Researcher Profile and Settings

Master

Affiliation (Master)

• Faculty of Medicine　Physiological Science　Biochemistry

Affiliation (Master)

• Faculty of Medicine　Physiological Science　Biochemistry

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Profile and Settings

Profile and Settings

• Name (Japanese)

  Watanabe

• Name (Kana)

  Masashi

• Name

  201301040083733805

Alternate Names

http://kaken.nii.ac.jp/d/r/10632424.ja.html

Achievement

Research Areas

• Life sciences / Physiology
• Life sciences / Pathobiochemistry
• Life sciences / Medical biochemistry

Research Experience

• 2018 - Today 北海道大学大学院 医学研究院 講師
• 2012 北海道大学大学院 医学研究院 助教
• 2009 日本学術振興会特別研究員(DC1)

Awards

• 2017 北海道医学会 北海道医学会研究奨励賞
   

  受賞者: 渡部昌
• 2015 北海道大学医学部同窓会 フラテ研究奨励賞
   

  受賞者: 渡部昌

Published Papers

• 自己免疫性小脳失調症に対する免疫沈降法と質量分析法を用いた網羅的自己抗体測定方法の開発

  工藤 彰彦, 矢口 裕章, 渡部 昌, 上床 尚, 白井 慎一, 岩田 育子, 松島 理明, 高橋 秀尚, 畠山 鎮次, 矢部 一郎

  神経免疫学 (一社)日本神経免疫学会 28 (1) 217 - 217 0918-936X 2023/09
• FBXO11 constitutes a major negative regulator of MHC class II through ubiquitin-dependent proteasomal degradation of CIITA.

  Yusuke Kasuga, Ryota Ouda, Masashi Watanabe, Xin Sun, Miki Kimura, Shigetsugu Hatakeyama, Koichi S Kobayashi

  Proceedings of the National Academy of Sciences of the United States of America 120 (24) e2218955120  2023/06/13 [Refereed]
   

  Major histocompatibility complex (MHC) class I and II molecules play critical roles in the activation and regulation of adaptive immunity through antigen presentation to CD8+ and CD4+ T cells, respectively. Strict regulation of MHC expression is critical for proper immune responses. CIITA (MHC class II transactivator), an NLR (nucleotide-binding domain, leucine-rich-repeat containing) protein, is a master regulator of MHC class II (MHC-II) gene transcription. Although it has been known that CIITA activity is regulated at the transcriptional and protein levels, the mechanism to determine CIITA protein level has not been elucidated. Here, we show that FBXO11 is a bona fide E3 ligase of CIITA and regulates CIITA protein level through ubiquitination-mediated degradation. A nonbiased proteomic approach for CIITA-binding protein identified FBXO11, a member of the Skp1-Cullin-1-F-box E3 ligase complex, as a binding partner of CIITA but not MHC class I transactivator, NLRC5. The cycloheximide chase assay showed that the half-life of CIITA is mainly regulated by FBXO11 via the ubiquitin-proteasome system. The expression of FBXO11 led to the reduced MHC-II at the promoter activity level, transcriptional level, and surface expression level through downregulation of CIITA. Moreover, human and mouse FBXO11-deficient cells display increased levels of MHC-II and related genes. In normal and cancer tissues, FBXO11 expression level is negatively correlated with MHC-II. Interestingly, the expression of FBXO11, along with CIITA, is associated with prognosis of cancer patients. Therefore, FBXO11 is a critical regulator to determine the level of MHC-II, and its expression may serve as a biomarker for cancer.
• Sez6l2 autoimmunity in a large cohort study.

  Megumi Abe, Hiroaki Yaguchi, Akihiko Kudo, Azusa Nagai, Shinichi Shirai, Ikuko Takahashi-Iwata, Masaaki Matsushima, Naoko Nakamura, Kenji Isahaya, Yoshihisa Yamano, Shinji Ashida, Takashi Kasai, Keiko Tanaka, Masashi Watanabe, Takeshi Kondo, Hidehisa Takahashi, Shigetsugu Hatakeyama, Akira Takekoshi, Akio Kimura, Takayoshi Shimohata, Ichiro Yabe

  Journal of neurology, neurosurgery, and psychiatry 94 (8) 667 - 668 2023/06/01 [Refereed]
  
• TRIM27 expression is associated with poor prognosis in sinonasal mucosal melanoma.

  S Kimura, M Suzuki, Y Nakamaru, S Kano, M Watanabe, A Honma, A Nakazono, N Tsushima, S Hatakeyama, A Homma

  Rhinology 2023/03/09 [Refereed]
   

  BACKGROUND: Tripartite motif-containing 27 (TRIM27) has been implicated in the progression of various cancers. However, the role of TRIM27 in sinonasal mucosal melanoma (SNMM) remains poorly understood. MATERIALS & METHODS: We retrospectively examined 28 patients with SNMM treated with between 2003 and 2021. We undertook immunohistochemical analysis of TRIM27, Ki-67, and p-Akt1 expression in SNMM tissues. We also investigated the relationship between TRIM27 expression and clinical characteristics, prognosis, Ki-67 as a tumor growth potential marker, and p-Akt1 as one of the prognostic factors in mucosal melanoma. RESULTS: TRIM27 expression was significantly higher in T4 disease than in T3 disease and was higher in stage IV than in stage III. Patients with high-TRIM27 SNMM had a significantly poorer prognosis in terms of overall survival (OS) and disease-free survival.There was also a significantly higher rate of distant metastasis. Univariate analysis for OS revealed that TRIM27 and T classification were significant poor prognostic factors. In addition, the Ki-67 positive score and the p-Akt1 total staining score were significantly higher in the high-TRIM27 group than in the low-TRIM27 group. CONCLUSIONS: High TRIM27 expression in SNMM was associated with advanced T classification, poor prognosis and distant metastasis. We suggest that TRIM27 has potential as a novel biomarker for prognosis in SNMM.
• TRIM22 negatively regulates MHC-II expression.

  Ayano Inoue, Masashi Watanabe, Takeshi Kondo, Satoshi Hirano, Shigetsugu Hatakeyama

  Biochimica et biophysica acta. Molecular cell research 1869 (10) 119318 - 119318 2022/06/28 [Refereed]
   

  The development of cancer treatment has recently achieved a remarkable breakthrough, and checkpoint blockade immunotherapy has received much attention. To enhance the therapeutic efficacy of checkpoint blockade immunotherapy, recent studies have revealed the importance of activation of CD4+ T cells via an increase in major histocompatibility complex (MHC) class II molecules in cancer cells. Here, we demonstrate that tripartite motif-containing (TRIM) 22, negatively regulates MHC-II expression. Gene knockout of TRIM22 using Cas9-sgRNAs led to an increase of MHC-II proteins, while TRIM22 overexpression remarkably decreased MHC-II proteins. mRNA levels of MHC-II and class II transactivator (CIITA), which plays an essential role in the regulation of MHC-II transcription, were not affected by TRIM22. Furthermore, TRIM22 knockout did not suppress the degradation of MHC-II protein but rather promoted it. These results suggest that TRIM22 decreases MHC-II protein levels through a combination of multiple mechanisms other than transcription or degradation. We showed that inhibition of TRIM22 can increase the amount of MHC-II expression in cancer cells, suggesting a possibility of providing the biological basis for a possible therapeutic target to potentiate checkpoint blockade immunotherapy.
• Loss of FAM83H promotes cell migration and invasion in cutaneous squamous cell carcinoma via impaired keratin distribution.

