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OKADA Hitoshi

    Department of Medicine　Professor/Senior Staff
Last Updated :2024/04/25

Researcher Information

Degree

  • MD, PhD（Tohoku Univ）

URL

J-Global ID

Research Interests

  • Aging   Metabolism   DNA damage response   Apoptosis   Epigenome   Tumor biology   

Research Areas

  • Life sciences / Laboratory animal science
  • Life sciences / Medical biochemistry
  • Life sciences / Tumor biology
  • Life sciences / Pathobiochemistry

Academic & Professional Experience

  • 2014/04 - Today  Kindai University, Faculty of MedicineDept. of BiochemistryProfessor
  • 2012/07 - 2014/03  University of TorontoDept. of Medical BiophysicsAssociate Professor
  • 2005/05 - 2014/03  Campbell Family Institute for Breast Cancer ResearchStaff Scientist
  • 2005/05 - 2014/03  Ontario Cancer InstituteScientist
  • 2005/11 - 2012/06  University of TorontoDept. of Medical BiophysicsAssistant Professor
  • 1999/03 - 2005/04  Ontario Cancer Institute博士研究員
  • 1998/02 - 1999/02  Columbia University博士研究員
  • 1995/04 - 1998/01  財団法人癌研究会癌研究所細胞生物部嘱託研究員

Education

  • 1991/04 - 1995/03  Tohoku University  大学院  医学研究科
  • 1981/04 - 1987/03  Tohoku University  School of Medicine

Association Memberships

  • THE JAPANESE ASSOCIATION FOR MOLECULAR TARGET THERAPY OF CANCER   American Association for Cancer Research   THE JAPANESE CANCER ASSOCIATION   THE MOLECULAR BIOLOGY SOCIETY OF JAPAN   THE JAPANESE BIOCHEMICAL SOCIETY   

Published Papers

  • Suman Dash; Takeshi Ueda; Akiyoshi Komuro; Masahiko Honda; Ryoichi Sugisawa; Hitoshi Okada
    Cell death & disease 15 (2) 131 - 131 2024/02 
    Pancreatic ductal adenocarcinoma (PDAC) is considered one of the most lethal forms of cancer. Although in the last decade, an increase in 5-year patient survival has been observed, the mortality rate remains high. As a first-line treatment for PDAC, gemcitabine alone or in combination (gemcitabine plus paclitaxel) has been used; however, drug resistance to this regimen is a growing issue. In our previous study, we reported MYC/glutamine dependency as a therapeutic target in gemcitabine-resistant PDAC secondary to deoxycytidine kinase (DCK) inactivation. Moreover, enrichment of oxidative phosphorylation (OXPHOS)-associated genes was a common property shared by PDAC cell lines, and patient clinical samples coupled with low DCK expression was also demonstrated, which implicates DCK in cancer metabolism. In this article, we reveal that the expression of most genes encoding mitochondrial complexes is remarkably upregulated in PDAC patients with low DCK expression. The DCK-knockout (DCK KO) CFPAC-1 PDAC cell line model reiterated this observation. Particularly, OXPHOS was functionally enhanced in DCK KO cells as shown by a higher oxygen consumption rate and mitochondrial ATP production. Electron microscopic observations revealed abnormal mitochondrial morphology in DCK KO cells. Furthermore, DCK inactivation exhibited reactive oxygen species (ROS) reduction accompanied with ROS-scavenging gene activation, such as SOD1 and SOD2. SOD2 inhibition in DCK KO cells clearly induced cell growth suppression. In combination with increased anti-apoptotic gene BCL2 expression in DCK KO cells, we finally reveal that venetoclax and a mitochondrial complex I inhibitor are therapeutically efficacious for DCK-inactivated CFPAC-1 cells in in vitro and xenograft models. Hence, our work provides insight into inhibition of mitochondrial metabolism as a novel therapeutic approach to overcome DCK inactivation-mediated gemcitabine resistance in PDAC patient treatment.
  • Suman Dash; Takeshi Ueda; Akiyoshi Komuro; Hisayuki Amano; Masahiko Honda; Masahito Kawazu; Hitoshi Okada
    Molecular cancer research : MCR 21 (5) 444 - 457 2023/02 
    Pancreatic ductal adenocarcinoma (PDAC) is one of the most life-threatening malignancies. Although the deoxycytidine analog gemcitabine has been used as the first-line treatment for PDAC, the primary clinical challenge arises because of an eventual acquisition of resistance. Therefore, it is crucial to elucidate the mechanisms underlying gemcitabine resistance to improve treatment efficacy. To investigate potential genes whose inactivation confers gemcitabine resistance, we performed CRISPR knockout library screening. We found that DCK deficiency is the primary mechanism of gemcitabine resistance, and the inactivation of CRYBA2, DMBX1, CROT, and CD36 slightly conferred gemcitabine resistance. In particular, gene expression analysis revealed that DCK-knockout (KO) cells displayed a significant enrichment of genes associated with MYC targets, folate/one-carbon metabolism and glutamine metabolism pathways. Evidently, chemically targeting each of these pathways significantly reduced the survival of DCK KO cells. Moreover, the pathways enriched in DCK KO cells represented a trend similar to those in PDAC cell lines and samples of patients with PDAC with low DCK expression. We further observed that short-term treatment of parental CFPAC-1 cells with gemcitabine induces the expression of several genes, which promote synthesis and transport of glutamine in a dose-dependent manner, which suggests glutamine availability as a potential mechanism of escaping drug toxicity in an initial response for survival. Thus, our findings provide insights into novel therapeutic approaches for gemcitabine-resistant PDAC and emphasize the involvement of glutamine metabolism in drug-tolerant persister cells. Implications: Our study revealed the key pathways involved in gemcitabine resistance in PDAC, thus providing potential therapeutic strategies.
  • Takeshi Ueda; Akinori Kanai; Akiyoshi Komuro; Hisayuki Amano; Kazushige Ota; Masahiko Honda; Masahito Kawazu; Hitoshi Okada
    FASEB bioAdvances 3 (12) 1020 - 1033 2021/12 [Refereed]
     
    Epigenetic alterations of chromatin structure affect chromatin accessibility and collaborate with genetic alterations in the development of cancer. Lysine demethylase 4B (KDM4B) has been identified as a JmjC domain-containing epigenetic modifier that possesses histone demethylase activity. Although recent studies have demonstrated that KDM4B positively regulates the pathogenesis of multiple types of solid tumors, the tissue specificity and context dependency have not been fully elucidated. In this study, we investigated gene expression profiles established from clinical samples and found that KDM4B is elevated specifically in acute myeloid leukemia (AML) associated with chromosomal translocation 8;21 [t(8;21)], which results in a fusion of the AML1 and the eight-twenty-one (ETO) genes to generate a leukemia oncogene, AML1-ETO fusion transcription factor. Short hairpin RNA-mediated KDM4B silencing significantly reduced cell proliferation in t(8;21)-positive AML cell lines. Meanwhile, KDM4B silencing suppressed the expression of AML1-ETO-inducible genes, and consistently perturbed chromatin accessibility of AML1-ETO-binding sites involving altered active enhancer marks and functional cis-regulatory elements. Notably, transduction of murine KDM4B orthologue mutants followed by KDM4B silencing demonstrated a requirement of methylated-histone binding modules for a proliferative surge. To address the role of KDM4B in leukemia development, we further generated and analyzed Kdm4b conditional knockout mice. As a result, Kdm4b deficiency attenuated clonogenic potential mediated by AML1-ETO and delayed leukemia progression in vivo. Thus, our results highlight a tumor-promoting role of KDM4B in AML associated with t(8;21).
  • Jianping Guo; Ji Cheng; Nana Zheng; Xiaomei Zhang; Xiaoming Dai; Linli Zhang; Changjiang Hu; Xueji Wu; Qiwei Jiang; Depei Wu; Hitoshi Okada; Pier Paolo Pandolfi; Wenyi Wei
    Advanced science (Weinheim, Baden-Wurttemberg, Germany) 8 (18) e2004303  2021/07 [Refereed]
     
    Copper plays pivotal roles in metabolic homoeostasis, but its potential role in human tumorigenesis is not well defined. Here, it is revealed that copper activates the phosphoinositide 3-kinase (PI3K)-protein kinase B (PKB, also termed AKT) oncogenic signaling pathway to facilitate tumorigenesis. Mechanistically, copper binds 3-phosphoinositide dependent protein kinase 1 (PDK1), in turn promotes PDK1 binding and subsequently activates its downstream substrate AKT to facilitate tumorigenesis. Blocking the copper transporter 1 (CTR1)-copper axis by either depleting CTR1 or through the use of copper chelators diminishes the AKT signaling and reduces tumorigenesis. In support of an oncogenic role for CTR1, the authors find that CTR1 is abnormally elevated in breast cancer, and is subjected by NEDD4 like E3 ubiquitin protein ligase (Nedd4l)-mediated negative regulation through ubiquitination and subsequent degradation. Accordingly, Nedd4l displays a tumor suppressive function by suppressing the CTR1-AKT signaling. Thus, the findings identify a novel regulatory crosstalk between the Nedd4l-CTR1-copper axis and the PDK1-AKT oncogenic signaling, and highlight the therapeutic relevance of targeting the CTR1-copper node for the treatment of hyperactive AKT-driven cancers.
  • Sun-Ju Yi; You-Jee Jang; Hye-Jung Kim; Kyubin Lee; Hyerim Lee; Yeojin Kim; Junil Kim; Seon Young Hwang; Jin Sook Song; Hitoshi Okada; Jae-Il Park; Kyuho Kang; Kyunghwan Kim
    Bone research 9 (1) 27 - 27 2021/05 [Refereed]
     
    Bone undergoes a constant and continuous remodeling process that is tightly regulated by the coordinated and sequential actions of bone-resorbing osteoclasts and bone-forming osteoblasts. Recent studies have shown that histone demethylases are implicated in osteoblastogenesis; however, little is known about the role of histone demethylases in osteoclast formation. Here, we identified KDM4B as an epigenetic regulator of osteoclast differentiation. Knockdown of KDM4B significantly blocked the formation of tartrate-resistant acid phosphatase-positive multinucleated cells. Mice with myeloid-specific conditional knockout of KDM4B showed an osteopetrotic phenotype due to osteoclast deficiency. Biochemical analysis revealed that KDM4B physically and functionally associates with CCAR1 and MED1 in a complex. Using genome-wide chromatin immunoprecipitation (ChIP)-sequencing, we revealed that the KDM4B-CCAR1-MED1 complex is localized to the promoters of several osteoclast-related genes upon receptor activator of NF-κB ligand stimulation. We demonstrated that the KDM4B-CCAR1-MED1 signaling axis induces changes in chromatin structure (euchromatinization) near the promoters of osteoclast-related genes through H3K9 demethylation, leading to NF-κB p65 recruitment via a direct interaction between KDM4B and p65. Finally, small molecule inhibition of KDM4B activity impeded bone loss in an ovariectomized mouse model. Taken together, our findings establish KDM4B as a critical regulator of osteoclastogenesis, providing a potential therapeutic target for osteoporosis.
  • Chen Zhu; Karen O Dixon; Kathleen Newcomer; Guangxiang Gu; Sheng Xiao; Sarah Zaghouani; Markus A Schramm; Chao Wang; Huiyuan Zhang; Kouichiro Goto; Elena Christian; Manu Rangachari; Orit Rosenblatt-Rosen; Hitoshi Okada; Tak Mak; Meromit Singer; Aviv Regev; Vijay Kuchroo
    Science advances 7 (18) 2021/04 [Refereed]
     
