プロジェクト紹介

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Professor Hiroyasu Nakano

1. Curriculum Vitae

Business address

Department of Biochemistry Toho University School of Medicine 5-21-16 Omori-Nishi, Ota-ku, Tokyo 143-8540 Japan
E-mail: hiroyasu.nakano@med.toho-u.ac.jp
Tel: 81-3-3762-4151 Ext. 2351
FAX: 81-3-5493-5412

中野教授

Education

1978-84
MD, Chiba University School of Medicine, Chiba, Japan
1984-86
Residency, Department of Medicine,
Chiba University School of Medicine, Chiba
1986-89
Research Fellow, Department of Medicine,
The Research Institute of Lung Cancer,
Chiba University School of Medicine, Chiba
1991-95
PhD, Division of Molecular Genetics (Prof. Takashi Saito),
Chiba University Graduate School of Medicine

Professional experience

1995-2001
Assistant Professor, Department of Immunology,
Juntendo University Graduate School of Medicine
2000-2003
An investigator, Precursory Research for Embryonic Science
and Technology (PRESTO), Japan Science and Technology
Corporation (JST)
2001-2014
Associate Professor, Department of Immunology,
Juntendo University Graduate School of Medicine
2014-present
Professor, Department of Biochemistry
Toho University School of Medicine

Honors and Awards

1999
Inohana young investigators’ award,
Chiba University Graduate School of Medicine
2001
Young Investigators’ grants award,
Human Frontier Science Program (HFSP)
2008
Medical Research Encouragement Prize of
The Japan Medical Association

Academic activities

1990-present
Member, Japanese Society for Immunology
1992-present
Member, Japanese Society for Oncology
1994-present
Member, Japanese Society for Molecular Biology
2002-2012
Member, American Society for Biochemistry and Molecular Biology
2003-present
Member, Japanese Society for Biochemistry
2003-present
Board of Councilors, Japanese Society for Immunology
2010-2014
Board of Councilors, Japanese Society for Cell Death
2015-present
Board of Councilors, Japanese Society for Biochemistry
2015-2019
Board of Directors, Japanese Society for Cell Death
2019-present
Chief Director, Japanese Society for Cell Death

Editorial Board Members

2007-2012
Editorial Board Member, Journal of Biological Chemistry
2011-present
Editorial Board Member, PLoS ONE
2015-2019
Editorial Board Member, Journal of Biochemistry

2. Team

Staff members

Soh Yamazaki, PhD, Associated Professor
Kenta Moriwaki, PhD, Associate Professor
Yuichi Tsuchiya, PhD, Assistant Professor
Shin Murai, PhD, Assistant Professor
Takashi Nishina, PhD, Assistant Professor
Takao Seki, PhD, Assistant Professor

Graduate Students

Kenta Kobayashi, PhD student
Ami Semba, PhD student

Undergraduate Students

Yuki Goto

Technical Staff

Sachiko Komazawa-Sakon

Assistant

Wakako Osanai

3. Research Projects

Our lab is interesting in the mechanism how regulated cell death maintains tissue homeostasis under physiological conditions, and also dysregulated cell death contributes to the development of many pathological conditions, and acute and chronic inflammatory diseases. To achieve these goals, we have been working on various murine models, especially focusing on cellular FLICE-inhibitory protein (cFLIP) and interleukin (IL)-11.