  Keiko Tokuchi, Shinya Kitamura, Takuya Maeda, Masashi Watanabe, Shigetsugu Hatakeyama, Satoshi Kano, Shinya Tanaka, Hideyuki Ujiie, Teruki Yanagi

  Journal of dermatological science 104 (2) 112 - 121 2021/09/30 [Refereed]
   

  BACKGROUNDS: FAM83H is essential for amelogenesis, but recent reports implicate that FAM83H is involved in the tumorigenesis. We previously clarified that TRIM29 binds to FAM83H to regulate keratin distribution and squamous cell migration. However, little is known about FAM83H in normal/malignant skin keratinocytes. OBJECTIVE: To investigate the expression of FAM83H in cutaneous squamous cell carcinoma (SCC) and its physiological function. METHODS: Immunohistochemical analysis and RT-PCR of human SCC tissues were performed. Next, we examined the effect of FAM83H knockdown/overexpression in SCC cell lines using cell proliferation, migration, and invasion assay. To investigate the molecular mechanism, immunoprecipitation of FAM83H was examined. Further, Immunofluorescence staining was performed. Finally, we examined the correlation between the expressions of FAM83H and the keratin distribution. RESULTS: FAM83H expression was lower in SCC lesions than in normal epidermis and correlated with differentiation grade. The mRNA expression levels of FAM83H in SCC tumors were also lower than in normal epidermis. The knockdown of FAM83H enhanced SCC cell migration and invasion, whereas the overexpression of FAM83H led to decreases in both. Furthermore, the knockdown of FAM83H enhanced the cancer cell metastasis in vivo. FAM83H formed a complex with TRIM29 and keratins. The knockdown of FAM83H altered keratin distribution and solubility. Clinically, the loss of FAM83H correlates with an altered keratin distribution. CONCLUSION: Our findings reveal a critical function for FAM83H in regulating keratin distribution, as well as in the migration/invasion of cutaneous SCC, suggesting that FAM83H could be a crucial molecule in the tumorigenesis of cutaneous SCC.
• Fluticasone Propionate Suppresses Poly(I:C)-Induced ACE2 in Primary Human Nasal Epithelial Cells.

  Akira Nakazono, Yuji Nakamaru, Mahnaz Ramezanpour, Takeshi Kondo, Masashi Watanabe, Shigetsugu Hatakeyama, Shogo Kimura, Aya Honma, P J Wormald, Sarah Vreugde, Masanobu Suzuki, Akihiro Homma

  Frontiers in cellular and infection microbiology 11 655666 - 655666 2021 [Refereed]
   

  Background: From the first detection in 2019, SARS-CoV-2 infections have spread rapidly worldwide and have been proven to cause an urgent and important health problem. SARS-CoV-2 cell entry depends on two proteins present on the surface of host cells, angiotensin-converting enzyme 2 (ACE2) and transmembrane protease serine 2 (TMPRSS2). The nasal cavity is thought to be one of the initial sites of infection and a possible reservoir for dissemination within and between individuals. However, it is not known how the expression of these genes is regulated in the nasal mucosa. Objective: In this study, we examined whether the expression of ACE2 and TMPRSS2 is affected by innate immune signals in the nasal mucosa. We also investigated how fluticasone propionate (FP), a corticosteroid used as an intranasal steroid spray, affects the gene expression. Methods: Primary human nasal epithelial cells (HNECs) were collected from the nasal mucosa and incubated with Toll-like receptor (TLR) agonists and/or fluticasone propionate (FP), followed by quantitative PCR, immunofluorescence, and immunoblot analyses. Results: Among the TLR agonists, the TLR3 agonist Poly(I:C) significantly increased ACE2 and TMPRSS2 mRNA expression in HNECs (ACE2 36.212±11.600-fold change, p<0.0001; TMPRSS2 5.598±2.434-fold change, p=0.031). The ACE2 protein level was also increased with Poly(I:C) stimulation (2.884±0.505-fold change, p=0.003). The Poly(I:C)-induced ACE2 expression was suppressed by co-incubation with FP (0.405±0.312-fold change, p=0.044). Conclusion: The activation of innate immune signals via TLR3 promotes the expression of genes related to SARS-CoV2 cell entry in the nasal mucosa, although this expression is suppressed in the presence of FP. Further studies are required to evaluate whether FP suppresses SARS-CoV-2 viral cell entry.
• Role of intracellular zinc in molecular and cellular function in allergic inflammatory diseases.

  Masanobu Suzuki, Takayoshi Suzuki, Masashi Watanabe, Shigetsugu Hatakeyama, Shogo Kimura, Akira Nakazono, Aya Honma, Yuji Nakamaru, Sarah Vreugde, Akihiro Homma

  Allergology international : official journal of the Japanese Society of Allergology 70 (2) 190 - 200 2020/10/27 [Refereed]
   

  Zinc is an essential micronutrient in human body and a vital cofactor for the function of numerous proteins encoded by the human genome. Zinc has a critical role in maintaining many biochemical and physiological processes at the molecular, cellular, and multiple organ and systemic levels. The alteration of zinc homeostasis causes dysfunction of many organs and systems. In the immune system, zinc regulates the differentiation, proliferation and function of inflammatory cells, including T cells, eosinophils, and B cells, by modifying several signaling pathways such as NFκB signaling pathways and TCR signals. An adequate zinc level is essential for proper immune responses and decreased zinc levels were reported in many allergic inflammatory diseases, including atopic dermatitis, bronchial asthma, and chronic rhinosinusitis. Decreased zinc levels often enhance inflammatory activation. On the other hand, the inflammatory conditions alter the intracellular homeostasis of zinc, often decreasing zinc levels. These findings implied that there could be a vicious cycle between zinc deficiency and inflammatory conditions. In this review, we present recent evidence on the involvement of zinc in atopic dermatitis, bronchial asthma, and chronic rhinosinusitis, with insights into the involvement of zinc in the underlying molecular and cellular mechanisms related to these allergic inflammatory diseases.
• A substrate-trapping strategy to find E3 ubiquitin ligase substrates identifies Parkin and TRIM28 targets.

  Masashi Watanabe, Yasushi Saeki, Hidehisa Takahashi, Fumiaki Ohtake, Yukiko Yoshida, Yusuke Kasuga, Takeshi Kondo, Hiroaki Yaguchi, Masanobu Suzuki, Hiroki Ishida, Keiji Tanaka, Shigetsugu Hatakeyama

  Communications biology 3 (1) 592 - 592 2020/10/20 [Refereed]
   

  The identification of true substrates of an E3 ligase is biologically important but biochemically difficult. In recent years, several techniques for identifying substrates have been developed, but these approaches cannot exclude indirect ubiquitination or have other limitations. Here we develop an E3 ligase substrate-trapping strategy by fusing a tandem ubiquitin-binding entity (TUBE) with an anti-ubiquitin remnant antibody to effectively identify ubiquitinated substrates. We apply this method to one of the RBR-type ligases, Parkin, and to one of the RING-type ligases, TRIM28, and identify previously unknown substrates for TRIM28 including cyclin A2 and TFIIB. Furthermore, we find that TRIM28 promotes cyclin A2 ubiquitination and degradation at the G1/S phase and suppresses premature entry into S phase. Taken together, the results indicate that this method is a powerful tool for comprehensively identifying substrates of E3 ligases.
• The role of Mediator and Little Elongation Complex in transcription termination.

  Hidehisa Takahashi, Amol Ranjan, Shiyuan Chen, Hidefumi Suzuki, Mio Shibata, Tomonori Hirose, Hiroko Hirose, Kazunori Sasaki, Ryota Abe, Kai Chen, Yanfeng He, Ying Zhang, Ichigaku Takigawa, Tadasuke Tsukiyama, Masashi Watanabe, Satoshi Fujii, Midori Iida, Junichi Yamamoto, Yuki Yamaguchi, Yutaka Suzuki, Masaki Matsumoto, Keiichi I Nakayama, Michael P Washburn, Anita Saraf, Laurence Florens, Shigeo Sato, Chieri Tomomori-Sato, Ronald C Conaway, Joan W Conaway, Shigetsugu Hatakeyama

  Nature communications 11 (1) 1063 - 1063 2020/02/26 [Refereed][Not invited]
   

  Mediator is a coregulatory complex that regulates transcription of Pol II-dependent genes. Previously, we showed that human Mediator subunit MED26 plays a role in the recruitment of Super Elongation Complex (SEC) or Little Elongation Complex (LEC) to regulate the expression of certain genes. MED26 plays a role in recruiting SEC to protein-coding genes including c-myc and LEC to small nuclear RNA (snRNA) genes. However, how MED26 engages SEC or LEC to regulate distinct genes is unclear. Here, we provide evidence that MED26 recruits LEC to modulate transcription termination of non-polyadenylated transcripts including snRNAs and mRNAs encoding replication-dependent histone (RDH) at Cajal bodies. Our findings indicate that LEC recruited by MED26 promotes efficient transcription termination by Pol II through interaction with CBC-ARS2 and NELF/DSIF, and promotes 3' end processing by enhancing recruitment of Integrator or Heat Labile Factor to snRNA or RDH genes, respectively.
• Loss of TRIM29 Alters Keratin Distribution to Promote Cell Invasion in Squamous Cell Carcinoma.