    T cell exhaustion has been associated with poor prognosis in persistent viral infection and cancer. Conversely, in the context of autoimmunity, T cell exhaustion has been favorably correlated with long-term clinical outcome. Understanding the development of exhaustion in autoimmune settings may provide underlying principles that can be exploited to quell autoreactive T cells. Here, we demonstrate that the adaptor molecule Bat3 acts as a molecular checkpoint of T cell exhaustion, with deficiency of Bat3 promoting a profound exhaustion phenotype, suppressing autoreactive T cell-mediated neuroinflammation. Mechanistically, Bat3 acts as a critical mTORC2 inhibitor to suppress Akt function. As a result, Bat3 deficiency leads to increased Akt activity and FoxO1 phosphorylation, indirectly promoting Prdm1 expression. Transcriptional analysis of Bat3 -/- T cells revealed up-regulation of dysfunction-associated genes, concomitant with down-regulation of genes associated with T cell effector function, suggesting that absence of Bat3 can trigger T cell dysfunction even under highly proinflammatory autoimmune conditions.
  • Naohiro Oka; Akiyoshi Komuro; Hisayuki Amano; Suman Dash; Masahiko Honda; Kazushige Ota; Shunji Nishimura; Takeshi Ueda; Masao Akagi; Hitoshi Okada
    PHARMACOLOGY RESEARCH & PERSPECTIVES JOHN WILEY & SONS LTD 8 (4) e00632  2052-1707 2020/08 [Refereed]
     
    Osteosarcoma (OS) is the most common malignant bone tumor and a leading cause of cancer-related deaths in children and adolescents. Current standard treatments for OS are a combination of preoperative chemotherapy, surgical resection, and adjuvant chemotherapy. Cisplatin is used as the standard chemotherapeutic for OS treatment, but it induces various adverse effects, limiting its clinical application. Improving treatment efficacy without increasing the cisplatin dosage is desirable. In the present study, we assessed the combined effect of ascorbate on cisplatin treatment using cultured human OS cells. Co-treatment with ascorbate induced greater suppression of OS cell but not nonmalignant cell proliferation. The chemosensitizing effect of ascorbate on cisplatin treatment was tightly linked to ROS production. Altered cellular redox state due to increased ROS production modified glycolysis and mitochondrial function in OS cells. In addition, OS cell sphere formation was markedly decreased, suggesting that ascorbate increased the treatment efficacy of cisplatin against stem-like cells in the cancer cell population. We also found that enhanced MYC signaling, ribosomal biogenesis, glycolysis, and mitochondrial respiration are key signatures in OS cells with cisplatin resistance. Furthermore, cisplatin resistance was reversed by ascorbate. Taken together, our findings provide a rationale for combining cisplatin with ascorbate in therapeutic strategies against OS.
  • Simone F. Glaser; Andreas W. Heumueller; Lukas Tombor; Patrick Hofmann; Marion Muhly-Reinholz; Ariane Fischer; Stefan Guenther; Karoline E. Kokot; Hitoshi Okada; David Hassel; Sandeep Kumar; Hanjoong Jo; Reinier A. Boon; Wesley Abplanalp; David John; Jes-Niels Boeckel; Stefanie Dimmeler
    PROCEEDINGS OF THE NATIONAL ACADEMY OF SCIENCES OF THE UNITED STATES OF AMERICA NATL ACAD SCIENCES 117 (15) 8657 - 8657 0027-8424 2020/04 [Refereed]
  • Kazushige Ota; Akiyoshi Komuro; Hisayuki Amano; Akinori Kenai; Kai Ge; Takeshi Ueda; Hitoshi Okada
    SCIENTIFIC REPORTS NATURE PUBLISHING GROUP 9 (1) 10036 - 10036 2045-2322 2019/07 [Refereed]
     
    Obesity increases the risk of metabolic disorders like diabetes mellitus and dyslipidemia. However, how metabolic status is sensed and regulates cellular behavior is unclear. Utx is an H3K27 demethylase that influences adipocyte function in vitro. To examine its role in vivo, we generated mice lacking Utx in adipocytes (UtxAKO). Although all UtxAKO mice grew normally on a normal chow diet (NCD), female UtxAKO mice on a high fat diet (HFD) showed striking reductions in body fat compared to control mice (Ctrl). Gene expression profiling of adipose tissues of HFD-fed UtxAKO female mice revealed decreased expression of rate-limiting enzymes of triacylglycerol synthesis but increased expression of those of cholesterol/steroid hormone synthesis. Moreover, these animals resisted adiposity induced by ovariectomy and exhibited increased estrogen in visceral adipose tissues. Thus, upon HFD feeding, Utx regulates lipid metabolism in adipose tissues by influencing the local hormonal microenvironment. Conversely, Utx deficiency skews lipid catabolism to enhance cholesterol/steroid hormone production and repress obesity.
  • Keisuke Hitachi; Hidehito Inagaki; Hiroki Kurahashi; Hitoshi Okada; Kunihiro Tsuchida; Masahiko Honda
    Frontiers in sports and active living 1 41 - 41 2019 [Refereed]
  • Changkeun Kang; Kayoko Saso; Kazushige Ota; Masahito Kawazu; Takeshi Ueda; Hitoshi Okada
    GENES TO CELLS WILEY 23 (9) 767 - 777 1356-9597 2018/09 [Refereed]
     
    Obesity is a serious global health issue; however, the roles of genetics and epigenetics in the onset and progression of obesity are still not completely understood. The aim of this study was to determine the role of Kdm4b, which belongs to a subfamily of histone demethylases, in adipogenesis and fat metabolism in vivo. We established conditional Kdm4b knockout mice. Inactivation of Kdm4b in adipocytes (K4bKO) induced profound obesity in mice on a high fat diet (HFD). The HFD-fed K4bKO mice exhibited an increased volume of fat mass and higher expression levels of adipogenesis-related genes. In contrast, the genes involved in energy expenditure and mitochondrial functions were down-regulated. Supporting these findings, the energy expenditure of Kdm4b-deficient cells was markedly decreased. In addition, progression of glucose intolerance and hepatic steatosis with hepatocellular damages was observed. These data indicate that Kdm4b is a critical regulator of systemic metabolism via enhancing energy expenditure in adipocytes.
  • Akiyoshi Komuro; Erna Raja; Caname Iwata; Manabu Soda; Kazunobu Isogaya; Keiko Yuki; Yasushi Ino; Masato Morikawa; Tomoki Todo; Hiroyuki Aburatani; Hiromichi Suzuki; Melissa Ranjit; Atsushi Natsume; Akitake Mukasa; Nobuhito Saito; Hitoshi Okada; Hiroyuki Mano; Kohei Miyazono; Daizo Koinuma
    INTERNATIONAL JOURNAL OF CANCER WILEY 142 (8) 1627 - 1639 0020-7136 2018/04 [Refereed]
     
    Glioblastoma is one of the most malignant forms of cancer, for which no effective targeted therapy has been found. Although The Cancer Genome Atlas has provided a list of fusion genes in glioblastoma, their role in progression of glioblastoma remains largely unknown. To search for novel fusion genes, we obtained RNA-seq data from TGS-01 human glioma-initiating cells, and identified a novel fusion gene (HMGA2-EGFR), encoding a protein comprising the N-terminal region of the high-mobility group AT-hook protein 2 (HMGA2) fused to the C-terminal region of epidermal growth factor receptor (EGFR), which retained the transmembrane and kinase domains of the EGFR. This fusion gene product showed transforming potential and a high tumor-forming capacity in cell culture and in vivo. Mechanistically, HMGA2-EGFR constitutively induced a higher level of phosphorylated STAT5B than EGFRvIII, an in-frame exon deletion product of the EGFR gene that is commonly found in primary glioblastoma. Forced expression of HMGA2-EGFR enhanced orthotopic tumor formation of the U87MG human glioma cell line. Furthermore, the EGFR kinase inhibitor erlotinib blocked sphere formation of TGS-01 cells in culture and inhibited tumor formation in vivo. These findings suggest that, in addition to gene amplification and in-frame exon deletion, EGFR signaling can also be activated by gene fusion, suggesting a possible avenue for treatment of glioblastoma.
  • Kazushige Ota; Kit I. Tong; Kouichiro Goto; Shuta Tomida; Akiyoshi Komuro; Zhong Wang; Kazuto Nishio; Hitoshi Okada
    PLOS ONE PUBLIC LIBRARY SCIENCE 12 (3) e0173713  1932-6203 2017/03 [Refereed]
     
    Understanding the molecular mechanisms that drive adipogenesis is important in developing new treatments for obesity and diabetes. Epigenetic regulations determine the capacity of adipogenesis. In this study, we examined the role of a histone H3 lysine 27 demethylase, the ubiquitously transcribed tetratricopeptide repeat protein on the X chromosome (Utx), in the differentiation of mouse embryonic stem cells (mESCs) to adipocytes. Using gene trapping, we examined Utx-deficient male mESCs to determine whether loss of Utx would enhance or inhibit the differentiation of mESCs to adipocytes. Utx-deficient mESCs showed diminished potential to differentiate to adipocytes compared to that of controls. In contrast, Utx-deficient preadipocytes showed enhanced differentiation to adipocytes. Microarray analyses indicated that the beta-catenin/c-Myc signaling pathway was differentially regulated in Utx-deficient cells during adipocyte differentiation. Therefore, our data suggest that Utx governs adipogenesis by regulating c-Myc in a differentiation stage-specific manner and that targeting the Utx signaling pathway could be beneficial for the treatment of obesity, diabetes, and congenital utx-deficiency disorders.
  • Kit I. Tong; Kazushige Ota; Akiyoshi Komuro; Takeshi Ueda; Akihiko Ito; C. Anne Koch; Hitoshi Okada
    CELL DEATH & DISEASE NATURE PUBLISHING GROUP 7 (10) e2401  2041-4889 2016/10 [Refereed]
     