(1) Necroptosis and cFLIP

Apoptosis is a prototype of programmed cell death and plays a crucial role in the development of various organs and elimination of unwanted cells. Recent studies have revealed another type of programmed cell death, which is referred to as necroptosis (Nakano, Curr Top Microbiol Immunol 2017).Necroptosis is executed by two related kinases, receptor-interacting kinase (RIPK)1 and RIPK3, and a downstream effector molecule, mixed lineage kinase domain-like (MLKL). Cellular FLICE-inhibitory protein (cFLIP) is a catalytically inactive homolog of the initiator caspase, caspase-8. CFLAR gene encodes two proteins, designated a long form cFLIP (cFLIPL) and a short from cFLIP (cFLIPs) due to an alternative splicing. Since Cflar-deficient mice exhibits embryonic lethality by enhanced apoptosis and necroptosis, it has been unclear whether cFLIP plays a role in maintaining tissue homeostasis. We previously reported that cFLIP is a very unstable protein and degraded rapidly in NF-KB-deficient cells upon TNF stimulation (Nakajima, EMBO J 2006; Oncogene 2008).Moreover, we recently generated conditional Cflar-deficient mice such as hepatocytes, intestinal epithelial cells, and epidermal cells, and reported that cFLIP plays a crucial role in preventing various types of cells from apoptosis and necroptosis (Piao, Sci Signal 2012; Piao, J Allergy Clin Immunol 2019). Using TNF-induced acute liver injury in hepatocyte-specific cFLIP-deficient mice as a model, we have very recently reported that depletion of myeloid cells exacerbated liver injury along with aberrant increase in serum histone H3 in mouse serum (Piao, Hepatology 2016). We are currently working on murine nonalcoholic steatohepatitis (NASH) using hepatocyte-specific cFLIP-deficient mice to test whether enhanced apoptosis of hepatocytes may exacerbate the development of NASH. If that is the case, we would like to identify factor (s) that critically contribute to compensatory proliferation of hepatocytes, ductular reaction, and liver fibrosis.
 A recent study has shown that cFLIPs blocks apoptosis, but promotes necroptosis, whereas cFLIPL blocks both apoptosis and necroptosis. To investigate what kind of cellular responses are induced by necroptosis in vivo, we have generated CFLARs transgenic (Tg) mice. To our surprise, CFLARs Tg mice develop severe ileitis due to enhanced apoptosis and succumb perinatally. Subsequent analysis reveals that necroptosis of intestinal epithelial cells (IECs) activate type 3 innate lymphoid cells that subsequently induce apoptosis of IECs (Shindo, iScience 2019).
 cFLIP is an unstable protein that is degraded by the proteasome/ubiquitin pathway. A previous study reported that an E3 ligase, ITCH might be responsible for degradation of cFLIP. However, it is unclear whether ITCH is a sole E3 ligase for cFLIPL. By using Alpha screening, we have identified Mind-bomb 2 (MIB2) as an E3 ligase that ubiquitylates cFLIPL, but not cFLIPs or caspase 8. MIB2 attaches K48- and K63-typed ubiquitin chains to cFLIPL and does not promote, but rather blocks degradation of cFLIPL. Moreover, cFLIP-deficient cells reconstituted with cFLIPL mutants lacking MIB2-binding region or MIB2-dependent ubiquitylation lysins, show an increase in susceptibility to TNF-induced apoptosis. These results suggest that MIB2-dependent ubiquitylation of cFLIPL is crucial for execution of full anti-apoptotic functions of cFLIPL through preventing high-ordered oligomer formation of cFLIPL and caspase 8. Moreover, TNF-induced autophosphorylation of RIPK1, a central player of TNF-induced apoptosis, is induced in MIB2 KO cells, suggesting that MIB2 suppresses autoactivation of RIPK1 via undetermined mechanisms. Thus, MIB2 prevents TNF-induced apoptosis via RIPK1-dependent and RIPK1-independent manner through ubiquitylation of cFLIPL (Nakabayashi, Commun Biol 2021).
 To further investigate the process of cells undergoing necroptosis and biological consequences of necroptosis, it is crucial to monitor necroptotic cells by live cell imaging. We have developed a fluorescence resonance energy transfer (FRET) biosensor, termed SMART (a sensor for MLKL activation by RIPK3 based on FRET). SMART is composed of a fragment of MLKL and monitors necroptosis, but not apoptosis or necrosis. Mechanistically, SMART monitors plasma membrane translocation of oligomerized MLKL, which is induced by RIPK3 or mutational activation. SMART in combination with imaging of the release of nuclear danger-associated molecular patterns (DAMPs) and Live-Cell Imaging for Secretion activity (LCI-S) reveals two different modes of the release of High Mobility Group Box 1 from necroptotic cells (Murai, Nat Commun 2018). We have recently generated SMART transgenic mice that enable us to necroptotic cells in vivo by multiphoton microscopy.