  Teruki Yanagi, Masashi Watanabe, Hiroo Hata, Shinya Kitamura, Keisuke Imafuku, Hiroko Yanagi, Akihiro Homma, Lei Wang, Hidehisa Takahashi, Hiroshi Shimizu, Shigetsugu Hatakeyama

  Cancer research 78 (24) 6795 - 6806 0008-5472 2018/12/15 [Refereed][Not invited]
   

  : TRIM29 (tripartite motif-containing protein 29) is a TRIM family protein that has been implicated in breast, colorectal, and pancreatic cancers. However, its role in stratified squamous epithelial cells and tumors has not been elucidated. Here, we investigate the expression of TRIM29 in cutaneous head and neck squamous cell carcinomas (SCC) and its functions in the tumorigenesis of such cancers. TRIM29 expression was lower in malignant SCC lesions than in adjacent normal epithelial tissue or benign tumors. Lower expression of TRIM29 was associated with higher SCC invasiveness. Primary tumors of cutaneous SCC showed aberrant hypermethylation of TRIM29. Depletion of TRIM29 increased cancer cell migration and invasion; conversely, overexpression of TRIM29 suppressed these. Comprehensive proteomics and immunoprecipitation analyses identified keratins and keratin-interacting protein FAM83H as TRIM29 interactors. Knockdown of TRIM29 led to ectopic keratin localization of keratinocytes. In primary tumors, lower TRIM29 expression correlated with the altered expression of keratins. Our findings reveal an unexpected role for TRIM29 in regulating the distribution of keratins, as well as in the migration and invasion of SCC. They also suggest that the TRIM29-keratin axis could serve as a diagnostic and prognostic marker in stratified epithelial tumors and may provide a target for SCC therapeutics. SIGNIFICANCE: These findings identify TRIM29 as a novel diagnostic and prognostic marker in stratified epithelial tissues.
• Brain-Derived Neurotrophic Factor Improves Limited Exercise Capacity in Mice With Heart Failure.

  Junichi Matsumoto, Shingo Takada, Shintaro Kinugawa, Takaaki Furihata, Hideo Nambu, Naoya Kakutani, Masaya Tsuda, Arata Fukushima, Takashi Yokota, Shinya Tanaka, Hidehisa Takahashi, Masashi Watanabe, Shigetsugu Hatakeyama, Masaki Matsumoto, Keiichi I Nakayama, Yutaro Otsuka, Hisataka Sabe, Hiroyuki Tsutsui, Toshihisa Anzai

  Circulation 138 (18) 2064 - 2066 0009-7322 2018/10/30 [Refereed][Not invited]
  
• Anti-Sez6l2 antibody detected in a patient with immune-mediated cerebellar ataxia inhibits complex formation of GluR1 and Sez6l2.

  Hiroaki Yaguchi, Ichiro Yabe, Hidehisa Takahashi, Masashi Watanabe, Taichi Nomura, Takahiro Kano, Masahiko Watanabe, Shigetsugu Hatakeyama

  Journal of neurology 265 (4) 962 - 965 1432-1459 2018/04 [Refereed][Not invited]
  
• Mutations in bassoon in individuals with familial and sporadic progressive supranuclear palsy-like syndrome.

  Ichiro Yabe, Hiroaki Yaguchi, Yasutaka Kato, Yasuo Miki, Hidehisa Takahashi, Satoshi Tanikawa, Shinichi Shirai, Ikuko Takahashi, Mari Kimura, Yuka Hama, Masaaki Matsushima, Shinsuke Fujioka, Takahiro Kano, Masashi Watanabe, Shin Nakagawa, Yasuyuki Kunieda, Yoshio Ikeda, Masato Hasegawa, Hiroshi Nishihara, Toshihisa Ohtsuka, Shinya Tanaka, Yoshio Tsuboi, Shigetsugu Hatakeyama, Koichi Wakabayashi, Hidenao Sasaki

  Scientific reports 8 (1) 819 - 819 2018/01/16 [Refereed][Not invited]
   

  Clinical diagnosis of progressive supranuclear palsy (PSP) is sometimes difficult because various phenotypes have been identified. Here, we report a mutation in the bassoon (BSN) gene in a family with PSP-like syndrome. Their clinical features resembled not only those of PSP patients but also those of individuals with multiple system atrophy and Alzheimer's disease. The neuropathological findings showed a novel three + four repeat tauopathy with pallido-luysio-nigral degeneration and hippocampal sclerosis. Whole-exome analysis of this family identified a novel missense mutation in BSN. Within the pedigree, the detected BSN mutation was found only in affected individuals. Further genetic analyses were conducted in probands from four other pedigrees with PSP-like syndrome and in 41 sporadic cases. Three missense mutations in BSN that are very rarely listed in databases of healthy subjects were found in four sporadic cases. Western blot analysis of tau following the overexpression of wild-type or mutated BSN revealed the possibility that wild-type BSN reduced tau accumulation, while mutated BSN lost this function. An association between BSN and neurological diseases has not been previously reported. Our results revealed that the neurodegenerative disorder associated with the original proband's pedigree is a novel tauopathy, differing from known dementia and parkinsonism syndromes, including PSP.
• Fine-tuning of thymocyte development by ubiquitination-mediated stability control of the ESCRT protein CHMP5

  Masashi Watanabe, Shigetsugu Hatakeyama

  CELLULAR & MOLECULAR IMMUNOLOGY 14 (12) 957 - 959 1672-7681 2017/12 [Refereed][Not invited]
  
• Sez6l2 regulates phosphorylation of ADD and neuritogenesis

  Hiroaki Yaguchi, Ichiro Yabe, Hidehisa Takahashi, Masashi Watanabe, Taichi Nomura, Takahiro Kano, Masaki Matsumoto, Keiichi I. Nakayama, Masahiko Watanabe, Shigetsugu Hatakeyama

  BIOCHEMICAL AND BIOPHYSICAL RESEARCH COMMUNICATIONS 494 (1-2) 234 - 241 0006-291X 2017/12 [Refereed][Not invited]
   

  Increasing evidence shows that immune-mediated mechanisms may contribute to the pathogenesis of central nervous system disorders including cerebellar ataxias, as indicated by the aberrant production of neuronal surface antibodies. We previously reported a patient with cerebellar ataxia associated with production of a new anti-neuronal antibody, anti-seizure-related 6 homolog like 2 (Sez6l2). Sez6l2 is a type 1 membrane protein that is highly expressed in the hippocampus and cerebellar cortex and mice lacking Sez6l2 protein family members develop ataxia. Here we used a proteomics-based approach to show that serum derived from this patient recognizes the extracellular domain of Sez6l2 and that Sez6l2 protein binds to both adducin (ADD) and glutamate receptor 1 (GluR1). Our results indicate that Sez6l2 is one of the auxiliary subunits of the AMPA receptor and acts as a scaffolding protein to link GluR1 to ADD. Furthermore, Sez6l2 overexpression upregulates ADD phosphorylation, whereas siRNA-mediated downregulation of Sez6l2 prevents ADD phosphorylation, suggesting that Sez6l2 modulates AMPAADD signal transduction. (C) 2017 Elsevier Inc. All rights reserved.
• TRIM proteins and diseases

  Masashi Watanabe, Shigetsugu Hatakeyama

  JOURNAL OF BIOCHEMISTRY 161 (2) 135 - 144 0021-924X 2017/02 [Refereed][Not invited]
   

  Ubiquitination is one of the posttranslational modifications that regulates a number of intracellular events including signal transduction, protein quality control, transcription, cell cycle, apoptosis and development. The ubiquitin system functions as a garbage machine to degrade target proteins and as a regulator for several signalling pathways. Biochemical reaction of ubiquitination requires several enzymes including E1, E2 and E3, and E3 ubiquitin ligases play roles as receptors for recognizing target proteins. Most of the tripartite motif (TRIM) proteins are E3 ubiquitin ligases. Recent studies have shown that some TRIM proteins function as important regulators for a variety of diseases including cancer, inflammatory diseases, infectious diseases, neuropsychiatric disorders, chromosomal abnormalities and developmental diseases. In this review, we summarize the involvement of TRIM proteins in the aetiology of various diseases.
• The novel heart-specific RING finger protein 207 is involved in energy metabolism in cardiomyocytes