    Therapy-related cancers are potentially fatal late life complications for patients who received radio-or chemotherapy. So far, the mouse model showing reduction or delay of these diseases has not been described. We found that the disruption of Aplf in mice moderately attenuated DNA damage repair and, unexpectedly, impeded myeloid neoplasms after exposure to ionizing radiation (IR). Irradiated mutant mice showed higher rates of p53-dependent cell death, fewer chromosomal translocations, and a delay in malignancy-induced mortality. Simultaneous deficiency of p53 abrogated IR-induced apoptosis and the benefit of impaired DNA repair on mortality in irradiated Aplf(-/-) mice. Depletion of APLF in non-tumorigenic human cells also markedly reduced the risk of radiation-induced chromosomal aberrations. We therefore conclude that proficient DNA damage repair may promote chromosomal aberrations in normal tissues after irradiation and induce malignant evolution, thus illustrating the potential benefit in sensitizing p53 function by manipulating DNA repair efficiency in cancer patients undergoing genotoxic therapies.
  • Eva Martini; Nadine Wittkopf; Claudia Guenther; Moritz Leppkes; Hitoshi Okada; Alastair J. Watson; Eva Podstawa; Ingo Backert; Kerstin Amann; Markus F. Neurath; Christoph Becker
    CELL REPORTS CELL PRESS 14 (5) 1062 - 1073 2211-1247 2016/02 [Refereed]
     
    A tightly regulated balance of proliferation and cell death of intestinal epithelial cells (IECs) is essential for maintenance of gut homeostasis. Survivin is highly expressed during embryogenesis and in several cancer types, but little is known about its role in adult gut tissue. Here, we show that Survivin is specifically expressed in transit-amplifying cells and Lgr5(+) stem cells. Genetic loss of Survivin in IECs resulted in destruction of intestinal integrity, mucosal inflammation, and death of the animals. Survivin deletion was associated with decreased epithelial proliferation due to defective chromosomal segregation. Moreover, Survivin-deficient animals showed induced phosphorylation of p53 and H2AX and increased levels of cell-intrinsic apoptosis in IECs. Consequently, induced deletion of Survivin in Lgr5(+) stem cells led to cell death. In summary, Survivin is a key regulator of gut tissue integrity by regulating epithelial homeostasis in the stem cell niche.
  • K. Fujiwara; Y. Fujita; A. Kasai; Y. Onaka; H. Hashimoto; H. Okada; T. Yamashita
    Translational Psychiatry NATURE PUBLISHING GROUP 6 (3) e766  2158-3188 2016 [Refereed]
     
    © 2016, Nature Publishing Group. All rights reserved. JMJD2B is a histone demethylase enzyme that regulates gene expression through demethylation of H3K9me3. Although mutations of JMJD2B have been suggested to be responsible for neurodevelopmental disorders, the function of JMJD2B in the central nervous system (CNS) remains to be elucidated. Here we show that JMJD2B has a critical role in the development of the CNS. We observed JMJD2B expression, which was especially strong in the hippocampus, throughout the CNS from embryonic periods through adulthood. We generated neuron-specific JMJD2B-deficient mice using the cre-loxP system. We found an increase in total spine number, but a decrease in mature spines, in the CA1 region of the hippocampus. JMJD2B-deficient mice exhibited hyperactive behavior, sustained hyperactivity in a novel environment, deficits in working memory and spontaneous epileptic-like seizures. Together these observations indicate that JMJD2B mutant mice display symptoms reminiscent of neurodevelopmental disorders. Our findings provide evidence for the involvement of histone demethylation in the formation of functional neural networks during development.
  • Jun Yang; Alaa M. AlTahan; Dongli Hu; Yingdi Wang; Pei-Hsin Cheng; Christopher L. Morton; Chunxu Qu; Amit C. Nathwani; Jason M. Shohet; Theodore Fotsis; Jan Koster; Rogier Versteeg; Hitoshi Okada; Adrian L. Harris; Andrew M. Davidoff
    JNCI-JOURNAL OF THE NATIONAL CANCER INSTITUTE OXFORD UNIV PRESS INC 107 (6) djv080  0027-8874 2015/06 [Refereed]
     
    Background: Epigenetic alterations, such as histone methylation, modulate Myc signaling, a pathway central to oncogenesis. We investigated the role of the histone demethylase KDM4B in N-Myc-mediated neuroblastoma pathogenesis.Methods: Spearman correlation was performed to correlate MYCN and KDM4B expression. RNA interference, microarray analysis, gene set enrichment analysis, and real-time polymerase chain reaction were used to define the functions of KDM4B. Immunoprecipitation and immunofluorescence were used to assess protein-protein interactions between N-Myc and KDM4B. Chromatin immunoprecipitation was used to assess the binding of Myc targets. Constitutive and inducible lentiviral-mediated KDM4B knockdown with shRNA was used to assess the effects on tumor growth. Kaplan-Meier survival analysis was used to assess the prognostic value of KDM4B expression. All statistical tests were two-sided.Results: KDM4B and MYCN expression were found to be statistically significantly correlated in a variety of cancers, including neuroblastoma (R = 0.396, P <.001). Functional studies demonstrated that KDM4B regulates the Myc pathway. N-Myc was found to physically interact with and recruit KDM4B. KDM4B was found to regulate neuroblastoma cell proliferation and differentiation in vitro and xenograft growth in vivo (5 mice/group, two-tailed t test, P <= 0.001). Finally, together with MYCN amplification, KDM4B was found to stratify a subgroup of poor-prognosis patients (122 case patients, P <.001).Conclusions: Our findings provide insight into the epigenetic regulation of Myc via histone demethylation and proof-of-concept for inhibition of histone demethylases to target Myc signaling in cancers such as neuroblastoma.
  • Hitoshi Okada
    ACTA MEDICA KINKI UNIVERSITY Kinki University Medical Association 39 (1) 1 - 10 0386-6092 2014/06 [Invited]
     
    [Abstract] Post-translational modifications of histones are important regulatory mechanisms of numerous biological processes such as gene transcription, DNA damage response, hormonal responses, and metabolism. Recent growing evidence indicates that dysregulation of these modifications has strong impact on onset and progression of human diseases including cancer, immune disorder, and obesity. Here we summarize regulatory mechanisms of histone lysine methylation with special emphasis on how the homeostasis of histone methylations are regulated and how deregulated demethylation process contributes to the pathological conditions. Since epigenetics is tightly linked to numerous biological phenomena, further understanding of regulatory mechanisms underlying epigenetic alterations may identify novel therapeutic intervention of human diseases and, eventually, lead to personalized medicine.
  • Eva Martini; Nadine Wittkopf; Claudia Guenther; Hitoshi Okada; Markus F. Neurath; Christoph Becker
    GASTROENTEROLOGY W B SAUNDERS CO-ELSEVIER INC 146 (5) S86 - S87 0016-5085 2014/05 [Refereed]
  • Eva Martini; Nadine Wittkopf; Claudia Guenther; Hitoshi Okada; Markus F. Neurath; Christoph Becker
    GASTROENTEROLOGY W B SAUNDERS CO-ELSEVIER INC 144 (5) S833 - S833 0016-5085 2013/05 [Refereed]
     
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  • Manu Rangachari; Chen Zhu; Kaori Sakuishi; Sheng Xiao; Jozsef Karman; Andrew Chen; Mathieu Angin; Andrew Wakeham; Edward A. Greenfield; Raymond A. Sobel; Hitoshi Okada; Peter J. McKinnon; Tak W. Mak; Marylyn M. Addo; Ana C. Anderson; Vijay K. Kuchroo
    NATURE MEDICINE NATURE PUBLISHING GROUP 18 (9) 1394 - + 1078-8956 2012/09 [Refereed]
     
    T cell immunoglobulin and mucin domain-containing 3 (Tim-3) is an inhibitory receptor that is expressed on exhausted T cells during infection with HIV-1 and hepatitis C virus. By contrast, Tim-3 expression and function are defective in multiple human autoimmune diseases. However, the molecular mechanisms modulating Tim-3 function are not well understood. Here we show that human leukocyte antigen B (HLA-B)-associated transcript 3 (Bat3) binds to, and represses the function of, Tim-3. Bat3 protects T helper type 1 (T(H)1) cells from galectin-9-mediated cell death and promotes both proliferation and proinflammatory cytokine production. Bat3-deficient T cells have elevated expression of exhaustion-associated molecules such as Tim-3, Lag3, Prdm1 and Pbx3, and Bat3 knockdown in myelin-antigen-specific CD4(+) T cells markedly inhibits the development of experimental autoimmune encephalomyelitis while promoting the expansion of a dysfunctional Tim-3(hi), interferon-gamma (IFN-gamma)(lo)CD4(+) cell population. Furthermore, expression of Bat3 is reduced in exhausted Tim-3(+) T cells from mouse tumors and HIV-1-infected individuals. These data indicate that Bat3 acts as an inhibitor of Tim-3-dependent exhaustion and cell death. Bat3 may thus represent a viable therapeutic target in autoimmune disorders, chronic infections and cancers.
  • Xiaohong Wu; Qinfeng Zhang; Xiaojing Wang; Jiayu Zhu; Kuangfeng Xu; Hitoshi Okada; Rennian Wang; Minna Woo
    PLOS ONE PUBLIC LIBRARY SCIENCE 7 (8) e41976  1932-6203 2012/08 [Refereed]
     
    Aims/Hypothesis: Pancreatic beta-cell mass expands through adulthood under certain conditions. The related molecular mechanisms are elusive. This study was designed to determine whether surviving (also known as Birc5), which is transiently expressed perinatally in islets, was required for beta-cell mass expansion in the pancreatic duct-ligated mouse model.Methods: Mice with beta cell-specific deletion of survivin (RIPCre(+) survivin(fl/fl)) and their control littermates (RIPCre(+) survivin(+/+)) were examined to determine the essential role of survivin in partial pancreatic duct ligation (PDL)-induced beta-cell proliferation, function and survival.Results: Resurgence of survivin expression occurred as early as day 3 post-PDL. By day 7 post-PDL, control mice showed significant expansion of beta-cell mass and increase in beta-cell proliferation and islet number in the ligated tail of the pancreas. However, mice deficient in beta-cell survivin showed a defect in beta-cell mass expansion and proliferation with a marked attenuation in the increase of total islet number, largely due to an impairment in the increase in number of larger islets while sparing the increase in number of small islets in the ligated tail of pancreas, resulting in insufficient insulin secretion and glucose intolerance. Importantly however, beta cell neogenesis and apoptosis were not affected by the absence of survivin in beta cells after PDL.Conclusions/Interpretation: Our results indicate that survivin is essential for beta-cell mass expansion after PDL. Survivin appears to exhibit a preferential requirement for proliferation of preexisting beta cells.
  • Feng Wang; Christopher B. Marshall; Kazuo Yamamoto; Guang-Yao Li; Genevieve M. C. Gasmi-Seabrook; Hitoshi Okada; Tak W. Mak; Mitsuhiko Ikura
    PROCEEDINGS OF THE NATIONAL ACADEMY OF SCIENCES OF THE UNITED STATES OF AMERICA NATL ACAD SCIENCES 109 (16) 6078 - 6083 0027-8424 2012/04 [Refereed]
     
