0882-8

PubMed

掲載種別：研究論文（学術雑誌）  

DOI： 10.1016/j.bbrc.2019.12.049

PubMed

掲載種別：研究論文（学術雑誌）  

DOI： 10.1093/jnen/nlz135

PubMed

掲載種別：研究論文（学術雑誌）   国際・国内誌：国際誌  

DOI： 10.1093/jnen/nlz135

PubMed

掲載種別：研究論文（学術雑誌）  

脱ユビキチン化酵素（DUB）は，タンパク質分解やシグナル伝達の活性化などのユビキチン化修飾を介した細胞応答を負に制御する重要な因子として注目されている．また，DUBの高発現や異常な活性化が，がんや神経疾患などを誘導することが明らかとなっており，創薬ターゲットとしても注目されている．我々はコムギ無細胞系を用いてヒトのDUBの約8割を網羅する83種類の組換えタンパク質を合成し，DUBタンパク質アレイを作製した．このDUBタンパク質アレイについて，8種類の重合様式の二量体ユビキチンを基質として，76種類のDUBについて活性を検出し，そのユビキチン鎖特異性を明らかにした．さらに，このDUBタンパク質アレイを応用することで，DUBを阻害する化合物のDUB特異性評価パネルを作製した．

DOI： 10.14952/seikagaku.2020.920064

CiNii Article

掲載種別：研究論文（学術雑誌）  

ユビキチンのN末端を介して形成される非定型型の直鎖状ユビキチン鎖は，炎症・免疫シグナル応答や細胞死を制御し，その生成を特異的に担うLUBACユビキチンリガーゼの機能不全は多くの疾患発症に関わる．筆者らは家族性筋萎縮性側索硬化症（ALS）の原因遺伝子の一つであるオプチニューリン（OPTN）やアルツハイマー病タウタンパク質の細胞機能解析から，LUBACによる直鎖状ユビキチン鎖生成が異常タンパク質封入体形成，神経炎症・細胞死の亢進に寄与することを見いだした．さらに，新規治療薬創出を最終目的としてLUBACに対する阻害剤の探索を行い，細胞毒性が低くLUBAC特異性の高い新規化合物（HOIPINs）を見いだした．本稿では，筆者らの研究を中心に国内外の動向を交えて，これらの知見を紹介したい．

DOI： 10.14952/seikagaku.2020.920028

CiNii Article

掲載種別：研究論文（学術雑誌）   共著区分：単著  

ユビキチンは,タンパク質に結合して選択的にプロテアソーム分解へ導く目印として発見されたが,ユビキチン分子間で多様な連結を形成することで分解以外にも数多くの生理機能制御を担うことが明らかになってきた.本稿では,我々が見出した直鎖状ユビキチン鎖を生成する酵素（LUBAC，linear ubiquitin chain assembly complex)の生理機能と疾患との連関,および創薬を目指した阻害剤開発の現状を紹介したい.

DOI： 10.14894/faruawpsj.56.1_26

CiNii Article

共著区分：共著  

共著区分：共著  

掲載種別：研究論文（学術雑誌）  

掲載種別：研究論文（学術雑誌）  

ユビキチンは,タンパク質に結合して選択的にプロテアソーム分解へ導く目印として発見されたが,ユビキチン分子間で多様な連結を形成することで分解以外にも数多くの生理機能制御を担うことが明らかになってきた.本稿では,我々が見出した直鎖状ユビキチン鎖を生成する酵素（LUBAC，linear ubiquitin chain assembly complex)の生理機能と疾患との連関,および創薬を目指した阻害剤開発の現状を紹介したい.

DOI： 10.14894/faruawpsj.56.1_26

CiNii Article

掲載種別：研究論文（学術雑誌）  

脱ユビキチン化酵素(DUB)は,タンパク質分解やシグナル伝達の活性化などのユビキチン化修飾を介した細胞応答を負に制御する重要な因子として注目されている.また,DUBの高発現や異常な活性化が,がんや神経疾患などを誘導することが明らかとなっており,創薬ターゲットとしても注目されている.我々はコムギ無細胞系を用いてヒトのDUBの約8割を網羅する83種類の組換えタンパク質を合成し,DUBタンパク質アレイを作製した.このDUBタンパク質アレイについて,8種類の重合様式の二量体ユビキチンを基質として,76種類のDUBについて活性を検出し,そのユビキチン鎖特異性を明らかにした.さらに,このDUBタンパク質アレイを応用することで,DUBを阻害する化合物のDUB特異性評価パネルを作製した.(著者抄録)

掲載種別：研究論文（学術雑誌）   国際・国内誌：国際誌  

DOI： 10.3389/fimmu.2020.601926

PubMed

掲載種別：研究論文（学術雑誌）  

DOI： 10.1016/j.bbrc.2019.12.049

PubMed

掲載種別：研究論文（学術雑誌）  

DOI： 10.1074/jbc.RA119.010119

PubMed

掲載種別：研究論文（学術雑誌）   国際・国内誌：国際誌  

DOI： 10.1074/jbc.RA119.010119

PubMed

掲載種別：研究論文（学術雑誌）   国際・国内誌：国際誌  

DOI： 10.1016/j.neulet.2019.03.017

PubMed

掲載種別：研究論文（学術雑誌）  

掲載種別：研究論文（学術雑誌）  

掲載種別：研究論文（学術雑誌）   共著区分：共著  

DOI： 10.1016/j.neulet.2019.03.017

PubMed

掲載種別：研究論文（学術雑誌）   共著区分：共著  

DOI： 10.1016/j.bbrc.2018.12.164

PubMed

掲載種別：研究論文（学術雑誌）  

DOI： 10.1016/j.bbrc.2018.12.164

PubMed

掲載種別：研究論文（学術雑誌）  

DOI： 10.1177/2472555218793066

PubMed

掲載種別：研究論文（学術雑誌）  

DOI： 10.1177/2472555218793066

PubMed

掲載種別：研究論文（学術雑誌）  

DOI： 10.1038/s41467-018-06922-7

PubMed

掲載種別：研究論文（学術雑誌）   国際・国内誌：国際誌  

DOI： 10.1038/s41467-018-06922-7

PubMed

掲載種別：研究論文（学術雑誌）  

DOI： 10.1089/mab.2018.0023

PubMed

掲載種別：研究論文（学術雑誌）  

DOI： 10.1089/mab.2018.0023

PubMed

掲載種別：研究論文（学術雑誌）  

KIT遺伝子はまだら症の原因遺伝子であり、これまでに60以上の変異が同定されているが、変異KIT蛋白質の機能と細胞内動態については十分に解明されていないことから、遺伝子解析とin vivo研究を行った。まだら症の発端者である3歳の日本人女児とその家族(両親、母方の祖母・祖父)の末梢血細胞を用いてKIT遺伝子の遺伝子解析を行った。さらに、野生型または変異KIT遺伝子でトランスフェクトしたHEK293T細胞における変異KIT蛋白質の細胞内局在を解析した。遺伝子解析により、KITの細胞外ドメインにおいて、Val216およびSer217のインフレーム欠失をもたらす新規ヘテロ接合変異c.645_650delTGTGTCが明らかになった。免疫沈降分析により、幹細胞因子(SCF)による処理後に野生型および変異KITがヘテロ二量体を形成することが確認されたが、下流のシグナル伝達因子のリン酸化は減少した。免疫蛍光研究では、変異KITは主に小胞体に蓄積し、細胞表面上にまばらに発現していることが示された。グリコシダーゼ消化研究により、変異KITは主にERに小胞体に局在することが明らかになった。

掲載種別：研究論文（学術雑誌）  

DOI： 10.1016/j.jdermsci.2018.03.012

PubMed

掲載種別：研究論文（学術雑誌）  

DOI： 10.1089/mab.2018.0005

PubMed

掲載種別：研究論文（学術雑誌）  

DOI： 10.1089/mab.2018.0005

PubMed

掲載種別：研究論文（学術雑誌）  

DOI： 10.1111/1346-8138.14149

PubMed

掲載種別：研究論文（学術雑誌）  

DOI： 10.1111/1346-8138.14149

PubMed

掲載種別：研究論文（学術雑誌）  

DOI： 10.1016/j.jdermsci.2018.03.012

PubMed

掲載種別：研究論文（学術雑誌）  

掲載種別：研究論文（学術雑誌）  

掲載種別：研究論文（学術雑誌）  

掲載種別：研究論文（学術雑誌）  

DOI： 10.1111/bjd.15342

掲載種別：研究論文（学術雑誌）  

掲載種別：研究論文（学術雑誌）  

DOI： 10.1111/bjd.15342

PubMed

掲載種別：研究論文（学術雑誌）  

掲載種別：研究論文（学術雑誌）  

DOI： 10.1016/J.bbrc.2017.02.040

PubMed

掲載種別：研究論文（学術雑誌）  

DOI： 10.1016/j.bbrc.2017.02.040

PubMed

掲載種別：研究論文（学術雑誌）  

DOI： 10.3390/ijms18020326

PubMed

掲載種別：研究論文（学術雑誌）  

DOI： 10.3390/ijms18020326

PubMed

掲載種別：研究論文（学術雑誌）  

DOI： 10.1371/journal.ppat.1006162

PubMed

掲載種別：研究論文（学術雑誌）  

DOI： 10.1371/journal.ppat.1006162

PubMed

掲載種別：研究論文（学術雑誌）   共著区分：単著  

LRBA(LPS-Responsive and Beige-like Anchor protein)は炎症・免疫制御に関わるタンパク質であり、その欠損がB細胞に機能不全をきたし、抗体産生低下を主徴とする分類不能型免疫不全(CVID)を発症するのではないかと考え検討を行った。CRISPR/Cas9法を用いてLrba遺伝子をノックアウトしたマウス胎児性線維芽細胞(MEF)を構築し、種々の刺激に対するシグナル応答性を評価した。各種自然・獲得免疫刺激によって惹起するシグナル伝達経路活性化への影響を野生型MEFとLrba-/-MEF間で比較した。その結果、polyI:C刺激で誘導されるIFN産生経路がLrba-/-MEFでは亢進していることが明らかとなり、Lrbaは生理的にはIFN産生経路を抑制することが示された。次いで、CRISPR/Cas9法を用いてLrba-KOマウスを作製し、Lrba mRNAの発現を解析したところ、Lrbaは各臓器で普遍的に発現しているが、免疫応答に関わる組織では特に発現レベルが高い可能性が示唆された。

掲載種別：研究論文（学術雑誌）  

LRBA(LPS-Responsive and Beige-like Anchor protein)は炎症・免疫制御に関わるタンパク質であり、その欠損がB細胞に機能不全をきたし、抗体産生低下を主徴とする分類不能型免疫不全(CVID)を発症するのではないかと考え検討を行った。CRISPR/Cas9法を用いてLrba遺伝子をノックアウトしたマウス胎児性線維芽細胞(MEF)を構築し、種々の刺激に対するシグナル応答性を評価した。各種自然・獲得免疫刺激によって惹起するシグナル伝達経路活性化への影響を野生型MEFとLrba-/-MEF間で比較した。その結果、polyI:C刺激で誘導されるIFN産生経路がLrba-/-MEFでは亢進していることが明らかとなり、Lrbaは生理的にはIFN産生経路を抑制することが示された。次いで、CRISPR/Cas9法を用いてLrba-KOマウスを作製し、Lrba mRNAの発現を解析したところ、Lrbaは各臓器で普遍的に発現しているが、免疫応答に関わる組織では特に発現レベルが高い可能性が示唆された。

CiNii Article

掲載種別：研究論文（学術雑誌）  

LRBA(LPS-Responsive and Beige-like Anchor protein)は炎症・免疫制御に関わるタンパク質であり、その欠損がB細胞に機能不全をきたし、抗体産生低下を主徴とする分類不能型免疫不全(CVID)を発症するのではないかと考え検討を行った。CRISPR/Cas9法を用いてLrba遺伝子をノックアウトしたマウス胎児性線維芽細胞(MEF)を構築し、種々の刺激に対するシグナル応答性を評価した。各種自然・獲得免疫刺激によって惹起するシグナル伝達経路活性化への影響を野生型MEFとLrba-/-MEF間で比較した。その結果、polyI:C刺激で誘導されるIFN産生経路がLrba-/-MEFでは亢進していることが明らかとなり、Lrbaは生理的にはIFN産生経路を抑制することが示された。次いで、CRISPR/Cas9法を用いてLrba-KOマウスを作製し、Lrba mRNAの発現を解析したところ、Lrbaは各臓器で普遍的に発現しているが、免疫応答に関わる組織では特に発現レベルが高い可能性が示唆された。

掲載種別：研究論文（学術雑誌）  

DOI： 10.1038/srep34756

PubMed

掲載種別：研究論文（学術雑誌）  

DOI： 10.1038/srep34756

PubMed

掲載種別：研究論文（学術雑誌）  

DOI： 10.1038/ncomms12547

PubMed

掲載種別：研究論文（学術雑誌）  

DOI： 10.1038/ncomms12547

PubMed

掲載種別：研究論文（学術雑誌）  

The Skp1-Cul1-F-box protein (SCF) complex catalyzes protein ubiquitination in diverse cellular processes and is one of the best-characterized ubiquitin ligases. F-box proteins determine the substrate specificities of SCF ubiquitin ligases. Among these, Fbs1/FBG1/FBXO2, Fbs2/FBG2/FBXO6, and Fbs3/FBG5/FBXO27 recognize the N-glycans of glycoproteins, whereas FBG3/FBXO44 has no sugar-binding activity, despite the high sequence homology and conservation of the residues necessary for oligosaccharide binding between Fbs1-3 and FBG3. Here we determined the crystal structure of the Skp1-FBG3 complex at a resolution of 2.6 angstrom. The substrate-binding domain of FBG3 is composed of a 10-stranded antiparallel beta-sandwich with three helices. Although the overall structure of FBG3 is similar to that of Fbs1, the residues that form the Fbs1 carbohydrate-binding pocket failed to be superposed with the corresponding residues of FBG3. Structure-based mutational analysis shows that distinct hydrogen bond networks of four FBG3 loops, i.e., beta 2-beta 3, beta 5-beta 6, beta 7-beta 8, and beta 9-beta 10, prevent the formation of the carbohydrate-binding pocket shown in Fbs1.

DOI： 10.1371/journal.pone.0140366

PubMed

掲載種別：研究論文（学術雑誌）  

DOI： 10.1038/jid.2015.12

PubMed

掲載種別：研究論文（学術雑誌）  

Monilethrix is a hair shaft anomaly characterized by beaded hair with periodic changes in hair thickness. Mutations in the desmoglein 4 (DSG4) gene reportedly underlie the autosomal recessive form of the disease. However, the pathogenesis and cellular basis for the DSG4 mutation induced monilethrix remained largely unknown. We report a Japanese female patient with monilethrix. Observation of her hair shaft by means of transmission electron microscopy showed fewer desmosomes and abnormal keratinization. Genetic analysis revealed a homozygous mutation, c.2119delG (p.Asp707Ilefs*109), in the DSG4 gene, which was predicted to cause a frameshift and premature termination in the intracellular region of the DSG4 protein. The mutation has not been reported previously. In the patient's hair shaft, we detected reduced but partial expression of the mutant DSG4 protein. Cellular analyses demonstrated that the mutant DSG4 lost its affinity to plakoglobin and accumulated in the endoplasmic reticulum (ER). The amounts of mutant DSG4 were increased by proteasome inhibitor treatment, and the expression of an ER chaperone, GRP78/BiP, was elevated in the patient's skin. Collectively, these results suggest that the dysfunctional mutated DSG4, tethered in the ER, undergoes ER-associated degradation, leading to unfolded protein response induction, and thus ER stress may have a role in the pathogenesis of monilethrix.