  Wataru Mizushima, Hidehisa Takahashi, Masashi Watanabe, Shintaro Kinugawa, Shouji Matsushima, Shingo Takada, Takashi Yokota, Takaaki Furihata, Junichi Matsumoto, Masaya Tsuda, Ikuru Chiba, Shun Nagashima, Shigeru Yanagi, Masaki Matsumoto, Keiichi I. Nakayama, Hiroyuki Tsutsui, Shigetsugu Hatakeyama

  JOURNAL OF MOLECULAR AND CELLULAR CARDIOLOGY 100 43 - 53 0022-2828 2016/11 [Refereed][Not invited]
   

  A failing heart shows severe energy insufficiency, and it is presumed that this energy shortage plays a critical role in the development of cardiac dysfunction. However, little is known about the mechanisms that cause energy metabolic alterations in the failing heart. Here, we show that the novel RING-finger protein 207 (RNF207), which is specifically expressed in the heart, plays a role in cardiac energy metabolism. Depletion of RNF207 in neonatal rat cardiomyocytes (NRCs) leads to a reduced cellular concentration of adenosine triphosphate (ATP) and mitochondrial dysfunction. Consistent with this result, we observed here that the expression of RNF207 was significantly reduced in mice with common cardiac diseases including heart failure. Intriguingly, proteomic approaches revealed that RNF207 interacts with the voltage-dependent anion channel (VDAC), which is considered to be a key regulator of mitochondria function, as an RNF207-interacting protein. Our findings indicate that RNF207 is involved in ATP production by cardiomyocytes, suggesting that RNF207 plays an important role in the development of heart failure. (C) 2016 Elsevier Ltd. All rights reserved.
• p53 represses the transcription of snRNA genes by preventing the formation of little elongation complex

  Delnur Anwar, Hidehisa Takahashi, Masashi Watanabe, Masanobu Suzuki, Satoshi Fukuda, Shigetsugu Hatakeyama

  BIOCHIMICA ET BIOPHYSICA ACTA-GENE REGULATORY MECHANISMS 1859 (8) 975 - 982 1874-9399 2016/08 [Refereed][Not invited]
   

  The regulation of transcription by RNA polymerase II (Pol II) is important for a variety of cellular functions. ELL/EAF-containing little elongation complex (LEC) was found to be required for transcription of Pol II-dependent small nuclear RNA (snRNA) genes. It was shown that the tumor suppressor p53 interacts with ELL and inhibits transcription elongation activity of ELL Here, we show that p53 inhibits interaction between ELL/EAF and ICE1 in LEC and thereby p53 represses transcription of Pol II-dependent snRNA genes through inhibiting LEC function. Furthermore, induction of p53 expression by ultraviolet (UV) irradiation decreases the occupancy of ICE1 at Pol II-dependent snRNA genes. Consistent with the results, knockdown of p53 increased both the expression of snRNA genes and the occupancy of Pol II and components of LEC at snRNA genes. Our results indicate that p53 interferes with the interaction between ELL/EAF and ICE1 and represses transcription of snRNA genes by Pol II. (C) 2016 Elsevier B.V. All rights reserved.
• TRIM39 negatively regulates the NF kappa B-mediated signaling pathway through stabilization of Cactin

  Masanobu Suzuki, Masashi Watanabe, Yuji Nakamaru, Dai Takagi, Hidehisa Takahashi, Satoshi Fukuda, Shigetsugu Hatakeyama

  CELLULAR AND MOLECULAR LIFE SCIENCES 73 (5) 1085 - 1101 1420-682X 2016/03 [Refereed][Not invited]
   

  NF kappa B is one of the central regulators of cell survival, immunity, inflammation, carcinogenesis and organogenesis. The activation of NF kappa B is strictly regulated by several posttranslational modifications including phosphorylation, neddylation and ubiquitination. Several types of ubiquitination play important roles in multi-step regulations of the NF kappa B pathway. Some of the tripartite motif-containing (TRIM) proteins functioning as E3 ubiquitin ligases are known to regulate various biological processes such as inflammatory signaling pathways. One of the TRIM family proteins, TRIM39, for which the gene has single nucleotide polymorphisms, has been identified as one of the genetic factors in Behcet's disease. However, the role of TRIM39 in inflammatory signaling had not been fully elucidated. In this study, to elucidate the function of TRIM39 in inflammatory signaling, we performed yeast two-hybrid screening using TRIM39 as a bait and identified Cactin, which has been reported to inhibit NF kappa B- and TLR-mediated transcriptions. We show that TRIM39 stabilizes Cactin protein and that Cactin is upregulated after TNF alpha stimulation. TRIM39 knockdown also causes activation of the NF kappa B signal. These findings suggest that TRIM39 negatively regulates the NF kappa B signal in collaboration with Cactin induced by inflammatory stimulants such as TNF alpha.
• The E3 ubiquitin ligase TRIM23 regulates adipocyte differentiation via stabilization of the adipogenic activator PPAR gamma

  Masashi Watanabe, Hidehisa Takahashi, Yasushi Saeki, Takashi Ozaki, Shihori Itoh, Masanobu Suzuki, Wataru Mizushima, Keiji Tanaka, Shigetsugu Hatakeyama

  ELIFE 4 2050-084X 2015/04 [Refereed][Not invited]
   

  Adipocyte differentiation is a strictly controlled process regulated by a series of transcriptional activators. Adipogenic signals activate early adipogenic activators and facilitate the transient formation of early enhanceosomes at target genes. These enhancer regions are subsequently inherited by late enhanceosomes. PPAR gamma is one of the late adipogenic activators and is known as a master regulator of adipogenesis. However, the factors that regulate PPAR gamma expression remain to be elucidated. Here, we show that a novel ubiquitin E3 ligase, tripartite motif protein 23 (TRIM23), stabilizes PPAR gamma protein and mediates atypical polyubiquitin conjugation. TRIM23 knockdown caused a marked decrease in PPAR gamma protein abundance during preadipocyte differentiation, resulting in a severe defect in late adipogenic differentiation, whereas it did not affect the formation of early enhanceosomes. Our results suggest that TRIM23 plays a critical role in the switching from early to late adipogenic enhanceosomes by stabilizing PPAR gamma protein possibly via atypical polyubiquitin conjugation.
• MED26 regulates the transcription of snRNA genes through the recruitment of little elongation complex

  Hidehisa Takahashi, Ichigaku Takigawa, Masashi Watanabe, Delnur Anwar, Mio Shibata, Chieri Tomomori-Sato, Shigeo Sato, Amol Ranjan, Chris W. Seidel, Tadasuke Tsukiyama, Wataru Mizushima, Masayasu Hayashi, Yasuyuki Ohkawa, Joan W. Conaway, Ronald C. Conaway, Shigetsugu Hatakeyama

  NATURE COMMUNICATIONS 6 5941  2041-1723 2015/01 [Refereed][Not invited]
   

  Regulation of transcription elongation by RNA polymerase II (Pol II) is a key regulatory step in gene transcription. Recently, the little elongation complex (LEC)-which contains the transcription elongation factor ELL/EAF-was found to be required for the transcription of Pol II-dependent small nuclear RNA (snRNA) genes. Here we show that the human Mediator subunit MED26 plays a role in the recruitment of LEC to a subset of snRNA genes through direct interaction of EAF and the N-terminal domain (NTD) of MED26. Loss of MED26 in cells decreases the occupancy of LEC at a subset of snRNA genes and results in a reduction in their transcription. Our results suggest that the MED26-NTD functions as a molecular switch in the exchange of TBP-associated factor 7 (TAF7) for LEC to facilitate the transition from initiation to elongation during transcription of a subset of snRNA genes.
• Pathology of frontotemporal dementia with limb girdle muscular dystrophy caused by a DNAJB6 mutation

  Ichiro Yabe, Mishie Tanino, Hiroaki Yaguchi, Akihiro Takiyama, Huaying Cai, Hiromi Kanno, Ikuko Takahashi, Yukiko K. Hayashi, Masashi Watanabe, Hidehisa Takahashi, Shigetsugu Hatakeyama, Shinya Tanak, Hidenao Sasaki

  CLINICAL NEUROLOGY AND NEUROSURGERY 127 10 - 12 0303-8467 2014/12 [Refereed][Not invited]
  
• TRIM29 as a novel prostate basal cell marker for diagnosis of prostate cancer

  Yukiko Kanno, Masashi Watanabe, Taichi Kimura, Katsuya Nonomura, Shinya Tanaka, Shigetsugu Hatakeyama