    Forkhead box class O 3a (FOXO3a) is a transcription factor and tumor suppressor linked to longevity that determines cell fate through activating transcription of cell differentiation, survival, and apoptotic genes. Recruitment of the coactivator CBP/p300 is a crucial step in transcription, and we revealed that in addition to conserved region 3 (CR3) of FOXO3a, the C-terminal segment of CR2 (CR2C) binds CBP/p300 and contributes to transcriptional activity. CR2C and CR3 of FOXO3a interact with the KIX domain of CBP/p300 at both "MLL" and "c-Myb" binding sites simultaneously. A FOXO3a CR2C-CR3 peptide in complex with KIX exists in equilibrium between two equally populated conformational states, one of which has CR2C bound to the MLL site and CR3 bound to the c-Myb site, whereas in the other, CR2C and CR3 bind the c-Myb and MLL sites, respectively. This promiscuous interaction between FOXO3a and CBP/p300 is further supported by additional binding sites on CBP/p300, namely, the TAZ1 and TAZ2 domains. In functional studies, our structure-guided mutagenesis showed that both CR2C and CR3 are involved in the activation of certain endogenous FOXO3a target genes. Further, phosphorylation of S626, a known AMP-dependent protein kinase target in CR3, increased affinity for CBP/p300 and the phosphomimetic mutation enhanced transactivation of luciferase. These findings underscore the significance of promiscuous multivalent interactions and posttranslational modification in the recruitment of transcriptional coactivators, which may allow transcription factors to adapt to various gene-specific genomic and chromatin structures and respond to cell signals.
  • K. Goto; K. I. Tong; J. Ikura; H. Okada
    Cell Death and Disease NATURE PUBLISHING GROUP 2 (12) e236  2041-4889 2011/12 [Refereed]
     
    Members of the transforming growth factor-β (TGF-β) superfamily participate in numerous biological phenomena in multiple tissues, including in cell proliferation, differentiation, and migration. TGF-β superfamily proteins therefore have prominent roles in wound healing, fibrosis, bone formation, and carcinogenesis. However, the molecular mechanisms regulating these signaling pathways are not fully understood. Here, we describe the regulation of bone morphogenic protein (BMP) signaling by Bat3 (also known as Scythe or BAG6). Bat3 overexpression in murine cell lines suppresses the activity of the Id1 promoter normally induced by BMP signaling. Conversely, Bat3 inactivation enhances the induction of direct BMP target genes, such as Id1, Smad6, and Smad7. Consequently, Bat3 deficiency accelerates the differentiation of primary osteoblasts into bone, with a concomitant increase in the bone differentiation markers Runx2, Osterix, and alkaline phosphatase. Using biochemical and cell biological analyses, we show that Bat3 inactivation sustains the C-terminal phosphorylation and nuclear localization of Smad1, 5, and 8 (Smad1/5/8), thereby enhancing biological responses to BMP treatment. At the mechanistic level, we show that Bat3 interacts with the nuclear phosphatase small C-terminal domain phosphatase (SCP) 2, which terminates BMP signaling by dephosphorylating Smad1/5/8. Notably, Bat3 enhances SCP2-Smad1 interaction only when the BMP signaling pathway is activated. Our results demonstrate that Bat3 is an important regulator of BMP signaling that functions by modulating SCP2-Smad interaction. © 2011 Macmillan Publishers Limited. All rights reserved.
  • Wenbin Ma; Blanca Ortiz-Quintero; Roberto Rangel; Morgan R. McKeller; Sara Herrera-Rodriguez; Eliseo F. Castillo; Kimberly S. Schluns; Mary Hall; Huiyuan Zhang; Woong-Kyung Suh; Hitoshi Okada; Tak W. Mak; Yang Zhou; Michael R. Blackburn; Hector Martinez-Valdez
    CELL RESEARCH INST BIOCHEMISTRY & CELL BIOLOGY 21 (11) 1564 - 1577 1001-0602 2011/11 [Refereed]
     
    Gene expression can be regulated by chromatin modifiers, transcription factors and proteins that modulate DNA architecture. Among the latter, AT-hook transcription factors have emerged as multifaceted regulators that can activate or repress broad A/ T-rich gene networks. Thus, alterations of AT-hook genes could affect the transcription of multiple genes causing global cell dysfunction. Here we report that targeted deletions of mouse AKNA, a hypothetical AT-hook-like transcription factor, sensitize mice to pathogen-induced inflammation and cause sudden neonatal death. Compared with wild-type littermates, AKNA KO mice appeared weak, failed to thrive and most died by postnatal day 10. Systemic inflammation, predominantly in the lungs, was accompanied by enhanced leukocyte infiltration and alveolar destruction. Cytologic, immunohistochemical and molecular analyses revealed CD11b(+)Gr1(+) neutrophils as major tissue infiltrators, neutrophilic granule protein, cathelin-related antimicrobial peptide and S100A8/9 as neutrophil-specific chemoattracting factors, interleukin-1 beta and interferon-gamma as proinflammatory mediators, and matrix metalloprotease 9 as a plausible proteolytic trigger of alveolar damage. AKNA KO bone marrow transplants in wildtype recipients reproduced the severe pathogen-induced reactions and confirmed the involvement of neutrophils in acute inflammation. Moreover, promoter/reporter experiments showed that AKNA could act as a gene repressor. Our results support the concept of coordinated pathway-specific gene regulation functions modulating the intensity of inflammatory responses, reveal neutrophils as prominent mediators of acute inflammation and suggest mechanisms underlying the triggering of acute and potentially fatal immune reactions.
  • X. Wu; X. Wang; J. Zhu; Q. Zhang; K. Xu; T. Yang; H. Okada; M. Woo
    DIABETOLOGIA SPRINGER 54 S25 - S25 0012-186X 2011/09
  • Masahito Kawazu; Kayoko Saso; Kit I. Tong; Tracy McQuire; Kouichiro Goto; Dong-Ok Son; Andrew Wakeham; Makoto Miyagishi; Tak W. Mak; Hitoshi Okada
    PLOS ONE PUBLIC LIBRARY SCIENCE 6 (3) e17830  1932-6203 2011/03 [Refereed]
     
    Estrogen is a key regulator of normal function of female reproductive system and plays a pivotal role in the development and progression of breast cancer. Here, we demonstrate that JMJD2B (also known as KDM4B) constitutes a key component of the estrogen signaling pathway. JMJD2B is expressed in a high proportion of human breast tumors, and that expression levels significantly correlate with estrogen receptor (ER) positivity. In addition, 17-beta-estradiol (E2) induces JMJD2B expression in an ER alpha dependent manner. JMJD2B interacts with ER alpha and components of the SWI/SNF-B chromatin remodeling complex. JMJD2B is recruited to ER alpha target sites, demethylates H3K9me3 and facilitates transcription of ER responsive genes including MYB, MYC and CCND1. As a consequence, knockdown of JMJD2B severely impairs estrogen-induced cell proliferation and the tumor formation capacity of breast cancer cells. Furthermore, Jmjd2b-deletion in mammary epithelial cells exhibits delayed mammary gland development in female mice. Taken together, these findings suggest an essential role for JMJD2B in the estrogen signaling, and identify JMJD2B as a potential therapeutic target in breast cancer.
  • Wei Qiu; Xinwei Wang; Brian Leibowitz; Hongtao Liu; Nick Barker; Hitoshi Okada; Naohide Oue; Wataru Yasui; Hans Clevers; Robert E. Schoen; Jian Yu; Lin Zhang
    PROCEEDINGS OF THE NATIONAL ACADEMY OF SCIENCES OF THE UNITED STATES OF AMERICA NATL ACAD SCIENCES 107 (46) 20027 - 20032 0027-8424 2010/11 [Refereed]
     
    Nonsteroidal anti-inflammatory drugs (NSAIDs) such as sulindac effectively prevent colon cancer in humans and rodent models. However, their cellular targets and underlying mechanisms have remained elusive. We found that dietary sulindac induced apoptosis to remove the intestinal stem cells with nuclear or phosphorylated beta-catenin in APC(Min/+) mice. NSAIDs also induced apoptosis in human colonic polyps and effectively removed cells with aberrant Wnt signaling. Furthermore, deficiency in SMAC, a mitochondrial apoptogenic protein, attenuated the tumor-suppressive effect of sulindac in APC(Min/+) mice by blocking apoptosis and removal of stem cells with nuclear or phosphorylated beta-catenin. These results suggest that effective chemoprevention of colon cancer by NSAIDs lies in the elimination of stem cells that are inappropriately activated by oncogenic events through induction of apoptosis.
  • X. Wu; Q. Zhang; C. Liu; K. Xu; X. Mao; T. Yang; H. Okada; M. Woo
    DIABETOLOGIA SPRINGER 53 S74 - S74 0012-186X 2010/09
  • X. Wu; L. Wang; S. Schroer; D. Choi; P. Chen; H. Okada; M. Woo
    Diabetologia SPRINGER 52 (10) 2130 - 2141 0012-186X 2009/10 [Refereed]
     
    Aims/hypothesis: Pancreatic beta cells undergo dynamic remodelling during the perinatal period, with enhanced neogenesis, proliferation and apoptosis observed. The molecular mechanisms responsible for these processes have yet to be elucidated. Survivin is an inhibitor of apoptosis, first described as being exclusively expressed in tumour and embryonic tissues with regulatory functions in mitosis and apoptosis. The aim of the present study was to define the essential physiological role of survivin in the pancreas. Methods: The expression profile of survivin was assessed in the mouse pancreas, and we generated a Pdx1 promoter-driven Survivin (also known as Birc5) knockout mouse using the Cre-loxP recombination system to determine the essential physiological function of survivin in the pancreas. Results: Survivin is transiently expressed in mouse pancreatic islets during the embryonic and neonatal periods. Targeted deletion of Survivin in the pancreas resulted in a significant decline in beta cell mass throughout the perinatal period, leading to glucose intolerance in the adult. Survivin-deficient islets showed decreased cell proliferation as a result of a delay in cell cycle progression with perturbations in cell cycle proteins. Survivin did not, however, play an essential role in beta cell apoptosis either during the physiological remodelling period or in response to streptozotocin. Islet development, islet architecture, microvasculature and apoptosis were not affected by the absence of survivin in the pancreas. Conclusions/interpretation: Survivin expression in the pancreatic islets during the perinatal remodelling period is essential for the establishment of beta cell mass through cell cycle regulation. © 2009 Springer-Verlag.
  • Toru Sasaki; Edyta Marcon; Tracy McQuire; Yoichi Arai; Peter B. Moens; Hitoshi Okada
    JOURNAL OF CELL BIOLOGY ROCKEFELLER UNIV PRESS 182 (3) 449 - 458 0021-9525 2008/08 [Refereed]
     