DOI： 10.1038/jid.2015.12

PubMed

掲載種別：研究論文（学術雑誌）  

DOI： 10.1038/nsmb.2970

PubMed

掲載種別：研究論文（学術雑誌）  

The tumor suppressor CYLD belongs to a ubiquitin (Ub)-specific protease (USP) family and specifically cleaves Met1- and Lys63-linked polyubiquitin chains to suppress inflammatory signaling pathways. Here, we report crystal structures representing the catalytic states of zebrafish CYLD for Met1- and Lys63-linked Ub chains and two distinct precatalytic states for Met1-linked chains. In both catalytic states, the distal Ub is bound to CYLD in a similar manner, and the scissile bond is located close to the catalytic residue, whereas the proximal Ub is bound in a manner specific to Met1- or Lys63-linked chains. Further structure-based mutagenesis experiments support the mechanism by which CYLD specifically cleaves both Met1- and Lys63-linked chains and provide insight into tumor-associated mutations of CYLD. This study provides new structural insight into the mechanisms by which USP family deubiquitinating enzymes recognize and cleave Ub chains with specific linkage types.

DOI： 10.1038/nsmb.2970

PubMed

掲載種別：研究論文（学術雑誌）  

DOI： 10.1155/2015/125380

PubMed

掲載種別：研究論文（学術雑誌）  

DOI： 10.1371/journal.pone.0140366

PubMed

掲載種別：研究論文（学術雑誌）  

Nonalcoholic steatohepatitis (NASH) is a disorder characterized by hepatic lipid accumulation followed by the inflammation-induced death of hepatocytes and fibrosis. In this process, oxidative stress contributes to the induction of several inflammatory cytokines including TNF-alpha and IL-1 beta in macrophages, while, in hepatocytes, NF-kappa B reportedly induces the expressions of cell survival genes for protection from apoptosis. Recently, it was reported that the new ubiquitin ligase complex termed linear ubiquitin chain assembly complex (LUBAC), composed of SHARPIN (SHANK-associated RH domain-interacting protein), HOIL-1L (longer isoform of heme-oxidized iron-regulatory protein 2 ubiquitin ligase-1), and HOIP (HOIL-1L interacting protein), forms linear ubiquitin on NF-kappa B essential modulator (NEMO) and thereby induces NF-kappa B pathway activation. In this study, we demonstrated the formation of LUBAC to be impaired in the livers of NASH rodent models produced by methionine and choline deficient (MCD) diet feeding, first by either gel filtration or Blue Native-PAGE, with subsequent confirmation by western blotting. The reduction of LUBAC is likely to be attributable to markedly reduced expression of SHARPIN, one of its components. Thus, impaired LUBAC formation, which would result in insufficient NF-kappa B activation, may be one of the molecular mechanisms underlying the enhanced apoptotic response of hepatocytes in MCD diet-induced NASH livers.

DOI： 10.1155/2015/125380

PubMed

掲載種別：研究論文（学術雑誌）  

掲載種別：研究論文（学術雑誌）  

掲載種別：研究論文（学術雑誌）  

掲載種別：研究論文（学術雑誌）   共著区分：単著  

B細胞リンパ腫におけるユビキチン系の寄与について検討した。NF-κB経路の制御に関わることが報告されていたOTU型のA20(TNFAIP3)とCezanne、USP型のCYLDに着目して抑制能を解析した。A20とCYLDは発現量を増加させるとユビキチンリガーゼ(LUBAC)発現によるNF-κB活性化を強く阻害するが、Cezanneは抑制効果が低かった。CYLDは脱ユビキチン化酵素活性依存的にLUBACによるNF-κB活性化を抑制した。A20がZF7を介して直鎖状ユビキチンに特異的に結合することでNF-κB活性を制御することが示唆された。A20はZF7の直鎖状ユビキチン結合能を介して受容体近傍に集積していること、その不全はNF-κB活性の持続的亢進となりB細胞リンパ腫を発症させることが示唆された。

掲載種別：研究論文（学術雑誌）  

B細胞リンパ腫におけるユビキチン系の寄与について検討した。NF-κB経路の制御に関わることが報告されていたOTU型のA20(TNFAIP3)とCezanne、USP型のCYLDに着目して抑制能を解析した。A20とCYLDは発現量を増加させるとユビキチンリガーゼ(LUBAC)発現によるNF-κB活性化を強く阻害するが、Cezanneは抑制効果が低かった。CYLDは脱ユビキチン化酵素活性依存的にLUBACによるNF-κB活性化を抑制した。A20がZF7を介して直鎖状ユビキチンに特異的に結合することでNF-κB活性を制御することが示唆された。A20はZF7の直鎖状ユビキチン結合能を介して受容体近傍に集積していること、その不全はNF-κB活性の持続的亢進となりB細胞リンパ腫を発症させることが示唆された。

CiNii Article

掲載種別：研究論文（学術雑誌）  

B細胞リンパ腫におけるユビキチン系の寄与について検討した。NF-κB経路の制御に関わることが報告されていたOTU型のA20(TNFAIP3)とCezanne、USP型のCYLDに着目して抑制能を解析した。A20とCYLDは発現量を増加させるとユビキチンリガーゼ(LUBAC)発現によるNF-κB活性化を強く阻害するが、Cezanneは抑制効果が低かった。CYLDは脱ユビキチン化酵素活性依存的にLUBACによるNF-κB活性化を抑制した。A20がZF7を介して直鎖状ユビキチンに特異的に結合することでNF-κB活性を制御することが示唆された。A20はZF7の直鎖状ユビキチン結合能を介して受容体近傍に集積していること、その不全はNF-κB活性の持続的亢進となりB細胞リンパ腫を発症させることが示唆された。

掲載種別：研究論文（学術雑誌）  

DOI： 10.1083/jcb.201304188

PubMed

掲載種別：研究論文（学術雑誌）   共著区分：単著  

DOI： 10.1093/jb/mvt079

PubMed

掲載種別：研究論文（学術雑誌）  

DOI： 10.1093/jb/mvt079

PubMed

掲載種別：研究論文（学術雑誌）  

The detection of cytosolic DNA, derived from pathogens or host cells, by cytosolic receptors is essential for appropriate host immune responses. Cyclic GMP-AMP synthase (cGAS) is a newly identified cytosolic DNA receptor that produces cyclic GMP-AMP, which activates stimulator of interferon genes (STING), resulting in TBK1-IRF3 pathway activation followed by the production of type I interferons. Here we report the crystal structure of human cGAS. The structure revealed that a cluster of lysine and arginine residues forms the positively charged DNA binding surface of human cGAS, which is important for the STING-dependent immune activation. A structural comparison with other previously determined cGASs and our functional analyses suggested that a conserved zinc finger motif and a leucine residue on the DNA binding surface are crucial for the DNA-specific immune response of human cGAS, consistent with previous work. These structural features properly orient the DNA binding to cGAS, which is critical for DNA-induced cGAS activation and STING-dependent immune activation. Furthermore, we showed that the cGAS-induced activation of STING also involves the activation of the NF-kappa B and IRF3 pathways. Our results indicated that cGAS is a DNA sensor that efficiently activates the host immune system by inducing two distinct pathways.

DOI： 10.1371/journal.pone.0076983

PubMed

掲載種別：研究論文（学術雑誌）  

Although ubiquitin is thought to be important for the autophagic sequestration of invading bacteria (also called xenophagy), its precise role remains largely enigmatic. Here we determined how ubiquitin is involved in this process. After invasion, ubiquitin is conjugated to host cellular proteins in endosomes that contain Salmonella or transfection reagent-coated latex (polystyrene) beads, which mimic invading bacteria. Ubiquitin is recognized by the autophagic machinery independently of the LC3-ubiquitin interaction through adaptor proteins, including a direct interaction between ubiquitin and Atg16L1. To ensure that invading pathogens are captured and degraded, Atg16L1 targeting is secured by two backup systems that anchor Atg16L1 to ubiquitin-decorated endosomes. Thus, we reveal that ubiquitin is a pivotal molecule that connects bacteria-containing endosomes with the autophagic machinery upstream of LC3.

DOI： 10.1083/jcb.201304188

PubMed

掲載種別：研究論文（学術雑誌）  

掲載種別：研究論文（学術雑誌）  

PubMed

掲載種別：研究論文（学術雑誌）  

DOI： 10.1371/journal.pone.0076983

PubMed

掲載種別：研究論文（学術雑誌）  

DOI： 10.1038/emboj.2012.241

PubMed

掲載種別：研究論文（学術雑誌）  

DOI： 10.1007/s12104-011-9350-1

PubMed

掲載種別：研究論文（学術雑誌）   国際・国内誌：国際誌  

HOIL-1L and its binding partner, HOIL-1L interacting protein (HOIP), are essential components of linear ubiquitin (Ub) chain assembly complex (LUBAC), a 600-kDa enzyme complex catalyzing elongation of a tandemly connected Ub chain, which serve as a regulator of NF-κB activation. Specific interaction between the N-terminal Ub-like domain (UBL) of HOIL-1L and the Ub-associated domain (UBA) located at the central region of HOIP is shown to be involved in the formation of LUBAC. For better understanding of the mechanisms underlying the generation of the linear Ub chains by LUBAC, it is necessary to characterize the UBL-UBA interaction on the basis of structural data, which, however, is not available to date. Here we report backbone and side chain NMR assignments of the UBL of human HOIL-1L. By inspection of chemical shift index, it was predicted that HOIL-1L-UBL assumes a Ub fold followed by an α-helical segment, offering the basis for determination its 3D structure and interaction with HOIP-UBA in solution.

DOI： 10.1007/s12104-011-9350-1

PubMed

掲載種別：研究論文（学術雑誌）  

DOI： 10.1038/emboj.2012.241

PubMed

掲載種別：研究論文（学術雑誌）  

DOI： 10.1507/endocrj.EJ12-0148

PubMed

掲載種別：研究論文（学術雑誌）  

DOI： 10.1074/jbc.M112.347195

PubMed

掲載種別：研究論文（学術雑誌）  

DOI： 10.1016/j.micinf.2012.01.011

PubMed

掲載種別：研究論文（学術雑誌）  

DOI： 10.1074/jbc.M112.347195

PubMed

掲載種別：研究論文（学術雑誌）  

LUBAC (linear ubiquitin chain assembly complex) is a ubiquitin ligase complex composed of SHARPIN, HOIL-IL and HOIP that generates linear polyubiquitin chains and regulates the NF-kappa B pathway, which is pivotal in inflammatory and immune responses. Recent findings on the regulation of NF-kappa B by LUBAC and the diseases associated with this process are the focus of this review. (C) 2012 Institut Pasteur. Published by Elsevier Masson SAS. All rights reserved.

DOI： 10.1016/j.micinf.2012.01.011

PubMed

掲載種別：研究論文（学術雑誌）  

DOI： 10.1038/embor.2012.24

PubMed

掲載種別：研究論文（学術雑誌）   国際・国内誌：国際誌  

HOIL-1L and its binding partner HOIP are essential components of the E3-ligase complex that generates linear ubiquitin (Ub) chains, which are critical regulators of NF-κB activation. Using crystallographic and mutational approaches, we characterize the unexpected structural basis for the specific interaction between the Ub-like domain (UBL) of HOIL-1L and the Ub-associated domain (UBA) of HOIP. Our data indicate the functional significance of this non-canonical mode of UBA-UBL interaction in E3 complex formation and subsequent NF-κB activation. This study highlights the versatility and specificity of protein-protein interactions involving Ub/UBLs and their cognate proteins.

DOI： 10.1038/embor.2012.24

PubMed

掲載種別：研究論文（学術雑誌）  

DOI： 10.3892/ijo.2011.1209

PubMed

掲載種別：研究論文（学術雑誌）  

NF-kappa B is involved in the metastasis of malignant cells. We have shown that NF-kappa B activation is involved in the pulmonary metastasis of LM8 cells, a highly metastatic subclone of Dunn murine osteosarcoma cells. Recently, it was determined that a newly identified type of polyubiquitin chain, a linear polyubiquitin chain, which is specifically generated by the linear ubiquitin chain assembly complex (LUBAC), plays a critical role in NF-kappa B activation. Here, we have evaluated the roles of LUBAC-mediated NF-kappa B activation in the development of lung metastasis of osteosarcoma cells. All three components of LUBAC (HOIL-1L, HOIP and SHARPIN) were highly expressed in LM8 cells compared to Dunn cells. Attenuation of LUBAC expression by stable knockdown of HOIL-1L in LM8 cells significantly suppressed NF-kappa B activity, invasiveness in vitro and lung metastasis. Induction of intracellular adhesion molecule-1 (ICAM-1) expression by LUBAC is involved in cell retention in the lungs after an intravenous inoculation of tumor cells. Moreover, we found that knockdown of LUBAC decreased not only the number but also the size of the metastatic nodules of LM8 cells in the lungs. These results indicate that LUBAC-mediated NF-kappa B activation plays crucial roles in several steps involved in metastasis, including extravasation and growth of osteosarcoma cells in the lung, and that suppression of LUBAC-mediated linear polyubiquitination activity may be a new approach to treat this life-threatening disease of young adolescents.

DOI： 10.3892/ijo.2011.1209

PubMed

掲載種別：研究論文（学術雑誌）  

DOI： 10.1126/science.1211667

PubMed

掲載種別：研究論文（学術雑誌）  

PubMed

掲載種別：研究論文（学術雑誌）  

DOI： 10.1038/nature09814

PubMed

掲載種別：研究論文（学術雑誌）  

DOI： 10.1038/nature09815

PubMed

掲載種別：研究論文（学術雑誌）  

DOI： 10.1038/nature09814

PubMed

掲載種別：研究論文（学術雑誌）  

DOI： 10.1038/nature09815

PubMed

掲載種別：研究論文（学術雑誌）  

DOI： 10.1016/j.molcel.2010.12.029

PubMed

掲載種別：研究論文（学術雑誌）  

Upon detection of viral RNA, retinoic acid-inducible gene I (RIG-I) undergoes TRIM25-mediated K63-linked ubiquitination, leading to type I interferon (IFN) production. In this study, we demonstrate that the linear ubiquitin assembly complex (LUBAC), comprised of two RING-IBR-RING (RBR)-containing E3 ligases, HOIL-1L and HOIP, independently targets TRIM25 and RIG-I to effectively suppress virus-induced IFN production. RBR E3 ligase domains of HOIL-1L and HOIP bind and induce proteasomal degradation of TRIM25, whereas the NZF domain of HOIL-1L competes with TRIM25 for RIG-I binding. Consequently, both actions by the HOIL-1L/HOIP LUBAC potently inhibit RIG-I ubiquitination and antiviral activity, but in a mechanistically separate manner. Conversely, the genetic deletion or depletion of HOIL-1L and HOIP robustly enhances virus-induced type I IFN production. Taken together, the HOIL-1L/HOIP LUBAC specifically suppresses RIG-I ubiquitination and activation by inducing TRIM25 degradation and inhibiting TRIM25 interaction with RIG-I, resulting in the comprehensive suppression of the IFN-mediated antiviral signaling pathway.

DOI： 10.1016/j.molcel.2010.12.029

PubMed

掲載種別：研究論文（学術雑誌）  

DOI： 10.1107/S1744309109050581

PubMed

掲載種別：研究論文（学術雑誌）  

F-box proteins are the substrate-recognition components of Skp1-Cullin1-F-box protein-Rbx1 (SCF) ubiquitin ligase complexes. Fbs1, an F-box protein, binds specifically to proteins modified with high-mannose oligosaccharides. Fbg3, another F-box protein, has 51% sequence identity to Fbs1. Although the residues that are necessary for binding to oligosaccharides are conserved between Fbs1 and Fbg3, Fbg3 does not bind glycoproteins. Skp1 and Fbg3 were co-expressed in Escherichia coli and their complex was purified to homogeneity and crystallized. Microseeding combined with the sandwiched hanging-drop technique improved the quality of the resulting crystals. The plate-shaped crystals belonged to space group P2(1)2(1)2(1), with unit-cell parameters a = 34.1, b = 76.6, c = 193.9 angstrom and one molecule per asymmetric unit.

DOI： 10.1107/S1744309109050581

PubMed

掲載種別：研究論文（学術雑誌）  

DOI： 10.1038/embor.2009.144

PubMed

掲載種別：研究論文（学術雑誌）  

The ubiquitin-conjugation system regulates a vast range of biological phenomena by affecting protein function mostly through polyubiquitin conjugation. The type of polyubiquitin chain that is generated seems to determine how conjugated proteins are regulated, as they are recognized specifically by proteins that contain chain-specific ubiquitin-binding motifs. An enzyme complex that catalyses the formation of newly described linear polyubiquitin chains-known as linear ubiquitin chain-assembly complex (LUBAC)-has recently been characterized, as has a particular ubiquitin-binding domain that specifically recognizes linear chains. Both have been shown to have crucial roles in the canonical nuclear factor-kappa B (NF-kappa B)-activation pathway. The ubiquitin system is intimately involved in regulating the NF-kappa B pathway, and the regulatory roles of K63-linked chains have been studied extensively. However, the role of linear chains in this process is only now emerging. This article discusses the possible mechanisms under lying linear polyubiquitin-mediated activation of NF-kappa B, and the different roles that K63-linked and linear chains have in NF-kappa B activation. Future directions for linear polyubiquitin research are also discussed.