  ACTA HISTOCHEMICA 116 (5) 708 - 712 0065-1281 2014 [Refereed][Not invited]
   

  Tripartite motif protein 29 (TRIM29) is one of the TRIM family proteins, some of which function as E3 ubiquitin ligases. In this study, we investigated the usefulness of TRIM29 for diagnosis of prostate cancer Prostate tissues including carcinoma and non-carcinoma tissues obtained by needle biopsy and radical prostatectomy were used. Immunohistochemistry was performed according to standard procedures using an antibody against TRIM29. Immunohistochemical staining with an antibody against 34 beta E12, which recognizes cytokeratins 1, 5, 10 and 14, was performed as a control. Basal cells of normal prostatic glands were stained with anti-TRIM29 antibody in all cases, whereas prostate cancer tissues had no or little staining with anti-TRIM29 antibody. TRIM29 is selectively expressed in basal cells of the normal prostate gland, and immunohistochemical staining with anti-TRIM29 antibody showed the same expression pattern as that with 34 beta E12 in prostate cancer and its benign mimics. Our data indicate that TRIM29 may be useful for distinguishing prostate cancers from benign tissues. (C) 2013 Elsevier GmbH. All rights reserved.
• TRIM59 interacts with ECSIT and negatively regulates NF-kappa B and IRF-3/7-mediated signal pathways

  Takeshi Kondo, Masashi Watanabe, Shigetsugu Hatakeyama

  BIOCHEMICAL AND BIOPHYSICAL RESEARCH COMMUNICATIONS 3 422 (3) 501 - 507 0006-291X 2012/06 [Refereed][Not invited]
   

  Innate immune responses are triggered by pathogen-associated molecular patterns (PAMPs) through pattern recognition receptors (PRRs) and then activate intracellular signaling pathways including NF-kappa B and interferon regulatory factors. Recently, it has been reported that tripartite motif (TRIM) proteins function as crucial regulators via ubiquitin-mediated modifications for these signaling pathways. In this study, we showed that one of the TRIM family ubiquitin ligases, TRIM59, interacts with ECSIT as an adaptor protein required for the TLR-mediated transduction pathway. Luciferase reporter assays using reporter plasmids including NF-kappa B responsive element, interferon beta (IFN-beta) promoter and interferon-sensitive response element (ISRE) showed that overexpression of TRIM59 repressed their transcriptional activities, whereas knockdown of TRIM59 enhanced their transcriptional activities. Furthermore, TRIM59 inhibited phosphorylation and dimerization of IRF3 and IRF7, suggesting that TRIM59 negatively regulates upstream kinases for IRFs. These findings indicate that TRIM59 may serve as a multifunctional regulator for innate immune signaling pathways. (c) 2012 Elsevier Inc. All rights reserved.
• TRIM24 mediates ligand-dependent activation of androgen receptor and is repressed by a bromodomain-containing protein, BRD7, in prostate cancer cells

  Misato Kikuchi, Fumihiko Okumura, Tadasuke Tsukiyama, Masashi Watanabe, Naoto Miyajima, Junji Tanaka, Masahiro Imamura, Shigetsugu Hatakeyama

  BIOCHIMICA ET BIOPHYSICA ACTA-MOLECULAR CELL RESEARCH 1793 (12) 1828 - 1836 0167-4889 2009/12 [Refereed][Not invited]
   

  The androgen receptor (AR) is a ligand-dependent transcription factor that belongs to the family of nuclear receptors, and its activity is regulated by numerous AR coregulators. AR plays an important role in prostate development and cancer. In this study, we found that TRIM24/transcriptional intermediary factor lot (T1F1 alpha), which is known as a ligand-dependent nuclear receptor co-regulator, interacts with AR and enhances transcriptional activity of AR by dihydrotestosterone in prostate cancer cells. We showed that TRIM24 functionally interacts with TIP60, which acts as a coactivator of AR and synergizes with TIP60 in the transactivation of AR. We also showed that TRIM24 binds to bromodomain containing 7 (BRD7), which can negatively regulate cell proliferation and growth. A luciferase assay indicated that BRD7 represses the AR transactivation activity upregulated by TRIM24. These findings indicate that TRIM24 regulates AR-mediated transcription in collaboration with TIP60 and BRD7. (C) 2009 Elsevier B.V. All rights reserved.
• ZNRF1 interacts with tubulin and regulates cell morphogenesis

  Koichi Yoshida, Masashi Watanabe, Shigetsugu Hatakeyama

  BIOCHEMICAL AND BIOPHYSICAL RESEARCH COMMUNICATIONS 389 (3) 506 - 511 0006-291X 2009/11 [Refereed][Not invited]
   

  The ubiquitin-proteasome system has been implicated in neuronal degeneration and regeneration. We demonstrated that overexpression of ZNRF1, which has been identified as a crucial molecule in nerve regeneration, causes morphological changes such as neurite-like elongation. Molecular dissections showed that both the RING finger domain and zinc finger domain are required for morphological changes. Furthermore, we identified beta-tubulin type 2 (Tubb2) as a ZNRF1-binding protein by yeast two-hybrid screening. In vivo binding assay showed that ZNRF1 interacts with Tubb2 and immunofluorescent staining Suggests that ZNRF1 is colocalized with Tubb2. These results suggest that ZNRF1 mediates regulation of neuritogenesis via interaction with tubulin. (C) 2009 Elsevier Inc. All rights reserved.
• TRIM31 interacts with p52(Shc) and inhibits Src-induced anchorage-independent growth

  Masashi Watanabe, Tadasuke Tsukiyama, Shigetsugu Hatakeyama

  BIOCHEMICAL AND BIOPHYSICAL RESEARCH COMMUNICATIONS 388 (2) 422 - 427 0006-291X 2009/10 [Refereed][Not invited]
   

  Tripartite motif-containing protein (TRIM) family proteins are involved in a broad range of biological processes and, consistently, their alterations result in diverse pathological conditions such as genetic diseases, viral infection and cancer development. In this study, we found that one of the TRIM family proteins, TRIM31, is highly expressed in the gastrointestinal tract and interacts with p52(Shc), one of the signal transducers. We also found by a binding assay that almost the whole region other than the RING domain is required for the binding to p52(Shc) but found by pulse-chase analysis that overexpression of TRIM31 does not affect the stability of p52(Shc). Moreover, we found that overexpression of TRIM31 suppresses anchorage-independent cell growth induced by the active form of c-Src. These results suggest that TRIM31 attenuates c-Src signaling via p52(Shc) under anchorage-independent growth conditions and is potentially associated with growth activity of cells in the gastrointestinal tract. (C) 2009 Elsevier Inc. All rights reserved.
• Inhibition of NF-kappa B signaling via tyrosine phosphorylation of Ymer

  Hiroyuki Kameda, Masashi Watanabe, Miyuki Bohgaki, Tadasuke Tsukiyama, Shigetsugu Hatakeyama

  BIOCHEMICAL AND BIOPHYSICAL RESEARCH COMMUNICATIONS 378 (4) 744 - 749 0006-291X 2009/01 [Refereed][Not invited]
   

  Cytoplasmic zinc finger protein A20 functionally dampens inflammatory signals and apoptosis via inhibition of NF-kappa B activation. We have reported that Ymer interacts with A20 and lysine (K)-63-linked polyubiquitin chain and that Ymer inhibits NF-kappa B signaling in collaboration with A20. It has also been reported that Ymer is phosphorylated by EGF stimulation. We found that Ymer was considerably phosphorylated on tyrosine residues also via Src family kinases such as Lck. A luciferase reporter assay showed that mutation of tyrosines on Ymer (YmerY217/279/304F) results in loss of the inhibitory activity for NF-kappa B signaling. Furthermore, a soft agar colony formation assay showed that the combination of SFCY527F and YmerY217/279/304F has no ability for anchorage-independent growth, suggesting that tyrosine phosphorylation of Ymer is important for inhibition of the NF-kappa B-mediated apoptotic pathway. These findings demonstrate that Ymer is likely to be a negative regulator for the NF-kappa B signaling pathway. (C) 2008 Elsevier Inc. All rights reserved
• Involvement of Ymer in suppression of NF-kappaB activation by regulated interaction with lysine-63-linked polyubiquitin chain.