    Meiosis is critical for sexual reproduction. During meiosis, the dynamics and integrity of homologous chromosomes are tightly regulated. The genetic and molecular mechanisms governing these processes in vivo, however, remain largely unknown. In this study, we demonstrate that Bat3/Scythe is essential for survival and maintenance of male germ cells (GCs). Targeted inactivation of Bat3/Scythe in mice results in widespread apoptosis of meiotic male GCs and complete male infertility. Pachytene spermatocytes exhibit abnormal assembly and disassembly of synaptonemal complexes as demonstrated by abnormal SYCP3 staining and sustained gamma-H2AX and Rad51/replication protein A foci. Further investigation revealed that a testis-specific protein, Hsp70-2/HspA2, is absent in Bat3-deficient male GCs at any stage of spermatogenesis; however, Hsp70-2 transcripts are expressed at normal levels. We found that Bat3 deficiency induces polyubiquitylation and subsequent degradation of Hsp70-2. Inhibition of proteasomal degradation restores Hsp70-2 protein levels. Our findings identify Bat3 as a critical regulator of Hsp70-2 in spermatogenesis, thereby providing a possible molecular target in idiopathic male infertility.
  • The Potentially Role of Survivin in Pancreatic Islet Development and Function
    Wu X; Wang L; Chen P; Okada H; Woo M
    Canadian Journal of Diabetes 32 (4) 339  2008 [Refereed]
  • Andrew T. Ho; Qin H. Li; Hitoshi Okada; Tak W. Mak; Eldad Zacksenhaus
    MOLECULAR AND CELLULAR BIOLOGY AMER SOC MICROBIOLOGY 27 (16) 5673 - 5685 0270-7306 2007/08 [Refereed]
     
    The current model for the intrinsic apoptotic pathway holds that mitochondrial activation of caspases in response to cytotoxic drugs requires both Apaf-I-induced dimerization of procaspase 9 and Smac/Diablo-mediated sequestration of inhibitors of apoptosis proteins (IAPs). Here, we showed that either pathway can independently promote caspase 9 activation in response to apoptotic stimuli. In drug-treated Apaf-1(-/-) primary myoblasts, but not fibroblasts, Smac/Diablo accumulates in the cytosol and sequesters X-linked IAP (XIAP), which is expressed at lower levels in myoblasts than in fibroblasts. Consequently, caspase 9 activation proceeds in Apaf-1(-/-) myoblasts; concomitant ablation of Apaf-1 and Smac is required to prevent caspase 9 activation and the onset of apoptosis. Conversely, in stimulated Apaf-1(-/-) fibroblasts, the ratio of XIAP to Smac/Diablo is high compared to that for myoblasts and procaspase 9 is not activated. Suppressing XIAP with exogenous Smac/Diablo or a pharmacological inhibitor can still induce caspase 9 in drug-treated Apaf-1-null fibroblasts. Thus, caspase 9 activation in response to intrinsic apoptotic stimuli can be uncoupled from Apaf-1 in vivo by XIAP antagonists.
  • Toru Sasaki; Eugene C. Gan; Andrew Wakeham; Sally Kornbluth; Tak W. Mak; Hitoshi Okada
    GENES & DEVELOPMENT COLD SPRING HARBOR LAB PRESS, PUBLICATIONS DEPT 21 (7) 848 - 861 0890-9369 2007/04 [Refereed]
     
    In response to DNA damage, p53 undergoes post-translational modifications (including acetylation) that are critical for its transcriptional activity. However, the mechanism by which p53 acetylation is regulated is still unclear. Here, we describe an essential role for HLA-B-associated transcript 3 (Bat3)/Scythe in controlling the acetylation of p53 required for DNA damage responses. Depletion of Bat3 from human and mouse cells markedly impairs p53-mediated transactivation of its target genes Puma and p21. Although DNA damage-induced phosphorylation, stabilization, and nuclear accumulation of p53 are not significantly affected by Bat3 depletion, p53 acetylation is almost completely abolished. Bat3 forms a complex with p300, and an increased amount of Bat3 enhances the recruitment of p53 to p300 and facilitates subsequent p53 acetylation. In contrast, Bat3-depleted cells show reduced p53-p300 complex formation and decreased p53 acetylation. Furthermore, consistent with our in vitro findings, thymocytes from Bat3-deficient mice exhibit reduced induction of puma and p21, and are resistant to DNA damage-induced apoptosis in vivo. Our data indicate that Bat3 is a novel and essential regulator of p53-mediated responses to genotoxic stress, and that Bat3 controls DNA damage-induced acetylation of p53.
  • G. I. Perez; B. M. Acton; A. Jurisicova; G. A. Perkins; A. White; J. Brown; A. M. Trbovich; M. R. Kim; R. Fissore; J. Xu; A. Ahmady; S. G. D'Estaing; H. Li; W. Kagawa; H. Kurumizaka; S. Yokoyama; H. Okada; T. W. Mak; M. H. Ellisman; R. F. Casper; J. L. Tilly
    Cell Death and Differentiation NATURE PUBLISHING GROUP 14 (3) 524 - 533 1350-9047 2007/03 [Refereed]
     
    Although the identification of specific genes that regulate apoptosis has been a topic of intense study, little is known of the role that background genetic variance plays in modulating cell death. Using germ cells from inbred mouse strains, we found that apoptosis in mature (metaphase II) oocytes is affected by genetic background through at least two different mechanisms. The first, manifested in AKR/J mice, results in genomic instability. This is reflected by numerous DNA double-strand breaks in freshly isolated oocytes, causing a high apoptosis susceptibility and impaired embryonic development following fertilization. Microinjection of Rad51 reduces DNA damage, suppresses apoptosis and improves embryonic development. The second, manifested in FVB mice, results in dramatic dimorphisms in mitochondrial ultrastructure. This is correlated with cytochrome c release and a high apoptosis susceptibility, the latter of which is suppressed by pyruvate treatment, Smac/DIABLO deficiency, or microinjection of 'normal' mitochondria. Therefore, background genetic variance can profoundly affect apoptosis in female germ cells by disrupting both genomic DNA and mitochondrial integrity.
  • Tomomasa Yokomizo; Satoru Takahashi; Naomi Mochizuki; Takashi Kuroha; Masatsugu Ema; Asami Wakamatsu; Ritsuko Shimizu; Osamu Ohneda; Motomi Osato; Hitoshi Okada; Toshihisa Komori; Minetaro Ogawa; Shin-Ichi Nishikawa; Yoshiaki Ito; Masayuki Yamamoto
    EMBO JOURNAL WILEY 26 (1) 184 - 196 0261-4189 2007/01 [Refereed]
     
    Hemangioblasts are thought to be one of the sources of hematopoietic progenitors, yet little is known about their localization and fate in the mouse embryo. We show here that a subset of cells co-expressing the hematopoietic marker GATA-1 and the endothelial marker VE-cadherin localize on the yolk sac blood islands at embryonic day 7.5. Clonal analysis demonstrated that GATA-1(+) cells isolated from E7.0-7.5 embryos include a common precursor for hematopoietic and endothelial cells. Moreover, this precursor possesses primitive and definitive hematopoietic bipotential. By using a transgenic complementation rescue approach, GATA-1(+) cell-derived progenitors were selectively restored in Runx1-deficient mice. In the rescued mice, definitive erythropoiesis was recovered but the rescued progenitors did not display multilineage hematopoiesis or intra-aortic hematopoietic clusters. These results provide evidence of the presence of GATA-1(+) hemangioblastic cells in the extra-embryonic region and also their functional contribution to hematopoiesis in the embryo.
  • Woong-Kyung Suh; Seng Wang; Gordon S. Duncan; Yoshiyuki Miyazaki; Elizabeth Cates; Tina Walker; Beata U. Gajewska; Elissa Deenick; Wojciech Dawicki; Hitoshi Okada; Andrew Wakeham; Annick Itie; Tania H. Watts; Pamela S. Ohashi; Manel Jordana; Hiroki Yoshida; Tak W. Mak
    MOLECULAR AND CELLULAR BIOLOGY AMER SOC MICROBIOLOGY 26 (17) 6403 - 6411 0270-7306 2006/09 [Refereed]
     
    Members of the B7 family of cosignaling molecules regulate T-cell proliferation and effector functions by engaging cognate receptors on T cells. In vitro and in vivo blockade experiments indicated that B7-H4 (also known as B7S1 or B7x) inhibits proliferation, cytokine production, and cytotoxicity of T cells. B7-H4 binds to an unknown receptor(s) that is expressed on activated T cells. However, whether B7-H4 plays nonredundant immune regulatory roles in vivo has not been tested. We generated B7-H4-deficient mice to investigate the roles of B7-H4 during various immune reactions. Consistent with its inhibitory function in vitro, B7-H4-deficient mice mounted mildly augmented T-helper 1 (Th1) responses and displayed slightly lowered parasite burdens upon Leishmania major infection compared to the wild-type mice. However, the lack of B7-H4 did not affect hypersensitive inflammatory responses in the airway or skin that are induced by either Thl or Th2 cells. Likewise, B7-H4-deficient mice developed normal cytotoxic T-lymphocyte reactions against viral infection. Thus, B7-H4 plays a negative regulatory role in vivo but the impact of B7-H4 deficiency is minimal. These results suggest that B7-H4 is one of multiple negative cosignaling molecules that collectively provide a fine-tuning mechanism for T-cell-mediated immune responses.
  • Andrew T. Ho; Hui-Qin Li; Hitoshi Okada; Tak W. Mak; Eldad Zacksenhaus
    CANCER RESEARCH AMER ASSOC CANCER RESEARCH 66 (8) 562 - 562 0008-5472 2006/04 [Refereed]
  • Laura Alonso; Hitoshi Okada; Hilda Amalia Pasolli; Andrew Wakeham; Annick Itie You-Ten; Tak W. Mak; Elaine Fuchs
    Journal of Cell Biology ROCKEFELLER UNIV PRESS 170 (4) 559 - 570 0021-9525 2005/08 [Refereed]
     
    Tyrosine kinase growth factor receptor signaling influences proliferation, survival, and apoptosis. Hair follicles undergo cycles of proliferation and apoptotic regression, offering an excellent paradigm to study how this transition is governed. Several factors are known to affect the hair cycle, but it remains a mystery whether Akt kinases that are downstream of growth factor signaling impact this equilibrium. We now show that an Akt relative, Sgk (serum and glucocorticoid responsive kinase) 3, plays a critical role in this process. Hair follicles of mice lacking Sgk3 fail to mature normally. Proliferation is reduced, apoptosis is increased, and follicles prematurely regress. Maintenance of the pool of transiently amplifying matrix cells is impaired. Intriguingly, loss of Sgk3 resembles the gain of function of epidermal growth factor signaling. Using cultured primary keratinocytes, we find that Sgk3 functions by negatively regulating phosphatidylinositol 3 kinase signaling. Our results reveal a novel and important function for Sgk3 in controlling life and death in the hair follicle. © The Rockefeller University Press.
  • Katsuya Tsuchihara; Valentina Lapin; Christopher Bakal; Hitoshi Okada; Lauren Brown; Masami Hirota-Tsuchihara; Kathrin Zaugg; Alexandra Ho; Annick Itie-YouTen; Marees Harris-Brandts; Robert Rottapel; Christopher D. Richardson; Samuel Benchimol; Tak Wah Mak
    Cancer Research AMER ASSOC CANCER RESEARCH 65 (15) 6685 - 6691 0008-5472 2005/08 [Refereed]
     