DOI： 10.1038/embor.2009.144

PubMed

掲載種別：研究論文（学術雑誌）  

PubMed

掲載種別：研究論文（学術雑誌）  

DOI： 10.1038/ncb1821

PubMed

掲載種別：研究論文（学術雑誌）  

Nuclear factor-kappa B (NF-kappa B) is a key transcription factor in inflammatory, anti-apoptotic and immune processes. The ubiquitin pathway is crucial in regulating the NF-kappa B pathway. We have found that the LUBAC ligase complex, composed of the two RING finger proteins HOIL-1L and HOIP, conjugates a head-to-tail- linked linear polyubiquitin chain to substrates. Here, we demonstrate that LUBAC activates the canonical NF-kappa B pathway by binding to NEMO (NF-kappa B essential modulator, also called IKK gamma) and conjugates linear polyubiquitin chains onto specific Lys residues in the CC2-LZ domain of NEMO in a Ubc13-independent manner. Moreover, in HOIL-1 knockout mice and cells derived from these mice, NF-kappa B signalling induced by pro-inflammatory cytokines such as TNF-alpha and IL-1 beta was suppressed, resulting in enhanced TNF-alpha-induced apoptosis in hepatocytes of HOIL-1 knockout mice. These results indicate that LUBAC is involved in the physiological regulation of the canonical NF-kappa B activation pathway through linear polyubiquitylation of NEMO.

DOI： 10.1038/ncb1821

PubMed

掲載種別：研究論文（学術雑誌）  

DOI： 10.1073/pnas.0806215105

PubMed

掲載種別：研究論文（学術雑誌）  

DOI： 10.1074/jbc.M710599200

PubMed

掲載種別：研究論文（学術雑誌）  

DOI： 10.1074/jbc.M710599200

PubMed

掲載種別：研究論文（学術雑誌）  

DOI： 10.1091/mbc.E07-06-0601

PubMed

掲載種別：研究論文（学術雑誌）  

Misfolded or improperly assembled proteins in the endoplasmic reticulum (ER) are exported into the cytosol and degraded via the ubiquitin-proteasome pathway, a process termed ER-associated degradation (ERAD). Saccharomyces cerevisiae Hrd1p/Der3p is an ER membrane-spanning ubiquitin ligase that participates in ERAD of the cystic fibrosis transmembrane conductance regulator (CFTR) when CFTR is exogenously expressed in yeast cells. Two mammalian orthologues of yeast Hrd1p/Der3p, gp78 and HRD1, have been reported. Here, we demonstrate that gp78, but not HRD1, participates in ERAD of the CFTR mutant CFTR Delta F508, by specifically promoting ubiquitylation of CFTR Delta F508. Domain swapping experiments and deletion analysis revealed that gp78 binds to CFTR Delta F508 through its ubiquitin binding region, the so-called coupling of ubiquitin to ER degradation (CUE) domain. Gp78 polyubiquitylated in vitro an N-terminal ubiquitin-glutathione-S-transferase (GST)-fusion protein, but not GST alone. This suggests that gp78 recognizes the ubiquitin that is already conjugated to CFTR Delta F508 and catalyzes further polyubiquitylation of CFTR Delta F508 in a manner similar to that of a multiubiquitin chain assembly factor (E4). Furthermore, we revealed by small interfering RNA methods that the ubiquitin ligase RMA1 functioned as an E3 enzyme upstream of gp78. Our data demonstrates that gp78 cooperates with RMA1 with E4-like activity in the ERAD of CFTR Delta F508.

DOI： 10.1091/mbc.E07-06-0601

PubMed

掲載種別：研究論文（学術雑誌）  

DOI： 10.1128/MCB.0241-06

掲載種別：研究論文（学術雑誌）  

The transcription repressor Bach1 is a sensor and effector of heme that regulates the expression of heme oxygenase I and globin genes. Heme binds to Bach1, inhibiting its DNA binding activity and inducing its nuclear export. We found that hemin further induced the degradation of endogenous Bach1 in NIH 3T3 cells, murine embryonic fibroblasts, and murine erythroleukemia cells. In contrast, succinylacetone, an inhibitor of heme synthesis, caused accumulation of Bach1 in murine embryonic fibroblasts, indicating that physiological levels of heme regulated the Bach1 turnover. Polyubiquitination and rapid degradation of overexpressed Bach1 were induced by hemin treatment. HOIL-1, an ubiquitin-protein ligase which recognizes heme-bound, oxidized iron regulatory protein 2, was found to bind with Bach1 when both were overexpressed in NIH 3T3 cells. HOIL-1 stimulated the polyubiquitination of Bach1 in a purified in vitro ubiquitination system depending on the intact heme binding motifs of Bach1. Expression of dominant-negative HOIL-1 in murine erythroleukemia cells resulted in higher stability of endogenous Bach1, raising the possibility that the heme-regulated degradation involved HOIL-1 in murine erythroleukemia cells. These results suggest that heme within a cell regulates the polyubiquitination and degradation of Bach1.

DOI： 10.1128/MCB.0241-06

PubMed

掲載種別：研究論文（学術雑誌）  

DOI： 10.1016/j.bbrc.2006.09.163

PubMed

掲載種別：研究論文（学術雑誌）  

DOI： 10.1016/j.bbrc.2006.09.163

PubMed

掲載種別：研究論文（学術雑誌）  

DOI： 10.1038/sj.emboj.7601360

PubMed

掲載種別：研究論文（学術雑誌）  

DOI： 10.1038/sj.emboj.7601360

PubMed

掲載種別：研究論文（学術雑誌）   共著区分：単著  

PubMed

掲載種別：研究論文（学術雑誌）  

PubMed

掲載種別：研究論文（学術雑誌）  

DOI： 10.1016/j.thromres.2005.02.017

PubMed

掲載種別：研究論文（学術雑誌）  

Introduction: Misfolded and unassembled glycoproteins are eliminated from the endoplasmic reticulum (ER) lumen by the ER-associated degradation (ERAD). We previously identified a Tyr595Cys (Y595C) mutation of protein S (PS) in a family of a quantitative PS deficiency. The mutation causes intracettular degradation and decreased secretion of the Y595C mutant PS. The aim of the present study was to further characterize the molecular basis of the intracellular degradation of the mutant.
Materials and methods: We stably Expressed the mutant in mammalian cells, and analyzed the intracellular localization of the protein. The intracellular degradation pathway was determined by pulse-chase analyses in the presence of various inhibitors of ERAD.
Results and conclusions: Endoglycosidase H digestion and immunofluorescence staining reveated the mutant being retained in the ER. Epoxomicin, a potent and specific proteasome inhibitor, and Ala-Ala-Phe-CH(2)Cl (AAF), an inhibitor of tripeptidyl peptidase II (TPPII), suppressed the intracellular degradation of the mutant by about 65% and 50%, respectively. When epoxomicin was combined with AAF, the inhibitory effect was substantially enhanced. Although castanospermine, an inhibitor of glucosidases I and II, did not affect the degradation, kifunensine, an inhibitor of ER mannosidase 1, suppressed it. Thus, it appears that the Y595C mutant is degraded through more than one pathway of ERAD, including the proteasome-dependent pathway and an alternate proteasome-independent pathway where proteases such as TPPII may be involved. Production of the critical B isoform of Man(8)GlcNAC(2) targets the mutant for ERAD, however, the interaction with calnexin/ catreticulin through monoglucosytated oLigosaccharides may not be required for the degradation of the mutant. (c) 2005 Etsevier Ltd. All rights reserved.

DOI： 10.1016/j.thromres.2005.02.017

PubMed

掲載種別：研究論文（学術雑誌）  

DOI： 10.1016/j.molcel.2005.05.027

PubMed

掲載種別：研究論文（学術雑誌）  

DOI： 10.1016/j.molcel.2005.05.027

PubMed

掲載種別：研究論文（学術雑誌）  

掲載種別：研究論文（学術雑誌）  

掲載種別：研究論文（学術雑誌）  

掲載種別：研究論文（学術雑誌）  

掲載種別：研究論文（学術雑誌）   共著区分：単著  

PubMed

掲載種別：研究論文（学術雑誌）  

PubMed

掲載種別：研究論文（学術雑誌）  

掲載種別：研究論文（学術雑誌）  

掲載種別：研究論文（学術雑誌）  

DOI： 10.1074/jbc.M304157200

PubMed

掲載種別：研究論文（学術雑誌）  

F-box proteins are substrate recognition components of Skp1-Cullin1-F-box protein-Roc1 (SCF) E3 ubiquitin-protein ligases. We reported previously that Fbs1 (F-box protein that recognizes sugar chains; equivalent to Fbx2 or NFB42) binds specifically to proteins attached with high mannose oligosaccharides and subsequently contributes to elimination of N-glycoproteins in cytosol (Yoshida, Y., Chiba, T., Tokunaga, F., Kawasaki, H., Iwai, K., Suzuki, T., Ito, Y., Matsuoka, K., Yoshida, M., Tanaka, K., and Tai, T. (2002) Nature 418, 438 - 442). Here we report the identification of another F-box protein that recognizes N-glycan, Fbs2 (called Fbx6b or FBG2 previously). Although the expression of Fbs1 was restricted to the adult brain and testis, the Fbs2 transcript was widely expressed. The Fbs2 protein forms an SCFFbs2 ubiquitin-ligase complex that targets sugar chains in N-glycoproteins for ubiquitylation. Only glycoproteins bound to concanavalin A lectin and not to wheat germ agglutinin or Ricinus communis agglutinin interacted with Fbs2 in various tissues and cell lines. Pull-down analysis using various oligosaccharides revealed that Man(3-9)GlcNAc(2) glycans were required for efficient Fbs2 binding, whereas modifications of mannose residues by other sugars or deletion of inner GlcNAc reduced Fbs2 binding. Fbs2 interacted with N-glycans of T-cell receptor alpha-subunit (TCRalpha), a typical substrate of the endoplasmic reticulum-associated degradation (ERAD) pathway, and the forced expression of mutant Fbs2DeltaF, which lacks the F-box domain essential for forming the SCF complex, and decrease of endogenous Fbs2 by small interfering RNA led to inhibition of TCRalpha degradation in cells. Thus, Fbs2 is a novel member of F-box protein family that recognizes N-glycans and plays a role in ERAD.

DOI： 10.1074/jbc.M304157200

PubMed

掲載種別：研究論文（学術雑誌）  

DOI： 10.1038/ncb952

PubMed

掲載種別：研究論文（学術雑誌）  

The ubiquitin system is involved in several basic cellular functions(1-3). Ubiquitination is carried out by a cascade of three reactions catalysed by the El, E2 and E3 enzymes. Among these, the E3 ubiquitin-protein ligases have a pivotal role in determining the specificity of the system by recognizing the target substrates through defined targeting motifs(1-3). Although RING finger proteins constitute an important family of E3 ligases(4), only a few post-transcriptional modifications, including phosphorylation(1), proline hydroxylation(5,6) and glycosylation(7), are known to function as recognition signals for E3. Iron regulatory protein 2 (IRP2), a modulator of iron metabolism, is regulated by iron-induced ubiquitination and degradation(8). Here we show that the RING finger protein HOIL-1 functions as an E3 ligase for oxidized IRP2, suggesting that oxidation is a specific recognition signal for ubiquitination. The oxidation of IRP2 is generated by haem, which binds to IRP2 in iron-rich cells, and by oxygen, indicating that the iron sensing of IRP2 depends on the synthesis and availability of haem.

DOI： 10.1038/ncb952

PubMed

掲載種別：研究論文（学術雑誌）  

DOI： 10.1016/s0003-9861(02)00717-8

PubMed

掲載種別：研究論文（学術雑誌）  

Previously we showed that two antithrombin mutants were degraded through an endoplasmic reticulum (ER)-associated degradation (ERAD) pathway [F. Tokunaga et al., FEBS Lett. 412 (1997) 65]. Here, we examined the combined effects of inhibitors of glycosidases, protein synthesis, proteasome, and tyrosine phosphatase on ERAD of a Glu313-deleted (DeltaGlu) mutant of antithrombin. We found that kifunensine, an ER marmosidase I inhibitor, suppressed ERAD, indicating that specific mannose trimming plays a critical role. Cycloheximide and puromycin, inhibitors of protein synthesis, also suppressed ERAD, the effects being cancelled by pretreatment with castanospermine. In contrast, kifunensine suppressed ERAD even in castanospermine-treated cells, suggesting that suppression of ERAD does not always require the binding of lectin-like ER chaperones-like calnexin and/or calreticulin. These results indicate that, besides proteasome inhibitors, inhibitors of ER mannosidase I and protein synthesis suppress ERAD of the antithrombin DeltaGlu mutant at different stages, and processing of N-linked oligosaccharides highly correlated with the efficiency of ERAD. (C) 2003 Elsevier Science (USA). All rights reserved.

DOI： 10.1016/S0003-9861(02)00717-8

PubMed

掲載種別：研究論文（学術雑誌）  

掲載種別：研究論文（学術雑誌）  

掲載種別：研究論文（学術雑誌）  

掲載種別：研究論文（学術雑誌）  

PubMed

掲載種別：研究論文（学術雑誌）  

PubMed

掲載種別：研究論文（学術雑誌）  

DOI： 10.1074/jbc.M206773200

PubMed

掲載種別：研究論文（学術雑誌）  

Monoclonal antibodies were raised against hemocytes of the horseshoe crab Tachypleus tridentatus. All of the antibodies obtained reacted with the same protein bands on SDS-PAGE of hemocyte lysate. Flow cytometry and biotinylation of surface substances on the hemocytes indicated that the antigens are major peripheral proteins of hemocytes. The antigens were purified from hemocyte lysate and were good substrates for the horseshoe crab hemocyte transglutaminase (HcTGase). Transglutaminases play an important role during the final stage of blood coagulation in mammals and crustaceans. Although HcTGase did not intermolecularly cross-link a clottable protein coagulogen or its proteolytic product coagulin, HcTGase promoted the cross-linking of coagulin with the surface antigens, resulting in the formation of a stable polymer. We determined the nucleotide sequences for two isoproteins of the antigens. The two proteins containing 271 and 284 residues (66% identity) were composed of tandem repeats of proline-rich segments. We named them proxins-1 and -2 after proline-rich proteins for protein cross-linking. Proxins may form a stable physical barrier against invading pathogens in cooperation with hemolymph coagulation at injured sites.

DOI： 10.1074/M206773200

PubMed

掲載種別：研究論文（学術雑誌）  

DOI： 10.1038/nature00890

PubMed

掲載種別：研究論文（学術雑誌）  

N-glycosylation of proteins in the endoplasmic reticulum (ER) has a central role in protein quality control(1-3). Here we report that N-glycan serves as a signal for degradation by the Skp1-Cullin1-Fbx2-Roc1 (SCF(Fbx2)) ubiquitin ligase complex. The F-box protein Fbx2 (ref. 4) binds specifically to proteins attached to N-linked high-mannose oligosaccharides and subsequently contributes to ubiquitination of N-glycosylated proteins. Pre-integrin beta1 is a target of Fbx2; these two proteins interact in the cytosol after inhibition of the proteasome. In addition, expression of the mutant Fbx2DeltaF, which lacks the F-box domain that is essential for forming the SCF complex, appreciably blocks degradation of typical substrates of the ER-associated degradation pathway(5,6). Our results indicate that SCF(Fbx2) ubiquitinates N-glycosylated proteins that are translocated from the ER to the cytosol by the quality control mechanism.