  Bohgaki M, Tsukiyama T, Nakajima A, Maruyama S, Watanabe M, Koike T, Hatakeyama S

  Biochimica et biophysica acta 5 1783 826 - 837 0006-3002 2008/05 [Refereed][Not invited]
  
• Involvement of Ymer in suppression of NF-kappa B activation by regulated interaction with lysine-63-linked polyubiquitin chain

  Miyuki Bohgaki, Tadasuke Tsukiyama, Ayako Nakajima, Satoru Maruyama, Masashi Watanabe, Takao Koike, Shigetsugu Hatakeyama

  BIOCHIMICA ET BIOPHYSICA ACTA-MOLECULAR CELL RESEARCH 1783 (5) 826 - 837 0167-4889 2008/05 [Refereed][Not invited]
   

  It is known that the cytoplasmic zinc finger protein A20 functionally dampens inflammatory signals and apoptosis via inhibition of NF-kappa B activation and biochemically acts as a unique ubiquitin-modifying protein with dcubiquitmating activity and ubiquitin ligase activity. However, the molecular mechanisms of A20-modulated signal transduction that influence normal immune responses or tumor immunity have not been fully elucidated. Using a yeast two-hybrid system to search for proteins interacting with A20, we identified one novel binding protein, Ymer. Ymer, which has been reported to be highly phosphorylated on tyrosine residues via EGF stimulation, bound to lysine (K)-63-linked polyubiquitin chain on receptor-interacting serine/threonine-protein kinase 1 (RIP 1), which is essential for NF-kappa B signaling in collaboration with A20. A luciferase assay showed that NF-kappa B signaling was down-regulated by overexpression of Ymer, whereas knock-down of Ymer up-egulated NF-kappa B signaling even without stimulation. These findings demonstrate that Ymer is likely to be a negative regulator for the NF-kappa B signaling pathway. (C)007 Elsevier B.V All rights reserved.
• Protection of vincristine-induced neuropathy by Wld(S) expression and the independence of the activity of Nmnat1

  Masashi Watanabe, Tadasuke Tsukiyama, Shigetsugu Hatakeyama

  NEUROSCIENCE LETTERS 411 (3) 228 - 232 0304-3940 2007/01 [Refereed][Not invited]
   

  The slow Wallerian degeneration protein (Wld(S)), a fusion protein containing amino-terminal E4B and full-length nicotinamide mononucleotide adenylyltransferase 1 (Nmnat1), delays axon degeneration caused by physical damages, toxins and genetic mutations which result in patients being diagnosed with neurodegenerative diseases. It is still controversial whether the suppression of axonal degeneration by Wld(S) is due to Nmnat1 or other portion. We generated Wld(S) or Nmnat1-overexpressing Neuro2A cell lines, in which neuronal differentiation including neurite elongation can be induced by retinoic acid. The overexpression of Wld(S) delayed the neurite degeneration by vincristine, whereas that of Nmnat1 did not delay it much. Taken together, Nmnat1 is considerably weaker than Wld(S) for protection from toxic injury in vitro, suggesting that amino-terminal region of Wld(S) is likely to be more significant for protection from axonal degeneration. (c) 2006 Elsevier Ireland Ltd. All rights reserved.
• Targeted destruction of c-Myc by an engineered ubiquitin ligase suppresses cell transformation and tumor formation

  S Hatakeyama, M Watanabe, Y Fujii, KL Nakayama

  CANCER RESEARCH 65 (17) 7874 - 7879 0008-5472 2005/09 [Refereed][Not invited]
   

  Given that expression of c-Myc is up-regulated in many human malignancies, targeted inactivation of this oncoprotein is a potentially effective strategy for cancer treatment The ubiquitin-proteasome pathway of protein degradation is highly specific and can be engineered to achieve the elimination of undesirable proteins such as oncogene products. We have now generated a ftision protein (designated Max-U) that is composed both of May, which forms a heterodimer with c-Myc, and of CRW, which is a U box-type ubiquitin ligase (E3). Max-U physically interacted with c-Myc in transfected cells and promoted the ubiquitylation of c-Myc in vitro. It also reduced the stability of c-Myc in vivo, resulting in suppression of transcriptional activity dependent on c-Myc. Expression of Max-U reduced both the abundance of endogenous c-Myc in and the proliferation rate of a Burkitt lymphoma cell line. Furthermore, expression of Max-U but not that of a catalytically inactive mutant thereof markedly inhibited both the anchorage-independent growth in vitro of NIH 3T3 cells that overexpress c-Myc as well as tumor formation by these cells in nude mice. These findings indicate that the targeted destruction of c-Myc by an artificial E3 may represent an effective therapeutic strategy for certain human malignancies.

MISC

• Proteolytic mechanism of the oncogene product PPM1D

  渡部昌, 高橋正樹, 畠山鎮次  日本分子生物学会年会プログラム・要旨集(Web)  46th-  2023
• Identification of a novel protein on MHC class I transcriptional regulation

  應田涼太, 坐間のゆり, XIN Sun, 渡部昌, 畠山鎮次, 小林弘一  日本分子生物学会年会プログラム・要旨集(Web)  45th-  2022
• 免疫介在性小脳失調症における新規自己抗体の検討

  工藤彰彦, 矢口裕章, 阿部恵, 江口克紀, 長井梓, 脇田雅大, 白井慎一, 岩田育子, 松島理明, 水戸泰紀, 田島康敬, 渡部昌, 畠山鎮次, 米田誠, 田中惠子, 矢部一郎  日本神経学会学術大会プログラム・抄録集  63rd-  2022
• TRIMファミリータンパク質と標的タンパク質分解

  渡部 昌, 畠山 鎮次  細胞  53-  (3)  160  -163  2021/03  

  タンパク質を選択的かつ急速に分解する薬剤の開発法として、Specific and Nongenetic Inhibitor of apoptosis proteindependent Protein Eraser(SNIPER)法やPROteolysis TArgeting Chimeras(PROTAC)法が近年注目を集めている。SNIPER/PROTAC法は、言わばユビキチン修飾系を小分子により言わば「ハイジャック」することで目的のタンパク質を分解する手法である。また、筆者らが着目しているTripartite Motif proteins(TRIM)型ユビキチンリガーゼの1つであるTRIM21に基づいたTrim-Away法もまた、新たな手法として注目されている。本稿ではまずユビキチン修飾系を概説し、TRIM型ユビキチンリガーゼのSNIPER/PROTAC法への応用の可能性およびTrim-Away法について紹介したい。(著者抄録)
• 扁平上皮癌におけるTripartite motif‐containing protein 29(TRIM29)の検討

  柳輝希, 秦洋郎, 北村真也, 清水宏, 柳紘子, 本間明宏, 渡部昌, 畠山鎮次  日本皮膚科学会雑誌  129-  (5)  1170  -1170  2019/05/15  [Not refereed][Not invited]
  
• 扁平上皮におけるTripartite motif‐containing protein 29(TRIM29)の機能解析

  柳輝希, 秦洋郎, 北村真也, 今福恵輔, 清水宏, 柳紘子, 本間明宏, 渡部昌, 畠山鎮次  日本皮膚科学会雑誌  129-  (1)  45  -45  2019/01/20  [Not refereed][Not invited]
  
• メディエーター複合体による転写終結制御機構

  高橋秀尚, 柴田美音, 瀧川一学, 渡部昌, 築山忠維, 山本淳一, 山口雄輝, 藤井聡, 飯田緑, RANJAN Amol, SATO Shigeo, TOMOMORI‐SATO Chieri, CONAWAY Joan, CONAWAY Ronald, 畠山鎮次  日本生化学会大会(Web)  90th-  ROMBUNNO.4P2T15‐06(3P‐0635) (WEB ONLY)  -0635)]  2017  [Not refereed][Not invited]
  
• 髄膜炎・臍周囲炎・呼吸障害を呈したOTULIN‐related autoinflammatory syndrome(ORAS)の1例

  植木将弘, 山田雅文, 戸澤雄介, 竹崎俊一郎, ABDRABOU ShimaaSaid MohamedAli, 小林一郎, 有賀正, 渡部昌, 畠山鎮次, 藤田宏明, 岩井一宏, 三宅紀子, 松本直通  日本免疫不全症研究会プログラム・抄録  10th-  12  2017  [Not refereed][Not invited]
  