    We used DNA microarray screening to identify Ckap2 (cytoskeleton associated protein 2) as a novel p53 target gene in a mouse erythroleukemia cell line. DNA damage induces human and mouse CKAP2 expression in a p53-dependent manner and p53 activates the Ckap2 promoter. Overexpressed Ckap2 colocalizes with and stabilizes microtubules. In p53-null cells, overexpression of Ckap2 induces tetraploidy with aberrant centrosome numbers, suggesting disturbed mitosis and cytokinesis. In p53-competent cells, Ckap2 does not induce tetraploidy but activates p53-mediated cell cycle arrest and apoptosis. Our data suggest the existence of a functional positive feedback loop in which Ckap2 activates the G1 tetraploidy checkpoint and prevents aneuploidy. ©2005 American Association for Cancer Research.
  • Zhenyue Hao; Gordon S. Duncan; Chia Che Chang; Andrew Elia; Min Fang; Andrew Wakeham; Hitoshi Okada; Thomas Calzascia; Yingju Jang; Annick You-Ten; Wen Chen Yeh; Pamela Ohashi; Xiaodong Wang; Tak W. Mak
    Cell CELL PRESS 121 (4) 579 - 591 0092-8674 2005/05 [Refereed]
     
    As components of the apoptosome, a caspase-activating complex, cytochrome c (Cyt c) and Apaf-1 are thought to play critical roles during apoptosis. Due to the obligate function of Cyt c in electron transport, its requirement for apoptosis in animals has been difficult to establish. We generated "knockin" mice expressing a mutant Cyt c (KA allele), which retains normal electron transfer function but fails to activate Apaf-1. Most KA/KA mice displayed embryonic or perinatal lethality caused by defects in the central nervous system, and surviving mice exhibited impaired lymphocyte homeostasis. Although fibroblasts from the KA/KA mice were resistant to apoptosis, their thymocytes were markedly more sensitive to death stimuli than Apaf-1 -/- thymocytes. Upon treatment with γ irradiation, procaspases were efficiently activated in apoptotic KA/KA thymocytes, but Apaf-1 oligomerization was not observed. These studies indicate the existence of a Cyt c- and apoptosome-independent but Apaf-1-dependent mechanism(s) for caspase activation. Copyright ©2005 by Elsevier Inc.
  • L. Miguel Martins; Alastair Morrison; Kristina Klupsch; Valentina Fedele; Nicoleta Moisoi; Peter Teismann; Alejandro Abuin; Evelyn Grau; Martin Geppert; George P. Livi; Caretha L. Creasy; Alison Martin; Iain Hargreaves; Simon J. Heales; Hitoshi Okada; Sebastian Brandner; Jörg B. Schulz; Tak Mak; Julian Downward
    Molecular and Cellular Biology AMER SOC MICROBIOLOGY 24 (22) 9848 - 9862 0270-7306 2004/11 [Refereed]
     
    The serine protease HtrA2/Omi is released from the mitochondrial intermembrane space following apoptotic stimuli. Once in the cytosol, HtrA2/Omi has been implicated in promoting cell death by binding to inhibitor of apoptosis proteins (IAPs) via its amino-terminal Reaper-related motif, thus inducing caspase activity, and also in mediating caspase-independent death through its own protease activity. We report here the phenotype of mice entirely lacking expression of HtrA2/Omi due to targeted deletion of its gene, Prss25. These animals, or cells derived from them, show no evidence of reduced rates of cell death but on the contrary suffer loss of a population of neurons in the striatum, resulting in a neurodegenerative disorder with a parkinsonian phenotype that leads to death of the mice around 30 days after birth. The phenotype of these mice suggests that it is the protease function of this protein and not its IAP binding motif that is critical. This conclusion is reinforced by the finding that simultaneous deletion of the other major IAP binding protein, Smac/DIABLO, does not obviously alter the phenotype of HtrA2/Omi knockout mice or cells derived from them. Mammalian HtrA2/Omi is therefore likely to function in vivo in a manner similar to that of its bacterial homologues DegS and DegP, which are involved in protection against cell stress, and not like the proapoptotic Reaper family proteins in Drosophila melanogaster.
  • Gaussin, V; H Okada; RS Foo; G Diaz; C Hong; J Thaisz; GP Yang; CF Peng; TW Mak; SF Vatner; RN Kitsis
    CIRCULATION LIPPINCOTT WILLIAMS & WILKINS 110 (17) 42 - 42 0009-7322 2004/10 [Refereed]
  • Han You; Ying Ju Jang; Annick Itie You-Ten; Hitoshi Okada; Jennifer Liepa; Andrew Wakeham; Kathrin Zaugg; Tak W. Mak
    Proceedings of the National Academy of Sciences of the United States of America NATL ACAD SCIENCES 101 (39) 14057 - 14062 0027-8424 2004/09 [Refereed]
     
    FKHRL1 (FOXO3a) and p53 are two potent stress-response regulators. Here we show that these two transcription factors exhibit "crosstalk" in vivo. In response to DNA damage, p53 activation led to FKHRL1 phosphorylation and subcellular localization change, which resulted in inhibition of FKHRL1 transcription activity. AKT was dispensable for p53-dependent suppression of FKHRL1. By contrast, serum- and glucocorticoid-inducible kinase 1 (SGK1) was significantly induced in a p53-dependent manner after DNA damage, and this induction was through extracellular signal-regulated kinase 1/2-mediated posttranslational regulation. Furthermore, inhibition of SGK1 expression by a small interfering RNA knockdown experiment significantly decreased FKHRL1 phosphorylation in response to DNA damage. Taken together, our observations reveal previously unrecognized crosstalk between p53 and FKHRL1. Moreover, our findings suggest a new pathway for understanding aging and the age dependency of human diseases governed by these two transcription factors.
  • Hitoshi Okada; Tak W. Mak
    Nature Reviews Cancer NATURE PUBLISHING GROUP 4 (8) 592 - 603 1474-175X 2004/08 [Refereed]
     
    Defects in cell-death pathways are hallmarks of cancer. Although resistance to apoptosis is closely linked to tumorigenesis, tumour cells can still be induced to die by non-apoptotic mechanisms, such as necrosis, senescence, autophagy and mitotic catastrophe. The molecular pathways that underlie these non-apoptotic responses remain unclear. Several apoptotic and non-apoptotic pathways of cell death have been defined in normal physiology and during tumorigenesis, and these could potentially be manipulated to develop new cancer therapies. The mitotic-checkpoint molecule survivin - the inactivation of which induces the death of p53-deficient cells by mitotic catastrophe - is of particular interest.
  • Woong Kyung Suh; Anna Tafuri; Nancy N. Berg-Brown; Arda Shahinian; Suzanne Plyte; Gordon S. Duncan; Hitoshi Okada; Andrew Wakeham; Bernhard Odermatt; Pamela S. Ohashi; Tak W. Mak
    Journal of Immunology AMER ASSOC IMMUNOLOGISTS 172 (10) 5917 - 5923 0022-1767 2004/05 [Refereed]
     
    CD28 plays crucial costimulatory roles in T cell proliferation, cytokine production, and germinal center response. Mice that are deficient in the inducible costimulator (ICOS) also have defects in cytokine production and germinal center response. Because the full induction of ICOS in activated T cells depends on CD28 signal, the T cell costimulatory capacity of ICOS in the absence of CD28 has remained unclear. We have clarified this issue by comparing humoral immune responses in wild-type, CD28 knockout (CD28 KO), and CD28-ICOS double-knockout (DKO) mice. DKO mice had profound defects in Ab responses against environmental Ags, T-dependent protein Ags, and vesicular stomatitis virus that extended far beyond those observed in CD28 KO mice. However, DKO mice mounted normal Ab responses against a T-independent Ag, indicating that B cell function itself was normal. Restimulated CD4+ DKO T cells that had been primed in vivo showed decreased proliferation and reduced IL-4 and IL-10 production compared with restimulated CD4+ T cells from CD28 KO mice. Thus, in the absence of CD28, ICOS assumes the major T cell costimulatory role for humoral immune responses. Importantly, CD28-mediated ICOS up-regulation is not essential for ICOS function in vivo.
  • Hitoshi Okada; Chris Bakal; Arda Shahinian; Andrew Elia; Andrew Wakeham; Woong Kyung Suh; Gordon S. Duncan; Maria Ciofani; Robert Rottapel; Juan Carlos Zúñiga-Pflücker; Tak W. Mak
    Journal of Experimental Medicine ROCKEFELLER UNIV PRESS 199 (3) 399 - 410 0022-1007 2004/02 [Refereed]
     
    Because survivin-null embryos die at an early embryonic stage, the role of survivin in thymocyte development is unknown. We have investigated the role by deleting the survivin gene only in the T lineage and show here that loss of survivin blocks the transition from CD4- CD8- double negative (DN) thymocytes to CD4+ CD8+ double positive cells. Although the pre-T cell receptor signaling pathway is intact in survivin-deficient thymocytes, the cells cannot respond to its signals. In response to proliferative stimuli, cycling survivin-deficient DN cells exhibit cell cycle arrest, a spindle formation defect, and increased cell death. Strikingly, loss of survivin activates the tumor suppressor p53. However, the developmental defects caused by survivin deficiency cannot be rescued by p53 inactivation or introduction of Bcl-2. These lines of evidence indicate that developing thymocytes depend on the cytoprotective function of survivin and that this function is tightly coupled to cell proliferation but independent of p53 and Bcl-2. Thus, survivin plays a critical role in early thymocyte development.
  • Woong Kyung Suh; Beata U. Gajewska; Hitoshi Okada; Matthew A. Gronski; Edward M. Bertram; Wojciech Dawicki; Gordon S. Duncan; Jacob Bukczynski; Suzanne Plyte; Andrew Elia; Andrew Wakeham; Annick Itie; Stephen Chung; Joan Da Costa; Sudha Arya; Tom Horan; Pauline Campbell; Kevin Gaida; Pamela S. Ohashi; Tania H. Watts; Steven K. Yoshinaga; Mark R. Bray; Manel Jordana; Tak W. Mak
    Nature Immunology NATURE PUBLISHING GROUP 4 (9) 899 - 906 1529-2908 2003/09 [Refereed]
     
    We investigated the in vivo function of the B7 family member B7-H3 (also known as B7RP-2) by gene targeting. B7-H3 inhibited T cell proliferation mediated by antibody to T cell receptor or allogeneic antigen-presenting cells. B7-H3-deficient mice developed more severe airway inflammation than did wild-type mice in conditions in which T helper cells differentiated toward type 1 (TH1) rather than type 2 (TH2). B7-H3 expression was consistently enhanced by interferon-γ but suppressed by interleukin 4 in dendritic cells. B7-H3-deficient mice developed experimental autoimmune encephalomyelitis several days earlier than their wild-type littermates, and accumulated higher concentrations of autoantibodies to DNA. Thus, B7-H3 is a negative regulator that preferentially affects TH1 responses.
  • Toshiaki Hamamoto; Hideyuki Beppu; Hitoshi Okada; Masahiro Kawabata; Tadaichi Kitamura; Kohei Miyazono; Mitsuyasu Kato
    Cancer research AMER ASSOC CANCER RESEARCH 62 (20) 5955 - 61 0008-5472 2002/10 [Refereed]
     