DOI： 10.1038/nature00890

PubMed

掲載種別：研究論文（学術雑誌）  

To examine the role of early carbohydrate recognition/trimming reactions in targeting endoplasmic reticulum (ER)-retained, misfolded glycoproteins for ER-associated degradation (ERAD), we have stably expressed the cog thyroglobulin (Tg) mutant cDNA in Chinese hamster ovary cells. We found that inhibitors of ER mannosidase I (but not other glycosidases) acutely suppressed Cog Tg degradation and also perturbed the ERAD process for Tg reduced with dithiothreitol as well as for gamma -carboxylation-deficient protein C expressed in warfarin-treated baby hamster kidney cells. Kifunensine inhibition of ER mannosidase I also suppressed ERAD in castanospermine-treated cells; thus, suppression of ERAD does not require lectin-like binding of ER chaperones calnexin and calreticulin to monoglucosylated oligosaccharides. Notably, the undegraded protein fraction remained completely microsome-associated. In pulse-chase studies, kifunensine-sensitive degradation was still inhibitable even 1 h after Tg synthesis. Intriguingly, chronic treatment with kifunensine caused a 3-fold accumulation of Cog Tg in Chinese hamster ovary cells and did not lead to significant induction of the ER unfolded protein response. We hypothesize that, in a manner not requiring lectin-like activity of calnexin/ calreticulin, the recognition or processing of a specific branched N-linked mannose structure enhances the efficiency of glycoprotein retrotranslocation from the ER lumen.

掲載種別：研究論文（学術雑誌）  

To examine the role of early carbohydrate recognition/trimming reactions in targeting endoplasmic reticulum (ER)-retained, misfolded glycoproteins for ER-associated degradation (ERAD), we have stably expressed the cog thyroglobulin (Tg) mutant cDNA in Chinese hamster ovary cells. We found that inhibitors of ER mannosidase I (but not other glycosidases) acutely suppressed Cog Tg degradation and also perturbed the ERAD process for Tg reduced with dithiothreitol as well as for gamma -carboxylation-deficient protein C expressed in warfarin-treated baby hamster kidney cells. Kifunensine inhibition of ER mannosidase I also suppressed ERAD in castanospermine-treated cells; thus, suppression of ERAD does not require lectin-like binding of ER chaperones calnexin and calreticulin to monoglucosylated oligosaccharides. Notably, the undegraded protein fraction remained completely microsome-associated. In pulse-chase studies, kifunensine-sensitive degradation was still inhibitable even 1 h after Tg synthesis. Intriguingly, chronic treatment with kifunensine caused a 3-fold accumulation of Cog Tg in Chinese hamster ovary cells and did not lead to significant induction of the ER unfolded protein response. We hypothesize that, in a manner not requiring lectin-like activity of calnexin/ calreticulin, the recognition or processing of a specific branched N-linked mannose structure enhances the efficiency of glycoprotein retrotranslocation from the ER lumen.

掲載種別：研究論文（学術雑誌）  

掲載種別：研究論文（学術雑誌）  

A glycoprotein (M(r)=43000) from horseshoe crab hemocytes with antimicrobial activity against Gram-negative bacteria was purified, The internal peptide sequences coincided exactly with the deduced amino acid sequence of a cDNA clone, designated limulus factor D, which was isolated by screening a hemocyte cDNA library with an anti-human plasminogen antibody, The open reading frame codes for a precursor of factor D of 394 amino acid residues, including an NH2-terminal signal sequence. The COOH-terminal domain of factor D has significant sequence homology with the catalytic domain of mammalian serine proteases, in particular with human tissue plasminogen activator (32% identity), except for the substitution of Ser of the active site tried to Gly. Factor D has a unique NH2-terminal domain with weak sequence homology with part of the mammalian interleukin-6 receptor alpha-chain. Factor D is likely to have an important role in host defense mechanisms.

DOI： 10.1016/S0014-5793(96)01224-0

PubMed

掲載種別：研究論文（学術雑誌）  

A glycoprotein (M(r)=43000) from horseshoe crab hemocytes with antimicrobial activity against Gram-negative bacteria was purified, The internal peptide sequences coincided exactly with the deduced amino acid sequence of a cDNA clone, designated limulus factor D, which was isolated by screening a hemocyte cDNA library with an anti-human plasminogen antibody, The open reading frame codes for a precursor of factor D of 394 amino acid residues, including an NH2-terminal signal sequence. The COOH-terminal domain of factor D has significant sequence homology with the catalytic domain of mammalian serine proteases, in particular with human tissue plasminogen activator (32% identity), except for the substitution of Ser of the active site tried to Gly. Factor D has a unique NH2-terminal domain with weak sequence homology with part of the mammalian interleukin-6 receptor alpha-chain. Factor D is likely to have an important role in host defense mechanisms.

DOI： 10.1016/S0014-5793(96)01224-0

PubMed

掲載種別：研究論文（学術雑誌）  

掲載種別：研究論文（学術雑誌）  

掲載種別：研究論文（学術雑誌）  

Heparin cofactor II (HCII) is a serine proteinase inhibitor in human plasma that rapidly inhibits thrombin in the presence of dermatan sulfate or heparin. To understand the molecular mechanism for HCII deficiency in a patient with reduced circulating HCII antigen, we studied a Japanese patient with type I HCII deficiency who suffered from angina pectoris and coronary artery disease, Polymerase chain reaction (PCR)-based sequence analysis showed that the propositus' gene for HCII (HCII Awaji gene) had a thymine insertion after codon (GAT) for Asp88 in exon II, resulting in a frameshift mutation. Consequently, the abnormal HCII Awaji protein was suggested to have an altered amino acid sequence from position 89 and terminate at 107, thus being composed of the NH2-terminal one fifth of normal HCII and dysfunctional for thrombin inhibition, The molecular weight and pI value of HCII Awaji were calculated to be 12,040 and 3.6, respectively, without posttranslational modification. Mutagenic PCR followed by the Tsp5091 digestion showed that a half of the PCR products derived from the propositus and his sister was cleaved, suggesting that his sister also has the same mutant allele, Crossed-immunoelectrophoresis and Western blot analyses of plasma and urine from the propositus and of plasma from his sister did not provide evidence for the existence of the abnormal HCII, suggesting that little truncated HCII was circulating in the patient's blood. However, stable expression assay using human kidney 293 cells transfected with the expression vector containing cDNA encoding wild-type or Awaji-type HCII showed that mutant as well as wildtype HCII was secreted into culture medium normally, These results suggest that the abnormal HCII Awaji protein is secreted normally, but rapidly degraded in the circulating blood. (C) 1996 by The American Society of Hematology.

掲載種別：研究論文（学術雑誌）  

Heparin cofactor II (HCII) is a serine proteinase inhibitor in human plasma that rapidly inhibits thrombin in the presence of dermatan sulfate or heparin. To understand the molecular mechanism for HCII deficiency in a patient with reduced circulating HCII antigen, we studied a Japanese patient with type I HCII deficiency who suffered from angina pectoris and coronary artery disease, Polymerase chain reaction (PCR)-based sequence analysis showed that the propositus' gene for HCII (HCII Awaji gene) had a thymine insertion after codon (GAT) for Asp88 in exon II, resulting in a frameshift mutation. Consequently, the abnormal HCII Awaji protein was suggested to have an altered amino acid sequence from position 89 and terminate at 107, thus being composed of the NH2-terminal one fifth of normal HCII and dysfunctional for thrombin inhibition, The molecular weight and pI value of HCII Awaji were calculated to be 12,040 and 3.6, respectively, without posttranslational modification. Mutagenic PCR followed by the Tsp5091 digestion showed that a half of the PCR products derived from the propositus and his sister was cleaved, suggesting that his sister also has the same mutant allele, Crossed-immunoelectrophoresis and Western blot analyses of plasma and urine from the propositus and of plasma from his sister did not provide evidence for the existence of the abnormal HCII, suggesting that little truncated HCII was circulating in the patient's blood. However, stable expression assay using human kidney 293 cells transfected with the expression vector containing cDNA encoding wild-type or Awaji-type HCII showed that mutant as well as wildtype HCII was secreted into culture medium normally, These results suggest that the abnormal HCII Awaji protein is secreted normally, but rapidly degraded in the circulating blood. (C) 1996 by The American Society of Hematology.

掲載種別：研究論文（学術雑誌）  

掲載種別：研究論文（学術雑誌）  

Warfarin is known to disrupt the microsomal vitamin K cycle, which results in a decrease of the plasma level of protein C, an anticoagulant factor, as well as some other vitamin K-dependent coagulation factors. We examined the effect of warfarin on secretion of recombinant protein C expressed in human kidney 293 or BHK cells. In transient expression, warfarin caused a two- to four-fold decrease in the quantity of protein C secreted, compared to findings with vitamin K-treated cells. Pulse-chase experiments using stable cells showed that, although recombinant protein C was secreted in the presence of vitamin K, the decrease in total amount of the radioactivity in the warfarin-treated cells suggested intracellular degradation. This degradation depended on the concentration of warfarin and was not inhibited by an endoplasmic reticulum (ER)-Golgi transport inhibitor or by lysosomotropic inhibitors. Thus, protein C synthesized in the presence of warfarin is selectively degraded and the degradation occurs in a pre-Golgi, nonlysosomal compartment. Among the protease inhibitors tested, N-acetyl-Leu-Leu-methioninal and N-acetyl-Leu-Leu-norleucinal blocked the degradation of protein C synthesized in the presence of warfarin and the protein C accumulated intracellularly, in a dose-dependent manner. Both inhibitors, however, did not disturb the secretion of protein C in the vitamin K-treated cells. Thus, a cysteine protease(s) appeared to be responsible for the degradation. These results suggest that protein C synthesized in the presence of warfarin was selectively degraded by a cysteine protease(s) in the ER through a 'quality control' mechanism.

PubMed

掲載種別：研究論文（国際会議プロシーディングス）  

掲載種別：研究論文（国際会議プロシーディングス）  

Warfarin is known to disrupt the microsomal vitamin K cycle, which results in a decrease of the plasma level of protein C, an anticoagulant factor, as well as some other vitamin K-dependent coagulation factors. We examined the effect of warfarin on secretion of recombinant protein C expressed in human kidney 293 or BHK cells. In transient expression, warfarin caused a two- to four-fold decrease in the quantity of protein C secreted, compared to findings with vitamin K-treated cells. Pulse-chase experiments using stable cells showed that, although recombinant protein C was secreted in the presence of vitamin K, the decrease in total amount of the radioactivity in the warfarin-treated cells suggested intracellular degradation. This degradation depended on the concentration of warfarin and was not inhibited by an endoplasmic reticulum (ER)-Golgi transport inhibitor or by lysosomotropic inhibitors. Thus, protein C synthesized in the presence of warfarin is selectively degraded and the degradation occurs in a pre-Golgi, nonlysosomal compartment. Among the protease inhibitors tested, N-acetyl-Leu-Leu-methioninal and N-acetyl-Leu-Leu-norleucinal blocked the degradation of protein C synthesized in the presence of warfarin and the protein C accumulated intracellularly, in a dose-dependent manner. Both inhibitors, however, did not disturb the secretion of protein C in the vitamin K-treated cells. Thus, a cysteine protease(s) appeared to be responsible for the degradation. These results suggest that protein C synthesized in the presence of warfarin was selectively degraded by a cysteine protease(s) in the ER through a 'quality control' mechanism.

PubMed

掲載種別：研究論文（学術雑誌）  

A new subunit, named RC6-I, of the rat 20 S proteasome was purified and the partial amino acid sequences of several peptide fragments obtained by digestion with lysyl-endopeptidase were determined by Edman degradation. Amplification of cDNAs encoding RC6-I by the polymerase chain reaction (PCR) technique revealed two types of cDNA, tentatively designated as RC6-IL and RC6-IS in order of size. The nucleotide sequences of the two cDNAs are identical except that RC6-IL contains an insertion of 18 nucleotides in the coding region compared with RC6-IS. The polypeptide predicted from the open reading frame of RC6-IS cDNA consists of 248 amino acid residues with a calculated molecular weight of 27783. These values are consistent with those obtained by protein chemical analyses. Computer-assisted homology analysis showed that RC6-I belongs to the α-type subfamily of the proteasome gene family, which shows similarity to the α-subunit of the archaebacterium Thermoplasma acidophilum proteasome, and that the 18 nucleotide insert, encoding six amino acid residues, VVASVS, appears to be unique to RC6-IL, because this motif has not been conserved in any other α-type subunit. By reverse transcription (RT)-PCR analysis, the mRNAs for both RC6-IL and RC6-IS were found in all the rat tissues examined. These results suggest that proteasomes are present as a heterogeneous population, possibly for acquisition of diversity of functions. © 1995.

DOI： 10.1016/0167-4781(95)00113-U

PubMed

掲載種別：研究論文（学術雑誌）  

A new subunit, named RC6-I, of the rat 20 S proteasome was purified and the partial amino acid sequences of several peptide fragments obtained by digestion with lysyl-endopeptidase were determined by Edman degradation. Amplification of cDNAs encoding RC6-I by the polymerase chain reaction (PCR) technique revealed two types of cDNA, tentatively designated as RC6-IL and RC6-IS in order of size. The nucleotide sequences of the two cDNAs are identical except that RC6-IL contains an insertion of 18 nucleotides in the coding region compared with RC6-IS. The polypeptide predicted from the open reading frame of RC6-IS cDNA consists of 248 amino acid residues with a calculated molecular weight of 27783. These values are consistent with those obtained by protein chemical analyses. Computer-assisted homology analysis showed that RC6-I belongs to the α-type subfamily of the proteasome gene family, which shows similarity to the α-subunit of the archaebacterium Thermoplasma acidophilum proteasome, and that the 18 nucleotide insert, encoding six amino acid residues, VVASVS, appears to be unique to RC6-IL, because this motif has not been conserved in any other α-type subunit. By reverse transcription (RT)-PCR analysis, the mRNAs for both RC6-IL and RC6-IS were found in all the rat tissues examined. These results suggest that proteasomes are present as a heterogeneous population, possibly for acquisition of diversity of functions. © 1995.

DOI： 10.1016/0167-4781(95)00113-U

PubMed

掲載種別：研究論文（学術雑誌）  

掲載種別：研究論文（学術雑誌）  

掲載種別：研究論文（学術雑誌）  

掲載種別：研究論文（学術雑誌）  

The nucleotide sequence of a cDNA that encodes anew subunit, named RC10-II, of the 20S proteasome of rat embryonic brain has been determined. The polypeptide predicted from the open reading frame consists of 205 amino acid residues with a calculated molecular weight of 22,965 and isoelectric point of 6.15. Computer analysis showed that RC10-II belongs to the β-type subgroup of proteasomes, differing clearly from α-type subunits of the proteasome gene family. The primary structure of RC10-II was found to contain the partial amino acid sequences of several fragments of subunit 13, which has a cysteinyl residue critical for the trypsin-like catalytic activity, as reported by L.R. Dick et al. [Biochemistry 31, 7347-7355, 1992], suggesting that RC10-II is a proteasomal subunit necessary for the expression of tryptic activity. © 1993.

DOI： 10.1016/0014-5793(93)80856-P

PubMed

掲載種別：研究論文（学術雑誌）  

The nucleotide sequence of a cDNA that encodes anew subunit, named RC10-II, of the 20S proteasome of rat embryonic brain has been determined. The polypeptide predicted from the open reading frame consists of 205 amino acid residues with a calculated molecular weight of 22,965 and isoelectric point of 6.15. Computer analysis showed that RC10-II belongs to the β-type subgroup of proteasomes, differing clearly from α-type subunits of the proteasome gene family. The primary structure of RC10-II was found to contain the partial amino acid sequences of several fragments of subunit 13, which has a cysteinyl residue critical for the trypsin-like catalytic activity, as reported by L.R. Dick et al. [Biochemistry 31, 7347-7355, 1992], suggesting that RC10-II is a proteasomal subunit necessary for the expression of tryptic activity. © 1993.

DOI： 10.1016/0014-5793(93)80856-P

PubMed

掲載種別：研究論文（学術雑誌）  

The nucleotide sequence of a cDNA that encodes a new subunit, named RC7-I, of the 20 S proteasome of rat hepatoma cells has been determined. The polypeptide predicted from the open reading frame consists of 201 amino acid residues with a calculated molecular weight of 22,912 and isoelectric point of 7.25. Approximately 80% of the partial amino acid sequences of several fragments of RC7-I, determined by protein chemical analyses, were found to be in excellent accordance with those deduced from the cDNA sequence. Computer analysis showed that RC7-I belongs to the beta-type subgroup of proteasomes with similarity to the beta-subunit of the archaebacterial proteasome, differing clearly from alpha-type subunits of the proteasome gene family. The overall structure of RC7-I was found to be homologous to that of yeast PRE1, which is necessary for chymotryptic activity.