• ユビキチンリガーゼTRIM23は非定型ポリユビキチン化によるPPARγの安定化を介して脂肪細胞分化を制御する

  渡部昌, 高橋秀尚, 畠山鎮次  日本生化学会大会(Web)  89th-  ROMBUNNO.3T11‐02(3P‐108) (WEB ONLY)  -02(3P  2016/09  [Not refereed][Not invited]
  
• The Novel Heart-specific Ring-finger Protein 207 Regulates Energy Metabolism in Cardiomyocytes

  Wataru Mizushima, Hidehisa Takahashi, Masashi Watanabe, Shintarou Kinugawa, Shouji Matsushima, Shingo Takada, Takashi Yokota, Takaaki Furuhata, Junichi Matsumoto, M. Tsuda, T. Katayama, T. Nakajima, S. Hatakeyama, H. Tsutsui  CIRCULATION RESEARCH  119-  2016/07
• ユビキチンE3リガーゼTRIM23は非定型ポリユビキチン化によるPPARγの安定化を介して脂肪細胞分化を制御する

  渡部昌, 高橋秀尚, 畠山鎮次  日本細胞生物学会大会(Web)  68th-  ROMBUNNO.P1‐14 (WEB ONLY)  -70  2016/05  [Not refereed][Not invited]
  
• ユビキチンリガーゼTRIM23はPPARγの安定化を介して脂肪細胞分化を制御する

  渡部昌, 高橋秀尚, 畠山鎮次  日本内分泌学会雑誌  92-  (1)  201  2016/04/01  [Not refereed][Not invited]
  
• Bechet病疾患感受性遺伝子Tripartite motif protein 39(TRIM39)の機能解析

  鈴木 正宣, 渡部 昌, 中丸 裕爾, 高木 大, 本間 あや, 畠山 鎮次, 福田 諭  耳鼻咽喉科免疫アレルギー  33-  (2)  38  -39  2015/06  [Not refereed][Not invited]
  
• ベーチェット病感受性遺伝子Tripartite motif protein 39(TRIM39)の機能解析

  SUZUKI MASANOBU, SUZUKI MASANOBU, WATANABE MASASHI, NAKAMARU YUJI, TAKAGI DAI, HONMA AYA, HATAKEYAMA SHIGETSUGU, FUKUDA SATOSHI  日本耳鼻咽喉科免疫アレルギー学会プログラム・抄録集  33rd-  82  2015  [Not refereed][Not invited]
  
• メディエーター複合体のサブユニットMED26はLittle elongation complexをリクルートすることでsnRNA遺伝子の転写を制御する

  高橋秀尚, 瀧川一学, 渡部昌, Delnur Anwar, 柴田美音, 佐藤チエリ, 佐藤滋生, Amol Ranjan, Chris W. Seidel, 築山忠維, 水島航, 林正康, 大川恭行, Joan, W. Conaway,Ronal, C. Conaway, 畠山鎮次  北海道医学雑誌  90-  (1)  76  -76  2015  [Not refereed][Not invited]
  
• Ubiquitin modification in otolaryngology

  SUZUKI MASANOBU, SUZUKI MASANOBU, WATANABE MASASHI, NAKAMARU YUJI, TAKAGI DAI, KANO SATOSHI, HONMA AYA, HATAKEYAMA SHIGETSUGU, FUKUDA SATOSHI  耳鼻咽喉科免疫アレルギー(Web)  33-  (3)  185-192 (J-STAGE)  2015  [Not refereed][Not invited]
  
• ベーチェット病感受性遺伝子Tripartite motif protein39(TRIM39)の機能解析

  SUZUKI MASANOBU, WATANABE MASASHI, TAKAGI DAI, NAKAMARU YUJI, HATAKEYAMA SHIGETSUGU, FUKUDA SATOSHI  日本耳鼻咽喉科学会会報  117-  (4)  504  -504  2014/04/20  [Not refereed][Not invited]
  
• Med26はLittle elongation complexをリクルートすることでsmall nuclear RNA遺伝子の発現を制御する

  高橋秀尚, 瀧川一学, 渡部昌, ANWAR Delnur, 柴田美音, 佐藤チエリ, 佐藤滋生, RANJAN Amol, SEIDEL Chris, 築山忠維, 林正康, 大川恭行, CONAWAY Joan, CONAWAY Ronald, 畠山鎮次  日本生化学会大会(Web)  87th-  WEB ONLY 2T15P-16(3P-502)  -16]  2014  [Not refereed][Not invited]
  
• メディエーター複合体による転写伸長制御

  高橋秀尚, 瀧川一学, 渡部昌, ANWAR Delnur, 柴田美音, 佐藤チエリ, 佐藤滋生, RANJAN Amol, SEIDEL Chris W, 築山忠維, 林正康, 大川恭行, CONAWAY Joan, CONAWAY Ronald C, 畠山鎮次  日本分子生物学会年会プログラム・要旨集(Web)  37th-  1W15-3(1P-0237) (WEB ONLY)  2014  [Not refereed][Not invited]
  
• Med26はLittle elongation complexをリクルートすることでsmall nuclear RNA遺伝子の発現を制御する

  TAKAHASHI Hidehisa, 瀧川一学, ANWAR Delnur, 柴田美音, TOMOMORI‐SATO Chieri, SATO Shigeo, RANJAN Amol, SEIDEL Chris, 築山忠維, 渡部昌, 林正康, 大川恭行, CONAWAY Joan, CONAWAY Ronald, 畠山鎮次  日本分子生物学会年会プログラム・要旨集(Web)  36th-  1P-0182 (WEB ONLY)  2013  [Not refereed][Not invited]
  
• ユビキチン修飾系の分子機構と疾患 TRIM型ユビキチンリガーゼと癌―TRIM蛋白質の多様性と特異性

  渡部昌, 畠山鎮次  医学のあゆみ  243-  (6)  508  -514  2012/11/10  [Not refereed][Not invited]
  
• ユビキチンの多彩な生理機能 3.複雑化するユビキチンリガーゼの機能と多様性

  畠山鎮次, 渡部昌  実験医学  29-  (12)  1921  -1932  2011/08/01  [Not refereed][Not invited]
  
• 「筋萎縮性側索硬化症の病態に基づく画期的治療法の開発」神経突起様伸長及び増殖を制御するTRIM‐EN1/2の機能解析

  佐々木秀直, 矢口裕章, 矢口裕章, 奥村文彦, 加納崇裕, 加納崇裕, 渡部昌, 内ケ島基政, 渡邊雅彦, 畠山鎮次  筋萎縮性側索硬化症の病態に基づく画期的治療法の開発 平成22年度 総括研究報告書  59-61  2011  [Not refereed][Not invited]
  
• TRIMファミリー分子によるPPARγシグナルの制御

  渡部昌, 畠山鎮次  生化学  ROMBUNNO.4P-0130  2011  [Not refereed][Not invited]
  

Books etc

• Advances in Medicine and Biology. Volume 120

  Masashi Watanabe, Shigetsugu Hatakeyama (Chapter 3. Ubiquitin-Conjugating Enzymes (E2s))
  Nova Science Publishers 2017 (ISBN: 9781536118070)

Presentations

• 自然免疫シグナルに関わるユビキチンリガーゼ群の網羅的な基質同定と解析  [Invited]

  渡部 昌, 畠山鎮次

  第92回日本生化学会大会  2019/09
• The E3 ubiquitin ligase TRIM23 regulates adipocyte differentiation via stabilization of the adipogenic activator PPARgamma  [Invited]

  Masashi Watanabe

  第39回日本分子生物学会年会  2016/12

Association Memberships

• PHYSIOLOGICAL SOCIETY OF JAPAN   THE MOLECULAR BIOLOGY SOCIETY OF JAPAN   THE JAPANESE BIOCHEMICAL SOCIETY   

Research Projects

• 新規に開発した基質同定法を用いたがんドライバーユビキチンリガーゼの機能解析

  日本学術振興会：科学研究費助成事業

  Date (from‐to) : 2021/04 -2024/03 

  Author : 渡部 昌, 近藤 豪

   