(2) Oxidative stress and IL-11

Interleukin-11 (IL-11) is a member of the IL-6 family cytokines, and controls various cellular responses, including hematopoiesis, bone development, tissue repair, and carcinogenesis. IL-11 binds to the IL-11 receptor α1 (IL-11Rα1) and gp130 complex and activates the family of signal transducer and activator of transcription (STAT) proteins. We previously reported that IL-11 is produced by hepatocytes in an oxidative stress-dependent manner and ameliorates acetaminophen-induced liver injury (Nishina, Sci Signal 2012). Moreover, oxidative stress-dependent IL-11 production largely depends on a transcription factor, Fra-1 that is upregulated by ERK-dependent phosphorylation of Fra-1.
 We have very recently reported that one electrophile, 1,2-Naphthoquinone (1,2-NQ), induces IL-11 production in vitro and in vivo. We also have found that IL-11 counteracts 1,2-NQ-induced intestinal toxicity through inducing proliferation of intestinal epithelial cells. Further analyses have revealed that NRF2, a critical transcription factor for oxidative stress, promotes translation of Fra-1 gene, thereby upregulating Fra-1 that subsequently induces IL-11 production (Nishina, J Biol Chem 2016). Thus, our present study has revealed an unexpected role for NRF2 in IL-11 production through upregulating Fra-1.
 Accumulating studies have shown that IL-11 plays a crucial role in the development of gastric and colorectal cancer in human and mice. However, the detailed characters of IL-11-producing (IL-11+) cells have not been determined yet. To characterize IL-11+ cells in vivo, we generate Il11 reporter mice. IL-11+ cells appear in the colon in murine tumor and acute colitis models. Il11ra1 or Il11 deletion attenuates the development of colitis-associated colorectal cancer. IL-11+ cells express fibroblast markers and genes associated with cell proliferation and tissue repair. IL-11 induces the activation of colonic fibroblasts and epithelial cells through phosphorylation of STAT3. Human cancer database analysis reveals that the expression of genes enriched in IL-11+ fibroblasts is elevated in human colorectal cancer and correlated with reduced recurrence-free survival. IL-11+ fibroblasts activate both tumor cells and fibroblasts via secretion of IL-11, thereby constituting a feed-forward loop between tumor cells and fibroblasts in the tumor microenvironment (Nishina, Nat Commun 2021). We are now testing whether depletion of IL-11+ cells attenuates the development of colorectal cancers or reduces the numbers and sizes of colorectal cancers after development of tumors in mice.