掲載種別：研究論文（学術雑誌）  

The nucleotide sequence of a cDNA that encodes a new subunit, named RC7-I, of the 20 S proteasome of rat hepatoma cells has been determined. The polypeptide predicted from the open reading frame consists of 201 amino acid residues with a calculated molecular weight of 22,912 and isoelectric point of 7.25. Approximately 80% of the partial amino acid sequences of several fragments of RC7-I, determined by protein chemical analyses, were found to be in excellent accordance with those deduced from the cDNA sequence. Computer analysis showed that RC7-I belongs to the beta-type subgroup of proteasomes with similarity to the beta-subunit of the archaebacterial proteasome, differing clearly from alpha-type subunits of the proteasome gene family. The overall structure of RC7-I was found to be homologous to that of yeast PRE1, which is necessary for chymotryptic activity.

掲載種別：研究論文（学術雑誌）  

掲載種別：研究論文（学術雑誌）  

The molecular properties of an ATP/ubiquitin-dependent ‗26S‘ proteasome complex purified from rat liver were examined by physicochemical, biochemical, and morphological analyses. On ultracentrifugation, the proteasome complex sedimented as almost a single component with a sedimentation coefficient of 30.3S. Dynamic light-scattering measurements indicated that it has a diffusion coefficient of 1.38 x 10-7 cm2/sec and a Stokes radius of 15.5 nm. From these two coefficients, the protein complex was estimated to have the high molecular weight of 2.02 x 106. Static light-scattering analysis indicated a molecular weight of 1.91 x 106 and a radius of gyration of 16.8 nm. The proteasome complex was found to be composed of multiple subunits of the 20S proteasome with molecular weights of 2.1-3.1 x 104 and 15-20 protein species with molecular weights of 3.5-11.0 x 104, which were directly associated with the 20S proteasome. The electron micrographic finding that the 26S proteasome complex had a caterpillar shape, direct electron-microscopic observations on the subunit arrangement of the 20S proteasome, and classification of the subunits of the latter into two groups with respect to sequence homology suggested that the 26S complex is a symmetrical assembly of two domains, each containing a large terminal subset and half the central 20S subset of components. For clarification of the molecular structure of the 26S proteasome complex in solution, its physicochemical parameters were calculated theoretically using a model based on this caterpillar-shaped complex. The values obtained for the Stokes radius and radius of gyration of 12.2 and 14.9 nm were consistent with the experimental values. These results provide evidence that the 26S proteasome complex is a cylindrical caterpillar-like structure of ‗30S‘ in solution, consisting of a 20S proteasome component with proteolytic function and multiple other components, which possibly have regulatory roles. © 1994 Academic Press. All rights reserved.

DOI： 10.1006/jsbi.1993.1050

掲載種別：研究論文（国際会議プロシーディングス）  

掲載種別：研究論文（学術雑誌）  

The molecular properties of an ATP/ubiquitin-dependent ‗26S‘ proteasome complex purified from rat liver were examined by physicochemical, biochemical, and morphological analyses. On ultracentrifugation, the proteasome complex sedimented as almost a single component with a sedimentation coefficient of 30.3S. Dynamic light-scattering measurements indicated that it has a diffusion coefficient of 1.38 x 10-7 cm2/sec and a Stokes radius of 15.5 nm. From these two coefficients, the protein complex was estimated to have the high molecular weight of 2.02 x 106. Static light-scattering analysis indicated a molecular weight of 1.91 x 106 and a radius of gyration of 16.8 nm. The proteasome complex was found to be composed of multiple subunits of the 20S proteasome with molecular weights of 2.1-3.1 x 104 and 15-20 protein species with molecular weights of 3.5-11.0 x 104, which were directly associated with the 20S proteasome. The electron micrographic finding that the 26S proteasome complex had a caterpillar shape, direct electron-microscopic observations on the subunit arrangement of the 20S proteasome, and classification of the subunits of the latter into two groups with respect to sequence homology suggested that the 26S complex is a symmetrical assembly of two domains, each containing a large terminal subset and half the central 20S subset of components. For clarification of the molecular structure of the 26S proteasome complex in solution, its physicochemical parameters were calculated theoretically using a model based on this caterpillar-shaped complex. The values obtained for the Stokes radius and radius of gyration of 12.2 and 14.9 nm were consistent with the experimental values. These results provide evidence that the 26S proteasome complex is a cylindrical caterpillar-like structure of ‗30S‘ in solution, consisting of a 20S proteasome component with proteolytic function and multiple other components, which possibly have regulatory roles. © 1994 Academic Press. All rights reserved.

DOI： 10.1006/jsbi.1993.1050

掲載種別：研究論文（学術雑誌）  

The nucleotide sequence of a cDNA that encodes a new subunit, named RC1, of rat proteasomes (multicatalytic proteinase complexes) has been determined. The polypeptide predicted from the open reading frame consisted of 208 amino acid residues with a calculated molecular mass of 23,130, which is consistent with the size obtained by electrophoretic analysis of purified RC1. The partial amino acid sequences of several fragments of RC1, obtained by protein chemical analyses, were found to be in excellent accordance with those deduced from the cDNA sequence. Surprisingly, the overall structure of RC1 was found to be almost identical to that of recently isolated RING10, whose gene is located in the class II region of the human MHC gene cluster. This finding suggests that RC1 is a homologue of human RING10, supporting the proposal that proteasomes are involved in the antigen processing pathway. © 1992.

DOI： 10.1016/0014-5793(92)80211-X

PubMed

掲載種別：研究論文（学術雑誌）  

The nucleotide sequence of a cDNA that encodes a new subunit, named RC1, of rat proteasomes (multicatalytic proteinase complexes) has been determined. The polypeptide predicted from the open reading frame consisted of 208 amino acid residues with a calculated molecular mass of 23,130, which is consistent with the size obtained by electrophoretic analysis of purified RC1. The partial amino acid sequences of several fragments of RC1, obtained by protein chemical analyses, were found to be in excellent accordance with those deduced from the cDNA sequence. Surprisingly, the overall structure of RC1 was found to be almost identical to that of recently isolated RING10, whose gene is located in the class II region of the human MHC gene cluster. This finding suggests that RC1 is a homologue of human RING10, supporting the proposal that proteasomes are involved in the antigen processing pathway. © 1992.

DOI： 10.1016/0014-5793(92)80211-X

PubMed

掲載種別：研究論文（学術雑誌）  

Seventeen murine monoclonal antibodies (mAbs) against horseshoe crab clotting factor, factor C, were prepared and characterized. When the binding sites of these mAbs were analyzed by immunoblotting, ten mAbs recognized nonreduced factor C, five mAbs were directed against the heavy chain, and two mAbs were directed against the B chain. Three mAbs, 1H4, 2C12, and 2A7, one selected from each group, were used for further study. The mAb 1H4, which recognized only nonreduced factor C molecule, inhibited the factor C activity in a dose-dependent manner. It also inhibited lipopolysaccharide (LPS)- and α-chymotrypsln-mediated activations of the zymogen factor C, suggesting that 1H4 binds close to the active site and/or the substrate-binding site located in the serine protease domain (B chain) of factor C. On the other hand, 2C12 and 2A7 recognized, respectively, an epitope located in the heavy and the B chains, and inhibited LPS-mediated activation of factor C, but not α-chymotrypsin-mediated activation of factor C or factor C¯ activity. Both F(ab')2 and Fab' fragments derived from 2C12 inhibited LPS-mediated activation in the same manner. These three mAbs did not bind with LPS, although a factor C-mAb complex was able to bind LPS, suggesting that the LPS-mediated activation of the zymogen factor C was induced through Intermolecular interaction between the LPS-bound factor C mole cules. The dissociation constants (Kd) for 1H4, 2C12, and 2A7 binding to factor C were determined as 1.9×10-90.6×10-10and 1.8×10 respectively. By using 2C12 and polyclonal antibody against factor C¯, an enzyme-linked immunosorbent assay for quantitative determination of factor C was established. © 1992 BY THE JOURNAL OF BIOCHEMISTRY.

DOI： 10.1093/oxfordjournals.jbchem.a123924

PubMed

掲載種別：研究論文（学術雑誌）  

Seventeen murine monoclonal antibodies (mAbs) against horseshoe crab clotting factor, factor C, were prepared and characterized. When the binding sites of these mAbs were analyzed by immunoblotting, ten mAbs recognized nonreduced factor C, five mAbs were directed against the heavy chain, and two mAbs were directed against the B chain. Three mAbs, 1H4, 2C12, and 2A7, one selected from each group, were used for further study. The mAb 1H4, which recognized only nonreduced factor C molecule, inhibited the factor C activity in a dose-dependent manner. It also inhibited lipopolysaccharide (LPS)- and α-chymotrypsln-mediated activations of the zymogen factor C, suggesting that 1H4 binds close to the active site and/or the substrate-binding site located in the serine protease domain (B chain) of factor C. On the other hand, 2C12 and 2A7 recognized, respectively, an epitope located in the heavy and the B chains, and inhibited LPS-mediated activation of factor C, but not α-chymotrypsin-mediated activation of factor C or factor C¯ activity. Both F(ab')2 and Fab' fragments derived from 2C12 inhibited LPS-mediated activation in the same manner. These three mAbs did not bind with LPS, although a factor C-mAb complex was able to bind LPS, suggesting that the LPS-mediated activation of the zymogen factor C was induced through Intermolecular interaction between the LPS-bound factor C mole cules. The dissociation constants (Kd) for 1H4, 2C12, and 2A7 binding to factor C were determined as 1.9×10-90.6×10-10and 1.8×10 respectively. By using 2C12 and polyclonal antibody against factor C¯, an enzyme-linked immunosorbent assay for quantitative determination of factor C was established. © 1992 BY THE JOURNAL OF BIOCHEMISTRY.

DOI： 10.1093/oxfordjournals.jbchem.a123924

PubMed

CiNii Article

掲載種別：研究論文（学術雑誌）  

The structure of hemocytes in the normal state and during blood coagulation, and the intracellular localization of three clotting factors and two antimicrobial factors were examined in the Japanese horseshoe crab Tachypleus tridentatus. Two types of hemocytes were found in the circulating blood: non-granular and granular hemocytes. The latter contained numerous dense granules classed into two major types: L- and D-granules. The L-granules were larger (up to 1.5-mu-m in diameter) and less electron-dense than the D-granules (less than 0.6-mu-m in diameter). The L-granules contained three clotting factors and one antimicrobial factor, whereas the D-granules exclusively contained the other antimicrobial factor. After treatment with endotoxin, the L-granules were released more rapidly than the D-granules, although almost all granules were finally exocytosed. The granular hemocyte possessed a single Golgi complex; possible precursor granules of L-granules and D-granules contained tubular and condensed dense material, respectively. These data are discussed in relation to the self-defense mechanisms of the horseshoe crab.

掲載種別：研究論文（学術雑誌）  

The structure of hemocytes in the normal state and during blood coagulation, and the intracellular localization of three clotting factors and two antimicrobial factors were examined in the Japanese horseshoe crab Tachypleus tridentatus. Two types of hemocytes were found in the circulating blood: non-granular and granular hemocytes. The latter contained numerous dense granules classed into two major types: L- and D-granules. The L-granules were larger (up to 1.5-mu-m in diameter) and less electron-dense than the D-granules (less than 0.6-mu-m in diameter). The L-granules contained three clotting factors and one antimicrobial factor, whereas the D-granules exclusively contained the other antimicrobial factor. After treatment with endotoxin, the L-granules were released more rapidly than the D-granules, although almost all granules were finally exocytosed. The granular hemocyte possessed a single Golgi complex; possible precursor granules of L-granules and D-granules contained tubular and condensed dense material, respectively. These data are discussed in relation to the self-defense mechanisms of the horseshoe crab.

掲載種別：研究論文（学術雑誌）  

Direct virus inactivation of tachyplesin I and related isopeptides, which are antimicrobial peptides isolated from the hemocytes of the horseshoe crab (Tachypleus tridentatus and Limulus polyphemus), was examined against several viruses. Vesicular stomatitis virus (VSV) was inactivated by incubation with tachyplesin I and its isopeptides. Influenza A (H1N1) virus was slightly inactivated by tachyplesin I, whereas herpes simplex virus 1 and 2, adenovirus 1, reovirus 2 and poliovirus 1 were resistant to inactivation. The inactivation of VSV by tachyplesin I depended on the concentration, the time and the temperature of incubation. Pretreatment of tachyplesin I with trypsin or lipopolysaccharide of gram-negative bacteria entirely abolished the antiviral activity. Electron microscopy of VSV treated with tachyplesin I showed naked and damaged virions. These data suggest that tachyplesin I directly inactivates the VSV by destroying its envelope subunits.

掲載種別：研究論文（学術雑誌）  

A novel thermolabile β-2 macroglycoprotein ('thermolabile substance' (TLS) or 'Hakata antigen' (HA)), which was detected by the precipitating (auto) antibodies of patients with systemic lupus erythematosus, was isolated and characterized. The purification procedure entailed the following steps: isoelectric precipitation in the range between pH 5.2-6.1, hydroxyapatite absorption chromatography, 35% saturated ammonium sulfate precipitation, Sephadex G-200 gel filtration, Pevikon block electrophoresis, lentil lectin affinity chromatography and immobilized rabbit anti-human whole serum IgG column chromatography. Utilizing these procedures, 0.1 mg of HA was purified from 3 I of pooled human serum. The molecular mass of HA was determined as 650 kDa by Sepharose 4B gel filtration. On SDS-PAGE analysis, HA showed a single band at 35 kDa under reduced conditions and numerous ladder bands between 35 kDa to more than 300 kDa under nonreduced conditions. On analytical ultracentrifugation, Ha gave a molecular mass of 520 kDa with a single meniscus and a sedimentation constant of 12.0. The amino acid and carbohydrate analysis of reduced and S-pyridylethylated HA revealed that it contained five residues of hydroxyproline and an N-linked type sugar chain. © 1991.

DOI： 10.1016/0167-4838(91)90158-V

PubMed

掲載種別：研究論文（学術雑誌）  

A novel thermolabile β-2 macroglycoprotein ('thermolabile substance' (TLS) or 'Hakata antigen' (HA)), which was detected by the precipitating (auto) antibodies of patients with systemic lupus erythematosus, was isolated and characterized. The purification procedure entailed the following steps: isoelectric precipitation in the range between pH 5.2-6.1, hydroxyapatite absorption chromatography, 35% saturated ammonium sulfate precipitation, Sephadex G-200 gel filtration, Pevikon block electrophoresis, lentil lectin affinity chromatography and immobilized rabbit anti-human whole serum IgG column chromatography. Utilizing these procedures, 0.1 mg of HA was purified from 3 I of pooled human serum. The molecular mass of HA was determined as 650 kDa by Sepharose 4B gel filtration. On SDS-PAGE analysis, HA showed a single band at 35 kDa under reduced conditions and numerous ladder bands between 35 kDa to more than 300 kDa under nonreduced conditions. On analytical ultracentrifugation, Ha gave a molecular mass of 520 kDa with a single meniscus and a sedimentation constant of 12.0. The amino acid and carbohydrate analysis of reduced and S-pyridylethylated HA revealed that it contained five residues of hydroxyproline and an N-linked type sugar chain. © 1991.

DOI： 10.1016/0167-4838(91)90158-V

PubMed

掲載種別：研究論文（学術雑誌）  

Factor C is an endotoxin-sensitive, intracellular serine protease zymogen which initiates the coagulation cascade system in the limulus hemolymph. We have determined the entire amino acid sequence of factor C using recombinant DNA technique. The zymogen consisted of 994 amino acid residues with a calculated molecular mass of 109,648 Da. Most interestingly, factor C has five repeating units ("Sushi" domain or short consensus repeat) of about 60 amino acid residues each, which have been found in many proteins participating in the mammalian complement system. In addition to a typical serine protease domain in the carboxyl-terminal portion, characteristic segments with an epidermal growth factor-like, a lectin-like, a cysteine-rich, and a proline-rich domain were also found, revealing a unique mosaic protein structure. The serine protease domain was most analogous to human thrombin. Factor C was identified to localize in large granules in the cell, indicating that it is released from the cell by lipopolysaccharide stimulation. Furthermore, we identified a transcript possibly derived by alternative splicing of factor C mRNA, which encodes a protein sharing the amino-terminal portion of factor C. We suggest that factor C, a newly discovered type of serine protease zymogen, is a "coagulation-complement factor" which may play important roles in both hemostasis and host defense mechanisms.