  (1)がん関連ユビキチンリガーゼ基質同定プローブの作製と安定発現細胞株の樹立：解析対象の１８種類のユビキチンリガーゼ遺伝子を入手し、基質を捕獲するためのユビキチンリガーゼプローブをレトロウイルス発現ベクター上に組み込んだ。今年度は変異していない野生型のユビキチンリガーゼ遺伝子について作製を行った。作製したプローブベクターを用いてレトロウイルスを作製し、プローブを安定に発現する細胞株作製を試みたところ、１３種類のプローブについて作製に成功した。残り５種については細胞毒性のために作製できなかった。 (2) イオントラップ・オービトラップ型質量分析器による基質・結合分子同定：樹立した１３種類の細胞からユビキチン化タンパク質を精製し、高感度質量分析計にて網羅的な同定を行った。具体的には、それぞれの細胞を可溶化し、抗FLAG抗体で免疫沈降を行い、第一段階の精製を行った。トリクロロ酢酸を用いて沈殿後、アセトンによる洗浄、乾燥、ジチオスレイトールにて還元、ヨードアセタミドにてアルキル化を行った後にトリプシンにてペプチドへと分解後、脱塩処理を行った。さらに抗ユビキチンレムナント抗体で再度免疫沈降を行い、第二段階の精製を行い、再度脱塩処理を行った。得られたサンプルについて質量分析を行った。実験は全て３回繰り返した。得られた結果は、過去に我々が行い蓄積している同様の基質同定結果と比較してスコアを算出することで、個々のユビキチンリガーゼ特異的な基質候補を抽出した。抗ユビキチンレムナント抗体について、これまではアガロースビーズに結合したものを使用していたが、今年度はマグネットビーズに結合した抗体についても検討を行い、アガロースビーズと同等またはそれ以上の結果が得られることを確認した。
• Comprehensive identification of polyubiquitinated substrates for elucidation of the "ubiquitin code"

  Japan Society for the Promotion of Science：Grants-in-Aid for Scientific Research

  Date (from‐to) : 2019/06 -2022/03 

  Author : watanabe masashi

   

  Reversible modification of proteins by ubiquitin is one of the important post-translational modifications that support various life phenomena. In recent years, it has been focused on elucidating the relationship between the diverse polyubiquitin chain and its function via the seven lysine residues and amino-terminal methionine in ubiquitin itself. In this study, we developed several probes that efficiently capture polyubiquitin chains and verified their chain specificity. As a result, we obtained several probes that retain various specificities.
• Establishment of method for identification of ubiquitinated protein

  Japan Society for the Promotion of Science：Grants-in-Aid for Scientific Research

  Date (from‐to) : 2017/06 -2019/03 

  Author : Hatakeyama Shigetsugu, TAKAHASHI Hidehisa, WATANABE Masashi

   

  Ubiquitination is a chemical modification that controls protein degradation and is one of the most important post-translational modifications that support many biological phenomena. The reaction system is mediated by ubiquitin ligases (E3) selectively recognizing substrate proteins. Therefore, it is important to identify specific substrates of each E3 and to determine the ubiquitination site of substrate proteins in understanding the biological phenomenon that they are associated with. However, due to various problems, it is still far from standardizing ubiquitinated substrate identification methods. In this study, we challenged to establish a novel technology using TUBE (artificial protein exhibiting high affinity binding to polyubiquitin chain) and ubiquitin remnant antibody allows comprehensive application to various types of ubiquitin ligase (E3) and the E3-substrates relationship was analyzed.
• ゲノムに局在するユビキチン様修飾分子Ufm1による生活習慣病防御機構の解明

  文部科学省：科学研究費補助金(若手研究(A))

  Date (from‐to) : 2016 -2019 

  Author : 渡部昌
• Regulation of signal transduction by TRIM family proteins

  Japan Society for the Promotion of Science：Grants-in-Aid for Scientific Research

  Date (from‐to) : 2015/04 -2018/03 

  Author : Hatakeyama Shigetsugu, TAKAHASHI Hidehisa, WATANABE Masashi

   

  Our previous studies have shown the importance of TRIM family ubiquitin ligase in intracellular signal transduction. Especially, we showed that TRIM family ubiquitin ligase regulates signal transduction involved in cell proliferation and differentiation. Therefore, in this application, we identified and analyzed TRIM family ubiquitin ligases which regulates the ubiquitination in various intracellular signalling pathways. Furthermore, we identified and analyzed candidate genes regulating metabolism and immune signal by comprehensive knockdown of TRIM family genes using their siRNA library.
• 自然免疫シグナルにおけるユビキチン化基質の網羅的同定と解析

  文部科学省：科学研究費補助金(新学術領域研究)

  Date (from‐to) : 2017 -2018 

  Author : 渡部昌
• Devolopment of a novel method to identify E3 ligase substrates

  Japan Society for the Promotion of Science：Grants-in-Aid for Scientific Research

  Date (from‐to) : 2015 -2016 

  Author : WATANABE MASASHI

   

  Since ubiquitin ligase (E3) has the substrate specificity of ubiquitination, identifications of specific substrates of each E3 enzymes and of their ubiquitination sites are important for understanding various biological phenomena. In recent years, Toczyski and colleagues have proposed a ligase trap method using a fusion probe of an ubiquitin-binding domain (UBA) and an E3, and Yoshida and colleagues have proposed a TR-TUBE method combining Tandem Ubiquitin Binding Entity (TUBE) and K-εGG-specific antibody. We attempted to develop a more advanced substrate identification method by adding some improvements in their methods and succeeded in improving the sensitivity. By using this method, it is expected that the functional analysis of E3 would be accelerated.
• The E3 ubiquitin ligase TRIM23 regulates adipocyte differentiation via stabilization of the adipogenic activator PPARgamma

  Japan Society for the Promotion of Science：Grants-in-Aid for Scientific Research

  Date (from‐to) : 2013 -2014 

  Author : WATANABE masashi

   

  Adipocyte differentiation is a tightly controlled process regulated by complex transcriptional activators. Multiple adipogenic signals activate early adipogenic activators and facilitate the transient formation of early enhanceosomes at target genes. These enhancer regions are subsequently inherited by the late adipogenic activators, which coordinate the formation of late enhanceosomes. PPARgamma is one of the late adipogenic activators and is considered the master regulator of adipogenesis. We showed that a novel ubiquitin E3 ligase, tripartite motif protein 23 (TRIM23), stabilizes PPARgamma protein and mediates atypical polyubiquitin conjugation. TRIM23 knockdown caused a marked decrease in PPARgamma protein abundance during preadipocyte differentiation, resulting in a severe defect in late adipogenic differentiation, whereas it did not affect the formation of early enhanceosomes. This adipogenic defect is rescued by ectopic expression of PPARgamma.
• ユビキチン化による脂質代謝制御転写因子PPARガンマーの活性化制御機構の解明

  文部科学省：科学研究費補助金(研究活動スタート支援)

  Date (from‐to) : 2012 -2012 

  Author : 渡部 昌

   

  ①PTRIMは直接PPARγをユビキチン化することを解明 申請者は、少なくとも細胞内ではPPARγと結合しユビキチン化を促進するユビキチンリガーゼPTRIMを同定していたが、直接結合することでユビキチン化を促進しているかは明らかではなかった。リコンビナントPTRIM、PPARγを作製し、In vitroユビキチン化アッセイを行ったところ、PTRIMによりPPARγを直接ユビキチン化することを明らかにした。 ②免疫沈降法と質量分析器を用いた新たなTRIM23相互作用タンパク質の探索 FLAG-TRIM23タンパク質を細胞内で発現させ、抗FLAG抗体にてプルダウンし、質量分析器にて相互作用分子を探索し、複数の結合タンパク質を得た。このうち、メディエーター複合体サブユニットのMed24が含まれていた。PTRIMがMed24のリクルートを介してPPARγの活性化を調節している可能性が示唆された。
• 疾患関連TRIMファミリーユビキチンリガーゼ群の網羅的解析

  文部科学省：科学研究費助成事業(特別研究員奨励費)

  Date (from‐to) : 2009 -2011 

  Author : 渡部昌

Industrial Property Rights

• 特願2021-170461:標的タンパク質分解誘導剤を介してユビキチン化されるタンパク質の同定方法  2021年/10/18

  渡部昌, 畠山鎮次, 佐伯泰

Social Contribution

• さっぽろサイエンスフェスティバル2015 in 北大

  Date (from-to) : 2015/12/19

  Role : Lecturer
• 立命館慶祥中学校 中学３年生キャリア教育 北海道大学キャンパスツアー「ヒトの遺伝子の異常によって起きる病気」

  Date (from-to) : 2015/11/18

  Role : Lecturer

  Sponser, Organizer, Publisher  : 立命館慶祥中学校