    Smad2 is a receptor-regulated Smad that is activated specifically by transforming growth factor beta and activin signaling. We disrupted the mouse Smad2 gene by gene targeting. Homozygous Smad2 mutant mice died around E8.5 with impaired visceral endoderm function and deficiency of mesoderm formation. Heterozygotes were fertile and had no apparent abnormality up to at least 1 1/2 year of age. To examine the role of Smad2 inactivation in the process of carcinogenesis, we prepared compound heterozygous mice, which carry both Apc and Smad2 mutations on the same chromosome in the cis-configuration. Compound inactivation of Smad2 in heterozygous Apc mutant mice did not change the total number of intestinal tumors but increased sudden death from intestinal obstruction caused by extremely large tumors. Furthermore, histological examination revealed that Apc/Smad2 cis-compound heterozygotes developed multiple invasive cancers that had never been observed in Apc single heterozygotes. These results indicate that loss of Smad2 does not initiate tumorigenesis by itself but accelerates malignant progression of tumors to invasive cancer in the late stages of carcinogenesis.
  • Atsushi Hirao; Alison Cheung; Gordon Duncan; Pierre Marie Girard; Andrew J. Elia; Andrew Wakeham; Hitoshi Okada; Talin Sarkissian; Jorge A. Wong; Takashi Sakai; Elisa De Stanchina; Robert G. Bristow; Toshio Suda; Scott W. Lowe; Penny A. Jeggo; Stephen J. Elledge; Tak W. Mak
    Molecular and Cellular Biology AMER SOC MICROBIOLOGY 22 (18) 6521 - 6532 0270-7306 2002/09 [Refereed]
     
    In response to ionizing radiation (IR), the tumor suppressor p53 is stabilized and promotes either cell cycle arrest or apoptosis. Chk2 activated by IR contributes to this stabilization, possibly by direct phosphorylation. Like p53, Chk2 is mutated in patients with Li-Fraumeni syndrome. Since the ataxia telangiectasia mutated (ATM) gene is required for IR-induced activation of Chk2, it has been assumed that ATM and Chk2 act in a linear pathway leading to p53 activation. To clarify the role of Chk2 in tumorigenesis, we generated genetargeted Chk2-deficient mice. Unlike ATM-/- and p53-/- mice, Chk2-/- mice do not spontaneously develop tumors, although Chk2 does suppress 7,12-dimethylbenzanthracene-induced skin tumors. Tissues from Chk2-/- mice, including those from the thymus, central nervous system, fibroblasts, epidermis, and hair follicles, show significant defects in IR-induced apoptosis or impaired G1/S arrest. Quantitative comparison of the G1/S checkpoint, apoptosis, and expression of p53 proteins in Chk2-/- versus ATM-/- thymocytes suggested that Chk2 can regulate p53-dependent apoptosis in an ATM-independent manner. IR-induced apoptosis was restored in Chk2-/- thymocytes by reintroduction of the wild-type Chk2 gene but not by a Chk2 gene in which the sites phosphorylated by ATM and ataxia telangiectasia and rad3+ related (ATR) were mutated to alanine. ATR may thus selectively contribute to p53-mediated apoptosis. These data indicate that distinct pathways regulate the activation of p53 leading to cell cycle arrest or apoptosis.
  • Hitoshi Okada; Woong-Kyung Suh; Jianping Jin; Minna Woo; Chunying Du; Andrew Elia; Gordon S Duncan; Andrew Wakeham; Annick Itie; Scott W Lowe; Xiaodong Wang; Tak W Mak
    Molecular and cellular biology AMER SOC MICROBIOLOGY 22 (10) 3509 - 17 0270-7306 2002/05 [Refereed]
     
    The mitochondrial proapoptotic protein Smac/DIABLO has recently been shown to potentiate apoptosis by counteracting the antiapoptotic function of the inhibitor of apoptosis proteins (IAPs). In response to apoptotic stimuli, Smac is released into the cytosol and promotes caspase activation by binding to IAPs, thereby blocking their function. These observations have suggested that Smac is a new regulator of apoptosis. To better understand the physiological function of Smac in normal cells, we generated Smac-deficient (Smac(-/-)) mice by using homologous recombination in embryonic stem (ES) cells. Smac(-/-) mice were viable, grew, and matured normally and did not show any histological abnormalities. Although the cleavage in vitro of procaspase-3 was inhibited in lysates of Smac(-/-) cells, all types of cultured Smac(-/-) cells tested responded normally to all apoptotic stimuli applied. There were also no detectable differences in Fas-mediated apoptosis in the liver in vivo. Our data strongly suggest the existence of a redundant molecule or molecules capable of compensating for a loss of Smac function.
  • Sumitaka Hasegawa; Tomoyuki Sato; Hiroshi Akazawa; Hitoshi Okada; Akiteru Maeno; Masaki Ito; Yoshinobu Sugitani; Hiroyuki Shibata; Jun Ichi Miyazaki; Motoya Katsuki; Yasutaka Yamauchi; Ken Ichi Yamamura; Shigeru Katamine; Tetsuo Noda
    Proceedings of the National Academy of Sciences of the United States of America NATL ACAD SCIENCES 99 (1) 297 - 302 0027-8424 2002/01 [Refereed]
     
    Apc is a gene associated with familial adenomatous polyposis coli (FAP) and its inactivation is a critical step in colorectal tumor formation. The protein product, adenomatous polyposis coli (APC), acts to down-regulate intracellular levels of β-catenin, a key signal transducer in the Wnt signaling. Conditional targeting of Apc in the neural crest of mice caused massive apoptosis of cephalic and cardiac neural crest cells at about 11.5 days post coitum, resulting in craniofacial and cardiac anomalies at birth. Notably, the apoptotic cells localized in the regions where β-catenin had accumulated, in contrast to its role in colorectal epithelial cells, inactivation of APC leads to dysregulation of β-catenin/Wnt signaling with resultant apoptosis in certain tissues including neural crest cells.
  • Damu Tang; Hitoshi Okada; Jurgen Ruland; Lieqi Liu; Vuk Stambolic; Tak W. Mak; Alistair J. Ingram
    Journal of Biological Chemistry AMER SOC BIOCHEMISTRY MOLECULAR BIOLOGY INC 276 (32) 30461 - 30466 0021-9258 2001/08 [Refereed]
     
    Akt is a serine-threonine kinase known to exert anti-apoptotic effects through several downstream targets. Akt is cleaved during mitochondrial-mediated apoptosis in a caspase-dependent manner. The reason for this is not clear, however, because Akt has not been demonstrated to be activated in response to mitochondrial apoptotic stimuli. Accordingly, we explored whether the well described mitochondrial apoptotic stimuli staurosporine (STS) and etoposide activate Akt and whether such activation impacts apoptosis. Both STS and etoposide activated Akt in NIH 3T3 cells, maximally at 8 and 2 h, respectively, preceding the onset of apoptosis and poly(ADP-ribose) polymerase cleavage. The overexpression of Akt delayed STS-induced apoptosis with an even more pronounced delay observed with overexpression of constitutively active Akt. Akt activation by proapoptotic stimuli lay upstream of mitochondria, because neither caspase inhibitors nor overexpression of Bcl-2 or Bcl-xL could prevent it. Activation depended on phosphatidylinositol 3-kinase activity, however. Conversely, inhibition of phosphatidylinositol 3-kinase with wort-mannin sensitized cells to apoptosis initiated by STS. These data demonstrate that mitochondrial apoptotic stimuli also activate Akt and such activation modulates apoptosis in this setting.
  • Tomomasa Yokomizo; Minetaro Ogawa; Motomi Osato; Tomohiko Kanno; Hisahiro Yoshida; Tetsuhiro Fujimoto; Stuart Fraser; Satomi Nishikawa; Hitoshi Okada; Masanobu Satake; Tetsuo Noda; Shin Ichi Nishikawa; Yoshiaki Ito
    Genes to Cells BLACKWELL SCIENCE LTD 6 (1) 13 - 23 1356-9597 2001 [Refereed]
     
    Background: Recent studies revealing that endothelial cells derived from E8.5-E10.5 mouse embryos give rise to haematopoietic cells appear to correspond to previous histological observations that haematopoietic cell clusters are attached to the ventral aspect of dorsal aorta in such a way as if they were budding from the endothelial cell layer. Gene disruption studies have revealed that Runx1/AML1 is required for definitive haematopoiesis but not for primitive haematopoiesis, but the precise stage of gene function is not yet known. Results: We found that mice deficient in Runx1/AML1 (an α subunit of the transcription factor PEBP2/CBF) lack c-Kit+ haematopoietic cell clusters in the dorsal aorta, omphalomesenteric and umbilical arteries, as well as yolk sac vessels. Moreover, endothelial cells sorted from the embryo proper and the yolk sac of AML1-/- embryos are unable to differentiate into haematopoietic cells on OP9 stromal cells, whereas colonies of AML1-/- endothelial cells can be formed in culture. Conclusions: These results strongly suggest that the emergence of haematopoietic cells from endothelial cells represents a major pathway of definitive haematopoiesis and is an event that also occurs in the yolk sac in vivo, as suggested by earlier in vitro experiments.
  • Kuniaki Ishii; Kazuo Nunoki; Toshio Yamagishi; Hitoshi Okada; Norio Taira
    Journal of Pharmacology and Experimental Therapeutics AMER SOC PHARMACOLOGY EXPERIMENTAL THERAPEUTICS 296 (2) 405 - 411 0022-3565 2001 [Refereed]
     
    We studied the effects of acidic pH on the currents through voltage-gated K+ channels, Kv1.2, Kv1.4, and their tandem construct (Kv1.4-Kv1.2). Kv1.4 currents were inhibited considerably under acidic pH in a voltage-independent manner, whereas Kv1.2 currents were less inhibited in an apparently voltage-dependent manner. The apparent voltage-dependent block of Kv1.2 currents was mostly ascribed to the shift of activation voltage, which is probably due to surface charge effects of H+ ions. Mutagenesis analysis identified the histidine residue at 508 (H508) in the S5-H5 linker as a molecular determinant of pH sensitivity of Kv1.4. Currents through the tandem channel showed intermediate characteristics between the two parent channels in both sensitivity and voltage dependence of pH effects. Our results suggest that 1) the H508 plays a critical role in determining pH sensitivity of Kv1.4; and 2) the two parent channels, Kv1.2 and Kv1.4, are involved in determining pH sensitivity and apparent voltage dependence of the tandem channel.
  • K. Hayashi; W. Natsume; T. Watanabe; N. Abe; N. Iwai; H. Okada; Y. Ito; M. Asano; Y. Iwakura; S. Habu; Y. Takahama; M. Satake
    Journal of Immunology AMER ASSOC IMMUNOLOGISTS 165 (12) 6816 - 6824 0022-1767 2000/12 [Refereed]
     