掲載種別：研究論文（学術雑誌）  

Factor C is an endotoxin-sensitive, intracellular serine protease zymogen which initiates the coagulation cascade system in the limulus hemolymph. We have determined the entire amino acid sequence of factor C using recombinant DNA technique. The zymogen consisted of 994 amino acid residues with a calculated molecular mass of 109,648 Da. Most interestingly, factor C has five repeating units ("Sushi" domain or short consensus repeat) of about 60 amino acid residues each, which have been found in many proteins participating in the mammalian complement system. In addition to a typical serine protease domain in the carboxyl-terminal portion, characteristic segments with an epidermal growth factor-like, a lectin-like, a cysteine-rich, and a proline-rich domain were also found, revealing a unique mosaic protein structure. The serine protease domain was most analogous to human thrombin. Factor C was identified to localize in large granules in the cell, indicating that it is released from the cell by lipopolysaccharide stimulation. Furthermore, we identified a transcript possibly derived by alternative splicing of factor C mRNA, which encodes a protein sharing the amino-terminal portion of factor C. We suggest that factor C, a newly discovered type of serine protease zymogen, is a "coagulation-complement factor" which may play important roles in both hemostasis and host defense mechanisms.

掲載種別：研究論文（学術雑誌）  

米国産カブトガニ(Limulus polyphemus)よりC因子を精製し,日本産カブトガニのC因子と比較した.米国産C因子は分子量およびアミノ酸組成,サブユニット構成のほか,免疫化学的にも日本産のそれと相同であった.日本産C因子同様,米国産C因子もリポ多糖や合成リピドAにより自己触媒的に活性化され,興味深いことに両C因子はトリプシンではなくα-キモトリプシンによって活性化され,活性型C因子はα-トロンビン様の基質特異性を示した.一方,体液凝固系に関与するすべての因子を含む血球抽出液もキモトリプシンにより活性化され,ゲルを形成した.哺乳類の凝固・線溶・補体系がすべてトリプシン様酵素によって活性化されるのに対して,カブドガニ凝固系はキモトリプシンによって開始される新しいタイプのカスケード系である

掲載種別：研究論文（学術雑誌）  

米国産カブトガニ(Limulus polyphemus)よりC因子を精製し,日本産カブトガニのC因子と比較した.米国産C因子は分子量およびアミノ酸組成,サブユニット構成のほか,免疫化学的にも日本産のそれと相同であった.日本産C因子同様,米国産C因子もリポ多糖や合成リピドAにより自己触媒的に活性化され,興味深いことに両C因子はトリプシンではなくα-キモトリプシンによって活性化され,活性型C因子はα-トロンビン様の基質特異性を示した.一方,体液凝固系に関与するすべての因子を含む血球抽出液もキモトリプシンにより活性化され,ゲルを形成した.哺乳類の凝固・線溶・補体系がすべてトリプシン様酵素によって活性化されるのに対して,カブドガニ凝固系はキモトリプシンによって開始される新しいタイプのカスケード系である

掲載種別：研究論文（学術雑誌）  

米国産カブトガニ(Limulus polyphemus)よりC因子を精製し,日本産カブトガニのC因子と比較した.米国産C因子は分子量およびアミノ酸組成,サブユニット構成のほか,免疫化学的にも日本産のそれと相同であった.日本産C因子同様,米国産C因子もリポ多糖や合成リピドAにより自己触媒的に活性化され,興味深いことに両C因子はトリプシンではなくα-キモトリプシンによって活性化され,活性型C因子はα-トロンビン様の基質特異性を示した.一方,体液凝固系に関与するすべての因子を含む血球抽出液もキモトリプシンにより活性化され,ゲルを形成した.哺乳類の凝固・線溶・補体系がすべてトリプシン様酵素によって活性化されるのに対して,カブドガニ凝固系はキモトリプシンによって開始される新しいタイプのカスケード系である

掲載種別：研究論文（学術雑誌）  

掲載種別：研究論文（学術雑誌）  

掲載種別：研究論文（国際会議プロシーディングス）  

掲載種別：研究論文（学術雑誌）  

掲載種別：研究論文（学術雑誌）  

掲載種別：研究論文（学術雑誌）  

DOI： 10.1016/0006-291X(90)91199-3

その他URL： http://orcid.org/0000-0003-2409-6556

掲載種別：研究論文（学術雑誌）  

DOI： 10.1016/0006-291X(90)91199-3

DOI： 10.1016/0006-291x(90)91199-3

掲載種別：研究論文（学術雑誌）  

オルニチントランスカルバミラーゼ(OTC)とOTC前駆体の性質を比較する目的で,ラットOTCを大腸菌で発現させ,沈澱画分より尿素またはグアニジン塩酸で抽出し,ほぼ均一に精製した.塩酸グアニジンで可溶化した成熟型OTC希釈後0°Cでインキュベートすることにより,比活性170 μmol/min/mg protein (native酵素の18%)まで活性を回復した.同様の条件下で,OTC前駆体は125 μmol/min/mgまで活性化された.nativeおよび再活性化OTCの沈降係数は6.2Sであるのに対し,活性化OTC前駆体は5.2Sと少し異なるが,ともに三量体を形成していることが示唆された.したがって,前駆体に附着している延長ペプチドは必ずしも前駆体のfoldingを阻害しないことが示された.精製前駆体を8M尿素で可溶化後希釈すると直ちに不溶化して沈澱するが,網状赤血球ライセート存在下では沈降係数6〜11Sの可溶性複合体を形成した.成熟OTCではこのような複合体の形成は見られない.この複合体形成は精製前駆体および化学合成したOTCの延長ペプチドによって阻害されたが,成熟型OTCでは阻害されなかった.これらの結果より,網状赤血球ライセートのなかには,OTCの延長ペプチド部分に結合して可溶性の複合体を形成するサイトソル因子が存在することが示唆された

掲載種別：研究論文（学術雑誌）  

掲載種別：研究論文（学術雑誌）  

掲載種別：研究論文（学術雑誌）  

オルニチントランスカルバミラーゼ(OTC)とOTC前駆体の性質を比較する目的で,ラットOTCを大腸菌で発現させ,沈澱画分より尿素またはグアニジン塩酸で抽出し,ほぼ均一に精製した.塩酸グアニジンで可溶化した成熟型OTC希釈後0°Cでインキュベートすることにより,比活性170 μmol/min/mg protein (native酵素の18%)まで活性を回復した.同様の条件下で,OTC前駆体は125 μmol/min/mgまで活性化された.nativeおよび再活性化OTCの沈降係数は6.2Sであるのに対し,活性化OTC前駆体は5.2Sと少し異なるが,ともに三量体を形成していることが示唆された.したがって,前駆体に附着している延長ペプチドは必ずしも前駆体のfoldingを阻害しないことが示された.精製前駆体を8M尿素で可溶化後希釈すると直ちに不溶化して沈澱するが,網状赤血球ライセート存在下では沈降係数6〜11Sの可溶性複合体を形成した.成熟OTCではこのような複合体の形成は見られない.この複合体形成は精製前駆体および化学合成したOTCの延長ペプチドによって阻害されたが,成熟型OTCでは阻害されなかった.これらの結果より,網状赤血球ライセートのなかには,OTCの延長ペプチド部分に結合して可溶性の複合体を形成するサイトソル因子が存在することが示唆された

掲載種別：研究論文（学術雑誌）  

オルニチントランスカルバミラーゼ(OTC)とOTC前駆体の性質を比較する目的で,ラットOTCを大腸菌で発現させ,沈澱画分より尿素またはグアニジン塩酸で抽出し,ほぼ均一に精製した.塩酸グアニジンで可溶化した成熟型OTC希釈後0°Cでインキュベートすることにより,比活性170 μmol/min/mg protein (native酵素の18%)まで活性を回復した.同様の条件下で,OTC前駆体は125 μmol/min/mgまで活性化された.nativeおよび再活性化OTCの沈降係数は6.2Sであるのに対し,活性化OTC前駆体は5.2Sと少し異なるが,ともに三量体を形成していることが示唆された.したがって,前駆体に附着している延長ペプチドは必ずしも前駆体のfoldingを阻害しないことが示された.精製前駆体を8M尿素で可溶化後希釈すると直ちに不溶化して沈澱するが,網状赤血球ライセート存在下では沈降係数6～11Sの可溶性複合体を形成した.成熟OTCではこのような複合体の形成は見られない.この複合体形成は精製前駆体および化学合成したOTCの延長ペプチドによって阻害されたが,成熟型OTCでは阻害されなかった.これらの結果より,網状赤血球ライセートのなかには,OTCの延長ペプチド部分に結合して可溶性の複合体を形成するサイトソル因子が存在することが示唆された

掲載種別：研究論文（学術雑誌）  

掲載種別：研究論文（学術雑誌）  

掲載種別：研究論文（学術雑誌）  

Proteasomes are multicatalytic proteinase complexes consisting of a set of non-identical polypeptide subunits. A cDNA for component C5 of rat proteasomes was isolated by screening a Reuber H4TG hepatoma cell cDNA library using synthetic oligodeoxynucleotide probes corresponding to partial amino acid sequences of the protein. The polypeptide deduced from the open reading frame consisted of 240 amino acid residues with a calculated molecular weight of 26479. Computer analysis revealed little similarity of C5 to other proteins reported so far. The primary structure of C5 showed partial sequence homology to that of another component C3, but no regions of homology with the sequence of component C2. Thus C5 is concluded to be a new type of subunit of the proteasome complex. © 1990.

DOI： 10.1016/0014-5793(90)80773-C

PubMed

掲載種別：研究論文（学術雑誌）  

Proteasomes are multicatalytic proteinase complexes consisting of a set of non-identical polypeptide subunits. A cDNA for component C5 of rat proteasomes was isolated by screening a Reuber H4TG hepatoma cell cDNA library using synthetic oligodeoxynucleotide probes corresponding to partial amino acid sequences of the protein. The polypeptide deduced from the open reading frame consisted of 240 amino acid residues with a calculated molecular weight of 26479. Computer analysis revealed little similarity of C5 to other proteins reported so far. The primary structure of C5 showed partial sequence homology to that of another component C3, but no regions of homology with the sequence of component C2. Thus C5 is concluded to be a new type of subunit of the proteasome complex. © 1990.

DOI： 10.1016/0014-5793(90)80773-C

PubMed

掲載種別：研究論文（学術雑誌）  

DOI： 10.1016/0014-5793(90)80267-M

その他URL： http://orcid.org/0000-0003-2409-6556

掲載種別：研究論文（学術雑誌）  

DOI： 10.1016/0014-5793(90)80267-M

DOI： 10.1016/0014-5793(90)80267-m

掲載種別：研究論文（学術雑誌）  

DOI： 10.1021/bi00467a026

J-GLOBAL

掲載種別：研究論文（学術雑誌）  

DOI： 10.1021/bi00467a026

J-GLOBAL

掲載種別：研究論文（学術雑誌）  

掲載種別：研究論文（学術雑誌）  

掲載種別：研究論文（学術雑誌）  

掲載種別：研究論文（学術雑誌）  

掲載種別：研究論文（学術雑誌）  

DOI： 10.1007/978-1-4757-5140-6_23

PubMed

掲載種別：研究論文（国際会議プロシーディングス）  

掲載種別：研究論文（国際会議プロシーディングス）  

掲載種別：研究論文（学術雑誌）  

掲載種別：研究論文（学術雑誌）  

日本産および米国産(Limulus polyphemus)の血球中に,抗菌性を示すタキプレシン2とポリフェムシン1,2を新たに見いだし,それらの一次構造を決定した.タキプレシン2は17アミノ酸残基から成り,先に見いだしたタキプレシン1(改名)との間では,僅かにNH2末端Lys残基がArg残基に,15番目のArg残基がLys残基に置換されている以外,同じ構造を示した.一方,米国産のポリフェムシン1と2は,18アミノ酸残基を含み,両者の違いはNH2末端から10番目が,1ではArg, 2ではLysに置換されていた.これらのペプチドは,いずれもC末端Arg-α-amideを有し,かつ強塩基性のものであった.ジスルフィド結合は2個所あり,その架橋結合はポリフェムシン1を用いて決定したのみであるが,タキプレシン1と同様であった.抗菌性については,タキプレシン1と同様,グラム陰性菌と陽性菌に対して強い成長阻止活性を示し,新たに抗カビ活性のあることを見いだした

掲載種別：研究論文（学術雑誌）  

日本産および米国産(Limulus polyphemus)の血球中に,抗菌性を示すタキプレシン2とポリフェムシン1,2を新たに見いだし,それらの一次構造を決定した.タキプレシン2は17アミノ酸残基から成り,先に見いだしたタキプレシン1(改名)との間では,僅かにNH2末端Lys残基がArg残基に,15番目のArg残基がLys残基に置換されている以外,同じ構造を示した.一方,米国産のポリフェムシン1と2は,18アミノ酸残基を含み,両者の違いはNH2末端から10番目が,1ではArg, 2ではLysに置換されていた.これらのペプチドは,いずれもC末端Arg-α-amideを有し,かつ強塩基性のものであった.ジスルフィド結合は2個所あり,その架橋結合はポリフェムシン1を用いて決定したのみであるが,タキプレシン1と同様であった.抗菌性については,タキプレシン1と同様,グラム陰性菌と陽性菌に対して強い成長阻止活性を示し,新たに抗カビ活性のあることを見いだした

掲載種別：研究論文（学術雑誌）  

日本産および米国産(Limulus polyphemus)の血球中に,抗菌性を示すタキプレシン2とポリフェムシン1,2を新たに見いだし,それらの一次構造を決定した.タキプレシン2は17アミノ酸残基から成り,先に見いだしたタキプレシン1(改名)との間では,僅かにNH2末端Lys残基がArg残基に,15番目のArg残基がLys残基に置換されている以外,同じ構造を示した.一方,米国産のポリフェムシン1と2は,18アミノ酸残基を含み,両者の違いはNH2末端から10番目が,1ではArg, 2ではLysに置換されていた.これらのペプチドは,いずれもC末端Arg-α-amideを有し,かつ強塩基性のものであった.ジスルフィド結合は2個所あり,その架橋結合はポリフェムシン1を用いて決定したのみであるが,タキプレシン1と同様であった.抗菌性については,タキプレシン1と同様,グラム陰性菌と陽性菌に対して強い成長阻止活性を示し,新たに抗カビ活性のあることを見いだした

掲載種別：研究論文（学術雑誌）  

掲載種別：研究論文（学術雑誌）  

掲載種別：研究論文（学術雑誌）  

掲載種別：研究論文（学術雑誌）  

掲載種別：研究論文（学術雑誌）  

掲載種別：研究論文（学術雑誌）  

Tachyplesin is an antimicrobial peptide recently found in the acid extract of hemocytes from the Japanese horseshoe crab (Tachypleus tridentatus) [Nakamura, T. et al. (1988) J. Biol Chem. 263, 16709-16713]. In our continuing studies on the peptide, we have found an isopeptide, tachyplesin II, and also polyphemusins I and II in hemocytes of the American horseshoe crab (Limulus polyphemus). The complete primary structures of these peptides, which are very similar to that of the previously isolated peptide, now named tachyplesin I, were determined to be as follows:Polyphemusin I NH2-R-R-W-C-F-R-V-C-Y-R-G-F-C-Y-R-K-C-R-CONH2Polyphemusin II NH2-R-R-W-C-F-R-V-C-Y-K-G-F-C-Y-R-K-C-R-CONH2Tachyplesin II NH2-R-W-C-F-R-V-C-Y-R-G-I- C-Y-R-K-C-R-CONH2The isopeptide, tachyplesin II, consists of 17 residues with a COOH-terminal arginine a-amide. On the other hand, both polyphemusins I and II were found to contain 18 residues due to an additional Arg residue at the NH2-terminal end as well as a COOH-terminal arginine α-amide. The disulfide linkages for polyphemusin I consisted of two bridges between Cys-4 and Cys-17 and between Cys-8 and Cys-13, which was identical to in the case of tachyplesin I. Moreover, all of these peptides inhibited the growth of not only Gram-negative and -positive bacteria but also fungi, such as Candida albicans M9. Furthermore, complex formation between these peptides and bacterial lipopolysaccharides was also observed in a double diffusion test. These results suggest that tachyplesins and poly-phemusins are probably located in the hemocyte membrane, where they act on antimi-crobial peptides as a self-defense mechanism in the horseshoe crab against invading microorganisms. © 1989 BY THE JOURNAL OF BIOCHEMISTRY.