4. Selected Publications

Original articles

  • 1. Nishina T, Deguchi Y, Ohshima D, Takeda W, Ohtsuka M, Shichino S, Ueha S, Yamazaki S, Kawauchi M, Nakamura E, Nishiyama C, Kojima Y, Adachi-Akahane S, Hasegawa M, Nakayama M, Oshima M, Yagita H, Shibuya K, Mikami T, Inohara N, Matsushima K, Tada N, Nakano H. Interleukin-11-expressing fibroblasts have a unique gene signature correlated with poor prognosis of colorectal cancer. Nat Commun 2021;12:2281.
  • 2. Nakabayashi O, Takahashi H, Moriwaki K, Komazawa-Sakon S, Ohtake F, Murai S, Tsuchiya Y, Koyahara Y, Saeki Y, Yoshida Y, Yamazaki S, Tokunaga F, Sawasaki T, Nakano H. MIND bomb 2 prevents RIPK1 kinase activity-dependent and -independent apoptosis through ubiquitylation of cFLIPL. Commun Biol 2021;4.
  • 3. Torii S, Yamaguchi H, Nakanishi A, Arakawa S, Honda S, Moriwaki K, Nakano H, Shimizu S. Identification of a phosphorylation site on Ulk1 required for genotoxic stress-induced alternative autophagy. Nat Commun 2020;11:1754.
  • 4. Shindo R, Ohmuraya M, Komazawa-Sakon S, Miyake S, Deguchi Y, Yamazaki S, Nishina T, Yoshimoto T, Kakuta S, Koike M, Uchiyama Y, Konishi H, Kiyama H, Mikami T, Moriwaki K, Araki K, Nakano H. Necroptosis of Intestinal Epithelial Cells Induces Type 3 Innate Lymphoid Cell-Dependent Lethal Ileitis. iScience 2019;15:536-551.
  • 5. Piao X, Miura R, Miyake S, Komazawa-Sakon S, Koike M, Shindo R, Takeda J, Hasegawa A, Abe R, Nishiyama C, Mikami T, Yagita H, Uchiyama Y, Nakano H. Blockade of TNF receptor superfamily 1 (TNFR1)-dependent and TNFR1-independent cell death is crucial for normal epidermal differentiation. J Allergy Clin Immunol 2019;143:213-228 e210.
  • 6. Katagiri T, Yamazaki S, Fukui Y, Aoki K, Yagita H, Nishina T, Mikami T, Katagiri S, Shiraishi A, Kimura S, Tateda K, Sumimoto H, Endo S, Kameda H, Nakano H. JunB plays a crucial role in development of regulatory T cells by promoting IL-2 signaling. Mucosal Immunol 2019;12:1104-1117.
  • 7. Murai S, Yamaguchi Y, Shirasaki Y, Yamagishi M, Shindo R, Hildebrand JM, Miura R, Nakabayashi O, Totsuka M, Tomida T, Adachi-Akahane S, Uemura S, Silke J, Yagita H, Miura M, Nakano H. A FRET biosensor for necroptosis uncovers two different modes of the release of DAMPs. Nat Commun 2018;9:4457.
  • 8. Piao X, Yamazaki S, Komazawa-Sakon S, Miyake S, Nakabayashi O, Kurosawa T, Mikami T, Tanaka M, Van Rooijen N, Ohmuraya M, Oikawa A, Kojima Y, Kakuta S, Uchiyama Y, Tanaka M, Nakano H. Depletion of myeloid cells exacerbates hepatitis and induces an aberrant increase in histone H3 in mouse serum. Hepatology 2017;65:237-252.
  • 9. Piao X, Komazawa-Sakon S, Nishina T, Koike M, Piao JH, Ehlken H, Kurihara H, Hara M, Van Rooijen N, Schutz G, Ohmuraya M, Uchiyama Y, Yagita H, Okumura K, He YW, Nakano H. c-FLIP Maintains Tissue Homeostasis by Preventing Apoptosis and Programmed Necrosis. Sci Signal 2012;5:ra93.
  • 10. Nishina T, Komazawa-Sakon S, Yanaka S, Piao X, Zheng DM, Piao JH, Kojima Y, Yamashina S, Sano E, Putoczki T, Doi T, Ueno T, Ezaki J, Ushio H, Ernst M, Tsumoto K, Okumura K, Nakano H. Interleukin-11 links oxidative stress and compensatory proliferation. Sci Signal 2012;5:ra5.
  • 11. Ushio H, Ueno T, Kojima Y, Komatsu M, Tanaka S, Yamamoto A, Ichimura Y, Ezaki J, Nishida K, Komazawa-Sakon S, Niyonsaba F, Ishii T, Yanagawa T, Kominami E, Ogawa H, Okumura K, Nakano H. Crucial role for autophagy in degranulation of mast cells. J Allergy Clin Immunol 2011;127:1267-1276 e1266.
  • 12. Tokunaga F, Nakagawa T, Nakahara M, Saeki Y, Taniguchi M, Sakata S, Tanaka K, Nakano H, Iwai K. SHARPIN is a component of the NF-KB-activating linear ubiquitin chain assembly complex. Nature 2011;471:633-636.
  • 8. Nakajima A, Kojima Y, Nakayama M, Yagita H, Okumura K, Nakano H. Downregulation of c-FLIP promotes caspase-dependent JNK activation and reactive oxygen species accumulation in tumor cells. Oncogene 2008;27:76-84.
  • 10. Nakajima A, Komazawa-Sakon S, Takekawa M, Sasazuki T, Yeh WC, Yagita H, Okumura K, Nakano H. An antiapoptotic protein, c-FLIP(L), directly binds to MKK7 and inhibits the JNK pathway. EMBO J 2006;25:5549-5559.
  • 12. Sakon S, Xue X, Takekawa M, Sasazuki T, Okazaki T, Kojima Y, Piao JH, Yagita H, Okumura K, Doi T, Nakano H. NF-KB inhibits TNF-induced accumulation of ROS that mediate prolonged MAPK activation and necrotic cell death. EMBO J 2003;22:3898-3909.
  • 13. Honda K, Nakano H, Yoshida H, Nishikawa S, Rennert P, Ikuta K, Tamechika M, Yamaguchi K, Fukumoto T, Chiba T, Nishikawa SI. Molecular basis for hematopoietic/mesenchymal interaction during initiation of Peyer's patch organogenesis. J Exp Med 2001;193:621-630.
  • 14. Nakano H, Sakon S, Koseki H, Takemori T, Tada K, Matsumoto M, Munechika E, Sakai T, Shirasawa T, Akiba H, Kobata T, Santee SM, Ware CF, Rennert PD, Taniguchi M, Yagita H, Okumura K. Targeted disruption of Traf5 gene causes defects in CD40- and CD27-mediated lymphocyte activation. Proc Natl Acad Sci U S A 1999;96:9803-9808.
  • 15. Nakano H, Shindo M, Sakon S, Nishinaka S, Mihara M, Yagita H, Okumura K. Differential regulation of IKB kinase alpha and beta by two upstream kinases, NF-KB-inducing kinase and mitogen-activated protein kinase/ERK kinase kinase-1. Proc Natl Acad Sci U S A 1998;95:3537-3542.
  • 16. Nakano H, Oshima H, Chung W, Williams-Abbott L, Ware CF, Yagita H, Okumura K. TRAF5, an activator of NF-KB and putative signal transducer for the lymphotoxin-beta receptor. J Biol Chem 1996;271:14661-14664.

Review

  • 1. Nakano H, Piao X, Shindo R, Komazawa-Sakon S. Cellular FLICE-Inhibitory Protein Regulates Tissue Homeostasis. Curr Top Microbiol Immunol 2017;403:119-141.
  • 2. Nakano H, Nakajima A, Sakon-Komazawa S, Piao JH, Xue X, Okumura K. Reactive oxygen species mediate crosstalk between NF-KB and JNK. Cell Death Differ 2006;13:730-737.
  • 3. Nakano H. Signaling crosstalk between NF-KB and JNK. Trends Immunol 2004;25:402-405.
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