DOI： 10.1093/oxfordjournals.jbchem.a122913

PubMed

掲載種別：研究論文（学術雑誌）  

Tachyplesin I, a 17-residue peptide amide with two disulfide bridges isolated from an acid extract of horseshoe crab hemocyte debris, was synthesized by the 9-fluorenylmethyloxycarbonyl (Fmoc)-based solid-phase method followed by two steps of deprotection and subsequent air-oxidation. The thioanisole-mediated deprotection with 1 M trifluoromethanesulfonic acid was first employed to cleave the peptide amide from the resin and, at the same time, to deprotect the side chain protecting groups employed, except for the S-Acm (acetamidomethyl) group. The 4 Acm groups attached were next cleaved by silver trifluoromethanesulfonate. In addition, two related peptides, tachyplesin II and polyphemusin I, were similarly synthesized. Synthetic tachyplesin I inhibited the lipopolysaccharide-mediated activation of clotting factor C to the same extent as did the corresponding natural peptide. The relative potencies of synthetic tachyplesin II and synthetic polyphemusin I with respect to natural tachyplesin I (taken as 1) were 2.1 and 0.61, respectively. © 1989, The Pharmaceutical Society of Japan. All rights reserved.

DOI： 10.1248/cpb.37.2661

掲載種別：研究論文（学術雑誌）  

Tachyplesin is an antimicrobial peptide recently found in the acid extract of hemocytes from the Japanese horseshoe crab (<I>Tachypleus tridentatus</I>)[Nakamura, T. et al.(1988) <I>J. Biol. Chem</I>. 263, 16709-16713]. In our continuing studies on the peptide, we have found an isopeptide, tachyplesin II, and also polyphemusins I and II in hemocytes of the American horseshoe crab (<I>Limulus polyphemus</I>). The complete primary structures of these peptides, which are very similar to that of the previously isolated peptide, now named tachyplesin I, were determined to be as follows:<BR>Polyphemusin I NH<I>2</I>-R-R-W-C-F-R-V-C-Y-R-G-F-C-Y-R-K-C-R-CONH<I>2</I><BR>Polyphemusin II NH<I>2</I>-R-R-W-C-F-R-V-C-Y-K-G-F-C-Y-R-K-C-R-CONH<I>2</I><BR>Tachyplesin II NH<I>2</I>-R-W-C-F-R-V-C-Y-R-G-I- C-Y-R-K-C-R-CONH<I>2</I><BR>The isopeptide, tachyplesin II, consists of 17 residues with a COOH-terminal arginine α-amide. On the other hand, both polyphemusins I and II were found to contain 18 residues due to an additional Arg residue at the NH, -terminal end as well as a COOH-terminal arginine α-amide. The disulfide linkages for polyphemusin I consisted of two bridges between Cys-4 and Cys-17 and between Cys-8 and Cys-13, which was identical to in the case of tachyplesin I. Moreover, all of these peptides inhibited the growth of not only Gramnegative and-positive bacteria but also fungi, such as <I>Candida albicans</I> M9. Furthermore, complex formation between these peptides and bacterial lipopolysaccharides was also observed in a double diffusion test. These results suggest that tachyplesins and polyphemusins are probably located in the hemocyte membrane, where they act on antimicrobial peptides as a self-defense mechanism in the horseshoe crab against invading microorganisms.

CiNii Article

掲載種別：研究論文（学術雑誌）  

Tachyplesin I, a 17-residue peptide amide with two disulfide bridges isolated from an acid extract of horseshoe crab hemocyte debris, was synthesized by the 9-fluorenylmethyloxycarbonyl (Fmoc)-based solid-phase method followed by two steps of deprotection and subsequent air-oxidation. The thioanisole-mediated deprotection with 1 M trifluoromethanesulfonic acid was first employed to cleave the peptide amide from the resin and, at the same time, to deprotect the side chain protecting groups employed, except for the S-Acm (acetamidomethyl) group. The 4 Acm groups attached were next cleaved by silver trifluoromethanesulfonate. In addition, two related peptides, tachyplesin II and polyphemusin I, were similarly synthesized. Synthetic tachyplesin I inhibited the lipopolysaccharide-mediated activation of clotting factor C to the same extent as did the corresponding natural peptide. The relative potencies of synthetic tachyplesin II and synthetic polyphemusin I with respect to natural tachyplesin I (taken as 1) were 2.1 and 0.61, respectively. © 1989, The Pharmaceutical Society of Japan. All rights reserved.

DOI： 10.1248/cpb.37.2661

掲載種別：研究論文（学術雑誌）  

Tachyplesin is an antimicrobial peptide recently found in the acid extract of hemocytes from the Japanese horseshoe crab (Tachypleus tridentatus) [Nakamura, T. et al. (1988) J. Biol Chem. 263, 16709-16713]. In our continuing studies on the peptide, we have found an isopeptide, tachyplesin II, and also polyphemusins I and II in hemocytes of the American horseshoe crab (Limulus polyphemus). The complete primary structures of these peptides, which are very similar to that of the previously isolated peptide, now named tachyplesin I, were determined to be as follows:Polyphemusin I NH2-R-R-W-C-F-R-V-C-Y-R-G-F-C-Y-R-K-C-R-CONH2Polyphemusin II NH2-R-R-W-C-F-R-V-C-Y-K-G-F-C-Y-R-K-C-R-CONH2Tachyplesin II NH2-R-W-C-F-R-V-C-Y-R-G-I- C-Y-R-K-C-R-CONH2The isopeptide, tachyplesin II, consists of 17 residues with a COOH-terminal arginine a-amide. On the other hand, both polyphemusins I and II were found to contain 18 residues due to an additional Arg residue at the NH2-terminal end as well as a COOH-terminal arginine α-amide. The disulfide linkages for polyphemusin I consisted of two bridges between Cys-4 and Cys-17 and between Cys-8 and Cys-13, which was identical to in the case of tachyplesin I. Moreover, all of these peptides inhibited the growth of not only Gram-negative and -positive bacteria but also fungi, such as Candida albicans M9. Furthermore, complex formation between these peptides and bacterial lipopolysaccharides was also observed in a double diffusion test. These results suggest that tachyplesins and poly-phemusins are probably located in the hemocyte membrane, where they act on antimi-crobial peptides as a self-defense mechanism in the horseshoe crab against invading microorganisms. © 1989 BY THE JOURNAL OF BIOCHEMISTRY.

DOI： 10.1093/oxfordjournals.jbchem.a122913

PubMed

掲載種別：研究論文（学術雑誌）  

Tachyplesin is an antimicrobial peptide recently found in the acid extract of hemocytes from the Japanese horseshoe crab (<I>Tachypleus tridentatus</I>)[Nakamura, T. et al.(1988) <I>J. Biol. Chem</I>. 263, 16709-16713]. In our continuing studies on the peptide, we have found an isopeptide, tachyplesin II, and also polyphemusins I and II in hemocytes of the American horseshoe crab (<I>Limulus polyphemus</I>). The complete primary structures of these peptides, which are very similar to that of the previously isolated peptide, now named tachyplesin I, were determined to be as follows:<BR>Polyphemusin I NH<I>2</I>-R-R-W-C-F-R-V-C-Y-R-G-F-C-Y-R-K-C-R-CONH<I>2</I><BR>Polyphemusin II NH<I>2</I>-R-R-W-C-F-R-V-C-Y-K-G-F-C-Y-R-K-C-R-CONH<I>2</I><BR>Tachyplesin II NH<I>2</I>-R-W-C-F-R-V-C-Y-R-G-I- C-Y-R-K-C-R-CONH<I>2</I><BR>The isopeptide, tachyplesin II, consists of 17 residues with a COOH-terminal arginine α-amide. On the other hand, both polyphemusins I and II were found to contain 18 residues due to an additional Arg residue at the NH, -terminal end as well as a COOH-terminal arginine α-amide. The disulfide linkages for polyphemusin I consisted of two bridges between Cys-4 and Cys-17 and between Cys-8 and Cys-13, which was identical to in the case of tachyplesin I. Moreover, all of these peptides inhibited the growth of not only Gramnegative and-positive bacteria but also fungi, such as <I>Candida albicans</I> M9. Furthermore, complex formation between these peptides and bacterial lipopolysaccharides was also observed in a double diffusion test. These results suggest that tachyplesins and polyphemusins are probably located in the hemocyte membrane, where they act on antimicrobial peptides as a self-defense mechanism in the horseshoe crab against invading microorganisms.

CiNii Article

掲載種別：研究論文（学術雑誌）  

掲載種別：研究論文（学術雑誌）  

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J-GLOBAL

掲載種別：記事・総説・解説・論説等（学術雑誌）  

掲載種別：記事・総説・解説・論説等（学術雑誌）  

ユビキチンは、タンパク質をプロテアソーム分解へ導く標識として発見されたが、ユビキチン分子間やユビキチン-基質間で多様に連結することで多彩な細胞機能を制御することが明らかになり、ユビキチンコード(ユビキチン暗号)と称されるようになった。Linear ubiquitin chain assembly complex(LUBAC)は、ユビキチンのN末端Met1を介する直鎖状ユビキチン鎖を生成する唯一のユビキチンリガーゼで、炎症・免疫制御に重要なnuclear factor-κB(NF-κB)シグナルを活性化する。また、直鎖状ユビキチン鎖を分解する脱ユビキチン化酵素や結合タンパク質も同定され、LUBAC制御がになう生理機能や破綻が引き起こす疾患の解明、創薬をめざした阻害剤開発が進められている。(著者抄録)

掲載種別：記事・総説・解説・論説等（学術雑誌）  

J-GLOBAL

掲載種別：記事・総説・解説・論説等（学術雑誌）  

J-GLOBAL

掲載種別：記事・総説・解説・論説等（学術雑誌）  

J-GLOBAL

掲載種別：記事・総説・解説・論説等（学術雑誌）  

脱ユビキチン化酵素(DUB)は,タンパク質分解やシグナル伝達の活性化などのユビキチン化修飾を介した細胞応答を負に制御する重要な因子として注目されている.また,DUBの高発現や異常な活性化が,がんや神経疾患などを誘導することが明らかとなっており,創薬ターゲットとしても注目されている.我々はコムギ無細胞系を用いてヒトのDUBの約8割を網羅する83種類の組換えタンパク質を合成し,DUBタンパク質アレイを作製した.このDUBタンパク質アレイについて,8種類の重合様式の二量体ユビキチンを基質として,76種類のDUBについて活性を検出し,そのユビキチン鎖特異性を明らかにした.さらに,このDUBタンパク質アレイを応用することで,DUBを阻害する化合物のDUB特異性評価パネルを作製した.(著者抄録)

掲載種別：記事・総説・解説・論説等（学術雑誌）  

ユビキチンのN末端を介して形成される非定型型の直鎖状ユビキチン鎖は,炎症・免疫シグナル応答や細胞死を制御し,その生成を特異的に担うLUBACユビキチンリガーゼの機能不全は多くの疾患発症に関わる.筆者らは家族性筋萎縮性側索硬化症(ALS)の原因遺伝子の一つであるオプチニューリン(OPTN)やアルツハイマー病タウタンパク質の細胞機能解析から,LUBACによる直鎖状ユビキチン鎖生成が異常タンパク質封入体形成,神経炎症・細胞死の亢進に寄与することを見いだした.さらに,新規治療薬創出を最終目的としてLUBACに対する阻害剤の探索を行い,細胞毒性が低くLUBAC特異性の高い新規化合物(HOIPINs)を見いだした.本稿では,筆者らの研究を中心に国内外の動向を交えて,これらの知見を紹介したい.(著者抄録)

掲載種別：記事・総説・解説・論説等（学術雑誌）  

脱ユビキチン化酵素(DUB)は,タンパク質分解やシグナル伝達の活性化などのユビキチン化修飾を介した細胞応答を負に制御する重要な因子として注目されている.また,DUBの高発現や異常な活性化が,がんや神経疾患などを誘導することが明らかとなっており,創薬ターゲットとしても注目されている.我々はコムギ無細胞系を用いてヒトのDUBの約8割を網羅する83種類の組換えタンパク質を合成し,DUBタンパク質アレイを作製した.このDUBタンパク質アレイについて,8種類の重合様式の二量体ユビキチンを基質として,76種類のDUBについて活性を検出し,そのユビキチン鎖特異性を明らかにした.さらに,このDUBタンパク質アレイを応用することで,DUBを阻害する化合物のDUB特異性評価パネルを作製した.(著者抄録)

掲載種別：記事・総説・解説・論説等（学術雑誌）  

ユビキチンのN末端を介して形成される非定型型の直鎖状ユビキチン鎖は,炎症・免疫シグナル応答や細胞死を制御し,その生成を特異的に担うLUBACユビキチンリガーゼの機能不全は多くの疾患発症に関わる.筆者らは家族性筋萎縮性側索硬化症(ALS)の原因遺伝子の一つであるオプチニューリン(OPTN)やアルツハイマー病タウタンパク質の細胞機能解析から,LUBACによる直鎖状ユビキチン鎖生成が異常タンパク質封入体形成,神経炎症・細胞死の亢進に寄与することを見いだした.さらに,新規治療薬創出を最終目的としてLUBACに対する阻害剤の探索を行い,細胞毒性が低くLUBAC特異性の高い新規化合物(HOIPINs)を見いだした.本稿では,筆者らの研究を中心に国内外の動向を交えて,これらの知見を紹介したい.(著者抄録)

掲載種別：記事・総説・解説・論説等（学術雑誌）   共著区分：単著  

掲載種別：記事・総説・解説・論説等（学術雑誌）  

掲載種別：記事・総説・解説・論説等（学術雑誌）  

掲載種別：記事・総説・解説・論説等（学術雑誌）  

J-GLOBAL

掲載種別：記事・総説・解説・論説等（学術雑誌）   共著区分：単著  

掲載種別：記事・総説・解説・論説等（学術雑誌）   共著区分：単著  

掲載種別：記事・総説・解説・論説等（学術雑誌）  

掲載種別：記事・総説・解説・論説等（学術雑誌）  

掲載種別：記事・総説・解説・論説等（学術雑誌）  

掲載種別：記事・総説・解説・論説等（学術雑誌）  

J-GLOBAL

掲載種別：記事・総説・解説・論説等（学術雑誌）  

J-GLOBAL

掲載種別：記事・総説・解説・論説等（学術雑誌）   共著区分：単著  

脱ユビキチン化酵素(DUB)は連結したユビキチンのC末端を加水分解するプロテアーゼで、ヒトでは約100種存在する。DUBはシステインプロテアーゼのUSP、UCH、ジョセフィン、OTU、MINDYファミリーと金属プロテアーゼのJAMM/MPN+ファミリーという6つのファミリーに分類され、ユビキチン修飾が制御する多彩な細胞機能変換に対して拮抗的に働き、フリーのユビキチン量を確保する。したがって、DUBの遺伝子異常はユビキチンシグナルの制御不全やさまざまな疾患を発症させる。本稿ではヒトDUB酵素群を俯瞰的に概説するとともに、炎症・免疫制御をつかさどり、遺伝子異常が疾患を引き起こすDUBとして知られるCYLD、A20、OTULINを紹介する。さらに、近年注目されているオートファジー制御に関わるDUBやDUBを標的とした最新の創薬研究を解説する。(著者抄録)

掲載種別：記事・総説・解説・論説等（学術雑誌）  

脱ユビキチン化酵素(DUB)は連結したユビキチンのC末端を加水分解するプロテアーゼで、ヒトでは約100種存在する。DUBはシステインプロテアーゼのUSP、UCH、ジョセフィン、OTU、MINDYファミリーと金属プロテアーゼのJAMM/MPN+ファミリーという6つのファミリーに分類され、ユビキチン修飾が制御する多彩な細胞機能変換に対して拮抗的に働き、フリーのユビキチン量を確保する。したがって、DUBの遺伝子異常はユビキチンシグナルの制御不全やさまざまな疾患を発症させる。本稿ではヒトDUB酵素群を俯瞰的に概説するとともに、炎症・免疫制御をつかさどり、遺伝子異常が疾患を引き起こすDUBとして知られるCYLD、A20、OTULINを紹介する。さらに、近年注目されているオートファジー制御に関わるDUBやDUBを標的とした最新の創薬研究を解説する。(著者抄録)

掲載種別：記事・総説・解説・論説等（学術雑誌）  

脱ユビキチン化酵素(DUB)は連結したユビキチンのC末端を加水分解するプロテアーゼで、ヒトでは約100種存在する。DUBはシステインプロテアーゼのUSP、UCH、ジョセフィン、OTU、MINDYファミリーと金属プロテアーゼのJAMM/MPN+ファミリーという6つのファミリーに分類され、ユビキチン修飾が制御する多彩な細胞機能変換に対して拮抗的に働き、フリーのユビキチン量を確保する。したがって、DUBの遺伝子異常はユビキチンシグナルの制御不全やさまざまな疾患を発症させる。本稿ではヒトDUB酵素群を俯瞰的に概説するとともに、炎症・免疫制御をつかさどり、遺伝子異常が疾患を引き起こすDUBとして知られるCYLD、A20、OTULINを紹介する。さらに、近年注目されているオートファジー制御に関わるDUBやDUBを標的とした最新の創薬研究を解説する。(著者抄録)

掲載種別：記事・総説・解説・論説等（学術雑誌）   共著区分：共著  

掲載種別：記事・総説・解説・論説等（学術雑誌）  

掲載種別：記事・総説・解説・論説等（学術雑誌）  

J-GLOBAL

掲載種別：記事・総説・解説・論説等（学術雑誌）  

J-GLOBAL

掲載種別：記事・総説・解説・論説等（学術雑誌）   共著区分：単著  

タンパク質のユビキチン修飾は、プロテアソーム分解に導く標識として見出されたが、その後の研究から細胞内輸送、DNA修復、シグナル伝達など多彩な生理機能を制御することが明らかになってきた。我々は、HOIL-1L、HOIP、SHARPINという3サブユニットからなる複合体型ユビキチンリガーゼ(LUBAC)を見出し、LUBACがユビキチンのN末端Met1を介する「直鎖状ユビキチン鎖」という全く新しいタイプのユビキチン連結鎖を特異的に生成することで細胞内シグナル伝達のうちNF-κB経路を制御することを見出した。NF-κBは炎症応答や自然・獲得免疫制御において中心的な働きを司る転写因子で、その活性化経路の破綻は多くの癌、自己免疫疾患、炎症性疾患、生活習慣病、神経変性疾患発症に関わる。また、LUBACによって生成される直鎖状ユビキチン鎖に特異的に結合するドメインをもつタンパク質は、NF-κB活性調節に寄与する。一例として我々は、直鎖状ユビキチン鎖結合ドメイン(UBANドメイン)を有するオプチニューリン(optineurin)がNF-κB活性化経路とアポトーシスを抑制し、そのユビキチン鎖結合性の破綻が筋萎縮性側索硬化症(ALS)発症に深く関わることを突き止めた。(著者抄録)

掲載種別：記事・総説・解説・論説等（学術雑誌）  

J-GLOBAL

掲載種別：記事・総説・解説・論説等（学術雑誌）  

タンパク質のユビキチン修飾は、プロテアソーム分解に導く標識として見出されたが、その後の研究から細胞内輸送、DNA修復、シグナル伝達など多彩な生理機能を制御することが明らかになってきた。我々は、HOIL-1L、HOIP、SHARPINという3サブユニットからなる複合体型ユビキチンリガーゼ(LUBAC)を見出し、LUBACがユビキチンのN末端Met1を介する「直鎖状ユビキチン鎖」という全く新しいタイプのユビキチン連結鎖を特異的に生成することで細胞内シグナル伝達のうちNF-κB経路を制御することを見出した。NF-κBは炎症応答や自然・獲得免疫制御において中心的な働きを司る転写因子で、その活性化経路の破綻は多くの癌、自己免疫疾患、炎症性疾患、生活習慣病、神経変性疾患発症に関わる。また、LUBACによって生成される直鎖状ユビキチン鎖に特異的に結合するドメインをもつタンパク質は、NF-κB活性調節に寄与する。一例として我々は、直鎖状ユビキチン鎖結合ドメイン(UBANドメイン)を有するオプチニューリン(optineurin)がNF-κB活性化経路とアポトーシスを抑制し、そのユビキチン鎖結合性の破綻が筋萎縮性側索硬化症(ALS)発症に深く関わることを突き止めた。(著者抄録)

掲載種別：記事・総説・解説・論説等（学術雑誌）  

タンパク質のユビキチン修飾は、プロテアソーム分解に導く標識として見出されたが、その後の研究から細胞内輸送、DNA修復、シグナル伝達など多彩な生理機能を制御することが明らかになってきた。我々は、HOIL-1L、HOIP、SHARPINという3サブユニットからなる複合体型ユビキチンリガーゼ(LUBAC)を見出し、LUBACがユビキチンのN末端Met1を介する「直鎖状ユビキチン鎖」という全く新しいタイプのユビキチン連結鎖を特異的に生成することで細胞内シグナル伝達のうちNF-κB経路を制御することを見出した。NF-κBは炎症応答や自然・獲得免疫制御において中心的な働きを司る転写因子で、その活性化経路の破綻は多くの癌、自己免疫疾患、炎症性疾患、生活習慣病、神経変性疾患発症に関わる。また、LUBACによって生成される直鎖状ユビキチン鎖に特異的に結合するドメインをもつタンパク質は、NF-κB活性調節に寄与する。一例として我々は、直鎖状ユビキチン鎖結合ドメイン(UBANドメイン)を有するオプチニューリン(optineurin)がNF-κB活性化経路とアポトーシスを抑制し、そのユビキチン鎖結合性の破綻が筋萎縮性側索硬化症(ALS)発症に深く関わることを突き止めた。(著者抄録)

掲載種別：記事・総説・解説・論説等（学術雑誌）  

J-GLOBAL

掲載種別：記事・総説・解説・論説等（学術雑誌）  

J-GLOBAL

掲載種別：記事・総説・解説・論説等（学術雑誌）  

CiNii Article

その他URL： http://search.jamas.or.jp/link/ui/2016004977

掲載種別：記事・総説・解説・論説等（学術雑誌）  

掲載種別：記事・総説・解説・論説等（学術雑誌）   共著区分：単著  

掲載種別：記事・総説・解説・論説等（学術雑誌）  

掲載種別：記事・総説・解説・論説等（学術雑誌）  

CiNii Article

掲載種別：記事・総説・解説・論説等（学術雑誌）   共著区分：単著  

掲載種別：記事・総説・解説・論説等（学術雑誌）  

DOI： 10.1271/kagakutoseibutsu.52.716

CiNii Article

掲載種別：記事・総説・解説・論説等（学術雑誌）  

掲載種別：記事・総説・解説・論説等（学術雑誌）  

掲載種別：研究発表ペーパー・要旨（国際会議）  

掲載種別：記事・総説・解説・論説等（学術雑誌）  

J-GLOBAL

掲載種別：記事・総説・解説・論説等（学術雑誌）  

掲載種別：記事・総説・解説・論説等（学術雑誌）  

J-GLOBAL

掲載種別：記事・総説・解説・論説等（学術雑誌）  

J-GLOBAL

掲載種別：記事・総説・解説・論説等（学術雑誌）   共著区分：単著  

NF-κB経路は免疫や炎症応答に重要なシグナル伝達経路で,ユビキチン化などの翻訳後修飾によって時空間特異的に制御される.本稿では,LUBACユビキチンリガーゼ複合体が生成する新規「直鎖状ポリユビキチン鎖」の生成機構とNF-κB活性化における役割,これに拮抗的に働く脱ユビキチン化酵素について解説する.さらに,LUBAC変異や直鎖状ユビキチン産生不全が関与する多様な疾患について最新の知見を紹介する.(著者抄録)

掲載種別：記事・総説・解説・論説等（学術雑誌）  

NF-κB経路は免疫や炎症応答に重要なシグナル伝達経路で,ユビキチン化などの翻訳後修飾によって時空間特異的に制御される.本稿では,LUBACユビキチンリガーゼ複合体が生成する新規「直鎖状ポリユビキチン鎖」の生成機構とNF-κB活性化における役割,これに拮抗的に働く脱ユビキチン化酵素について解説する.さらに,LUBAC変異や直鎖状ユビキチン産生不全が関与する多様な疾患について最新の知見を紹介する.(著者抄録)

掲載種別：記事・総説・解説・論説等（学術雑誌）   共著区分：単著  

NF-κB(nuclear factor-κB)経路は、免疫制御や炎症応答に中心的なシグナル伝達経路で、リン酸化やユビキチン化などの翻訳後修飾がその時空間特異的なシグナル制御に深く関与する。筆者らはユビキチンのN末端を介する新規「直鎖状ポリユビキチン鎖」を生成するユビキチンリガーゼ(LUBAC)を発見し、LUBACがNF-κB制御に必須であることを明らかにした。さらに、LUBACを介するNF-κB活性化を抑制する脱ユビキチン化酵素としてA20を同定し、A20は直鎖状ユビキチンに結合することでNF-κB活性を調節していること、その制御不全が悪性リンパ腫惹起に連関することを見いだした。本稿では、拡大する直鎖状ポリユビキチン鎖生成によるNF-κB制御の生理的意義とその破綻による疾患との関連について紹介する。(著者抄録)

掲載種別：記事・総説・解説・論説等（学術雑誌）  

NF-κB(nuclear factor-κB)経路は、免疫制御や炎症応答に中心的なシグナル伝達経路で、リン酸化やユビキチン化などの翻訳後修飾がその時空間特異的なシグナル制御に深く関与する。筆者らはユビキチンのN末端を介する新規「直鎖状ポリユビキチン鎖」を生成するユビキチンリガーゼ(LUBAC)を発見し、LUBACがNF-κB制御に必須であることを明らかにした。さらに、LUBACを介するNF-κB活性化を抑制する脱ユビキチン化酵素としてA20を同定し、A20は直鎖状ユビキチンに結合することでNF-κB活性を調節していること、その制御不全が悪性リンパ腫惹起に連関することを見いだした。本稿では、拡大する直鎖状ポリユビキチン鎖生成によるNF-κB制御の生理的意義とその破綻による疾患との関連について紹介する。(著者抄録)

掲載種別：記事・総説・解説・論説等（学術雑誌）  

NF-κB(nuclear factor-κB)経路は、免疫制御や炎症応答に中心的なシグナル伝達経路で、リン酸化やユビキチン化などの翻訳後修飾がその時空間特異的なシグナル制御に深く関与する。筆者らはユビキチンのN末端を介する新規「直鎖状ポリユビキチン鎖」を生成するユビキチンリガーゼ(LUBAC)を発見し、LUBACがNF-κB制御に必須であることを明らかにした。さらに、LUBACを介するNF-κB活性化を抑制する脱ユビキチン化酵素としてA20を同定し、A20は直鎖状ユビキチンに結合することでNF-κB活性を調節していること、その制御不全が悪性リンパ腫惹起に連関することを見いだした。本稿では、拡大する直鎖状ポリユビキチン鎖生成によるNF-κB制御の生理的意義とその破綻による疾患との関連について紹介する。(著者抄録)

掲載種別：記事・総説・解説・論説等（学術雑誌）  

掲載種別：記事・総説・解説・論説等（学術雑誌）  

掲載種別：記事・総説・解説・論説等（学術雑誌）   共著区分：単著  

NF-κB経路は炎症応答や自然・獲得免疫制御にきわめて重要なシグナル伝達経路で、その機能不全は癌、慢性炎症性疾患、自己免疫疾患などの多くの病態発症に深く関連する。NF-κB経路では、キナーゼによるリン酸化とともにLys48、Lys63、Lys11、直鎖状など多様なポリユビキチン鎖形成が重要な役割を果たす。本稿では筆者らが見出した直鎖状ポリユビキチン鎖を生成するLUBACユビキチンリガーゼ複合体の生理機能を中心に、NF-κB経路の多様なユビキチン修飾による制御と病態との連関を解説する。(著者抄録)

掲載種別：記事・総説・解説・論説等（学術雑誌）  

NF-κB経路は炎症応答や自然・獲得免疫制御にきわめて重要なシグナル伝達経路で、その機能不全は癌、慢性炎症性疾患、自己免疫疾患などの多くの病態発症に深く関連する。NF-κB経路では、キナーゼによるリン酸化とともにLys48、Lys63、Lys11、直鎖状など多様なポリユビキチン鎖形成が重要な役割を果たす。本稿では筆者らが見出した直鎖状ポリユビキチン鎖を生成するLUBACユビキチンリガーゼ複合体の生理機能を中心に、NF-κB経路の多様なユビキチン修飾による制御と病態との連関を解説する。(著者抄録)

掲載種別：記事・総説・解説・論説等（学術雑誌）  

NF-κB経路は炎症応答や自然・獲得免疫制御にきわめて重要なシグナル伝達経路で、その機能不全は癌、慢性炎症性疾患、自己免疫疾患などの多くの病態発症に深く関連する。NF-κB経路では、キナーゼによるリン酸化とともにLys48、Lys63、Lys11、直鎖状など多様なポリユビキチン鎖形成が重要な役割を果たす。本稿では筆者らが見出した直鎖状ポリユビキチン鎖を生成するLUBACユビキチンリガーゼ複合体の生理機能を中心に、NF-κB経路の多様なユビキチン修飾による制御と病態との連関を解説する。(著者抄録)

掲載種別：記事・総説・解説・論説等（学術雑誌）  

J-GLOBAL

掲載種別：記事・総説・解説・論説等（学術雑誌）  

J-GLOBAL

掲載種別：記事・総説・解説・論説等（学術雑誌）  

J-GLOBAL

掲載種別：記事・総説・解説・論説等（学術雑誌）  

J-GLOBAL

掲載種別：記事・総説・解説・論説等（学術雑誌）  

J-GLOBAL

掲載種別：記事・総説・解説・論説等（学術雑誌）  

NF-κBシグナル経路は、免疫制御や炎症に関連する遺伝子群の発現を司るシグナル伝達経路で、リン酸化とともに多様なポリユビキチン鎖形成によって制御される。われわれはSHARPIN/HOIL-1L/HOIPからなるユビキチンリガーゼ複合体(LUBAC)が、ユビキチンのN末端アミノ基を介する新規直鎖状ポリユビキチン鎖の生成を介して、NF-κB経路を特異的に制御するという新しい細胞機構を見出した。(著者抄録)

掲載種別：記事・総説・解説・論説等（学術雑誌）  

NF-κBシグナル経路は、免疫制御や炎症に関連する遺伝子群の発現を司るシグナル伝達経路で、リン酸化とともに多様なポリユビキチン鎖形成によって制御される。われわれはSHARPIN/HOIL-1L/HOIPからなるユビキチンリガーゼ複合体(LUBAC)が、ユビキチンのN末端アミノ基を介する新規直鎖状ポリユビキチン鎖の生成を介して、NF-κB経路を特異的に制御するという新しい細胞機構を見出した。(著者抄録)

掲載種別：記事・総説・解説・論説等（学術雑誌）  

NF-κBシグナル経路は、免疫制御や炎症に関連する遺伝子群の発現を司るシグナル伝達経路で、リン酸化とともに多様なポリユビキチン鎖形成によって制御される。われわれはSHARPIN/HOIL-1L/HOIPからなるユビキチンリガーゼ複合体(LUBAC)が、ユビキチンのN末端アミノ基を介する新規直鎖状ポリユビキチン鎖の生成を介して、NF-κB経路を特異的に制御するという新しい細胞機構を見出した。(著者抄録)