    In the thymic cortex, T lymphocytes are positively selected to survive and committed either to the CD4 single-positive (SP) or the CD8 SP lineage. The SP cells then pass through a step of maturation in the medulla and are delivered to peripheral lymphoid tissues. We examined the role of AML1, the gene encoding a transcription factor, in the above processes by using the transgenic mice expressing a dominant interfering form of AML1 as well as mice targeted heterozygously for AML1. One phenotypic change seen in the AML1-diminished mice was the reduction in the numbers of both CD4 SP and CD8 SP thymocytes, reflecting the partial impairment of the transition from the double-positive to SP stage. In addition, distinct from the above abnormality, perturbed were several aspects of SP cells, including the maturation of SP thymoeytes, the recent thymic emigration, and the proliferative responsiveness of peripheral T cells to TCR stimulation. Interestingly, the AML1 diminution caused inhibitory and enhancing effects on the CD4 SP and CD8 SP cells, respectively. These differential effects are most likely related to the reduction in the peripheral CD4 SP/CD8 SP ratio observed in the AML1-diminished mice. The AML1 transcription factor tiros maintains the homeostasis of each SP subset by functioning at the later stages of T lymphocyte differentiation.
  • Yoh suke Mukouyama; Natsuko Chiba; Takahiko Hara; Hitoshi Okada; Yoshiaki Ito; Ryunosuke Kanamaru; Atsushi Miyajima; Masanobu Satake; Toshio Watanabe
    Developmental Biology ACADEMIC PRESS INC 220 (1) 27 - 36 0012-1606 2000/04 [Refereed]
     
    We examined the role of the AML1 transcription factor in the development of hematopoiesis in the paraaortic splanchnopleural (P-Sp) and the aorta- gonad-mesonephros (AGM) regions of mouse embryos. The activity of colony- forming units of colonies from the P-Sp/AGM region was reduced severalfold by heterozygous disruption of the AML1 gene, indicating that AML1 functioned in a dosage-dependent manner to generate hematopoietic progenitors. In addition, no hematopoietic progenitor activity was detected in the P-Sp/AGM region of embryos with an AML1 null mutation. Similar results were obtained when a dispersed culture was first prepared from the P-Sp/AGM region before assay of the activity of the colony-forming units. In a culture of cells with the AML1(+/+) genotype, both hematopoietic and endothelial-like cell types emerged, but in a culture of cells with the AML1(-/-) genotype, only endothelial-like cells emerged. Interestingly, introduction of AML1 cDNA into the P-Sp/AGM culture with the AML1(-/-) genotype partially restored the production of hematopoietic cells. This restoration was observed for cultures prepared from 9.5-day postcoitum (dpc) embryos but not for cultures prepared from 11.5-dpc embryos. Therefore, the population of endothelial-like cells capable of growing in the AML1(-/-) culture would appear to contain inert but nonetheless competent hematogenic precursor cells up until at least the 9.5- dpc period. All these results support the notion that the AML1 transcription factor functions to develop and maintain hematogenic precursor cells in the embryonic P-Sp/AGM region. (C) 2000 Academic Press.
  • Hitoshi Okada; Toshio Watanabe; Masaru Niki; Hiroshi Takano; Natsuko Chiba; Nobuaki Yanai; Kenzaburo Tani; Hitoshi Hibino; Shigetaka Asano; Michael L. Mucenski; Yoshiaki Ito; Tetsuo Noda; Masanobu Satake
    Oncogene STOCKTON PRESS 17 (18) 2287 - 2293 0950-9232 1998/11 [Refereed]
     
    The AML1 and PEBP2β/CBFβ genes encode the DNA-binding and non-binding subunits, respectively, of the heterodimeric transcription factor, PEBP2/CBF. Targeting each gene results in an almost identical phenotype, namely the complete lack of definitive hematopoiesis in the fetal liver on embryonic day 11.5 (E11.5). We examined and compared the expression levels of various hematopoiesis-related genes in wild type embryos and in embryos mutated for AML1 or PEBP2β/CBFβ. The RNAs were prepared from the yolk sacs of E9.5 embryos, from the aorta-gonad- mesonephros regions of E11.5 embryos and from the livers of E11.5 embryos and RT-PCR was performed to detect various gene transcripts. Transcripts were detected for most of the hematopoiesis-related genes that encode transcription factors, cytokines and cytokine receptors, even in tissues from homozygously targeted embryos. On the other hand, PU.1 transcripts were never detected in any tissue of AML1(-/-) or PEBP2β/CBFβ(-/-) embryos. In addition, transcripts for the Vav, flk-2/flt-3, M-CSF receptor G-CSF receptor and c-Myb genes were not detected in certain tissues of the (-/-) embryos. The results suggest that the expression of a particular set of hematopoiesis-related genes is closely correlated with the PEBP2/CBF function.
  • Masaru Niki; Hitoshi Okada; Hiroshi Takano; Junko Kuno; Kenzaburo Tani; Hitoshi Hibino; Shigetaka Asano; Yoshiaki Ito; Masanobu Satake; Tetsuo Noda
    Proceedings of the National Academy of Sciences of the United States of America NATL ACAD SCIENCES 94 (11) 5697 - 5702 0027-8424 1997/05 [Refereed]
     
    The Pebpb2 gene encodes a non-DNA binding subunit of the heterodimeric transcription factor, polyomavirus enhancer binding protein 2/core binding factor (PEBP2/CBF), and is rearranged in inversion of chromosome 16 associated with human acute myeloid leukemia. To investigate its physiological function, Pebpb2 was mutated by a targeting strategy to generate a null mutant. The homozygous mutation in mice proved lethal in embryos around embryonic day 12.5, apparently due to massive hemorrhaging in the central nervous system. In addition, definitive hematopoiesis in the liver was severely impaired. The observed phenotype was indistinguishable from that reported for homozygous disruption of AML1, which encodes a DNA binding subunit of PEBP2/CBF. Thus, the results indicate that the two subunits function together as a heterodimeric PEBP2/CBF in vivo and that PEBP2/CBF plays an essential role in the development of definitive hematopoiesis.
  • Suehiro Sakaguchi; Shigeru Katamine; Noriyuki Nishida; Ryozo Moriuchi; Kazuto Shigematsu; Tetsuo Sugimoto; Akira Nakatani; Yasufumi Kataoka; Takeshi Houtani; Susumu Shirabe; Hitoshi Okada; Sumitaka Hasegawa; Tsutomu Miyamoto; Tetsuo Noda
    Nature MACMILLAN MAGAZINES LTD 380 (6574) 528 - 531 0028-0836 1996/04 [Refereed]
     
    Prion protein (PrP) is a glycoprotein constitutively expressed on the neuronal cell surface. A protease-resistant isoform of prion protein is implicated in the pathogenesis of a series of transmissible spongiform encephalopathies. We have developed a line of mice homozygous for a disrupted. PrP gene in which the whole PrP-coding sequence is replaced by a drug-resistant gene. In keeping with previous results, we find that homozygous loss of the PrP gene has no deleterious effect on the development of these mice and renders them resistant to prion. The PrP-null mice grew normally after birth, but at about 70 weeks of age all began to show progressive symptoms of ataxia. Impaired motor coordination in these ataxic mice was evident in a rotorod test. Pathological examination revealed an extensive loss of Purkinje cells in the vast majority of cerebellar folia, suggesting that PrP plays a role in the long-term survival of Purkinje neurons.
  • Hitoshi Okada; Kuniaki Ishii; Kazuo Nunoki; Norio Taira
    BBA - Biomembranes ELSEVIER SCIENCE BV 1234 (1) 145 - 148 0005-2736 1995/03 
    Recently, pIcln has been reported to be a regulator of a swelling-induced chloride conductance. We have cloned a cDNA RCL-H1 from rabbit heart, of which primary structure is highly homologous to that of pIcln. Outwardly rectifying currents were recorded in oocytes expressing RCL-H1, which is consistent with the result of pIcln. RNA blot analysis revealed the widespread expression of RCL-H1 mRNA in rabbit tissues. RCL-H1 may play an important role in regulating cell volume and give a clue to revealing molecular structure of swelling-induced chloride channel(s). © 1995.
  • K ISHII; T YAMAGISHI; K NUNOKI; H OKADA; N TAIRA
    HEART AND VESSELS SPRINGER VERLAG 12 - 13 0910-8327 1995 [Refereed]
     
    The effects of lowering the external pH on the currents of the cloned cardiac K+ channels (Kv1.2 and Kv1.4) were investigated. The currents of Kv1.2 (I-K(V1.2)) and Kv1.4 (I-K(V1.4)) were suppressed by external protons in a concentration-dependent manner. However. the sensitivity to protons and the voltage-dependence of the block were different between Kv1.2 and Kv1.4. The block of I-K(V1.2) was voltage-dependent but that of I-K(V1.4) was not. I-K(V1.4) was much more sensitive to external protons than I-K(V1.Z) The tandem construct, in which the C-terminus of Kv1.4 was ligated to the N-terminus of Kv1.2, had intermediate characteristics between the parent K+ channels. To identify the amino acid residues responsible for the high sensitivity to pH of Kv1.4, a chimera in which the part of putative pore-forming region (S5-S6 linker) of Kv1.4 was replaced with that of Kv1.2 was made. Studies on the chimera localized the responsible amino acid residues within the first half of the S5-S6 linker region.
  • K. Nunoki; K. Ishii; H. Okada; T. Yamagishi; H. Murakoshi; N. Taira
    Journal of Biological Chemistry AMER SOC BIOCHEMISTRY MOLECULAR BIOLOGY INC 269 (39) 24138 - 24142 0021-9258 1994 [Refereed]
     
    We constructed tandem cDNA by linking the 5' end of a delayed rectifier- type (Kv1.2) clone to the 3' end of a transient-type (Kv1.4) K+ channel clone. Fusion genes were also constructed, consisting of Kv1.4 and mutants of Kv1.2, which have a single amino acid substitution in the S4-S5 loop. From electrophysiological characterization, it is likely that two pairs of tandem heterodimer constructs can form hybrid channels. In addition, it has been revealed that the wild-type hybrid channel shows a time constant of inactivation very similar to that observed in the homotetrameric Kv1.4 channel. Difference of inactivation kinetics between wild-type and mutant hybrid K+ channels suggests that not only the S4-S5 loop of Kv1.4 but also that of Kv1.2 can serve as the acceptor sites for the inactivation gates, and that all of four sets of loops should be functional for rapid inactivation. From these results, in the hybrid channels the structure and composition of the acceptor sites could be important factors for determining the rate of inactivation.
  • H Okada; K Ishii; K Nunoki; T Abe; N Taira
    Biochemical and biophysical research communications ACADEMIC PRESS INC JNL-COMP SUBSCRIPTIONS 189 (1) 430 - 6 0006-291X 1992/11 [Refereed]
     
    We have cloned a transient type K channel from rat heart (RH10) and coexpressed a metabotropic glutamate receptor (mGluR5) to study the functional modulation of RH10 coupled to the phosphatidylinositol (PI) hydrolysis. Stimulation of mGluR5 suppressed peak amplitude of RH10 current and affected voltage dependence of activation and inactivation of the channel.

Books etc

  • ジーンターゲティングの実際
    岡田 斉; 野田哲生 (Joint work)実験医学 1996

Conference Activities & Talks

  • エピジェネティクスから考える環境因子と肥満  [Invited]
    岡田 斉
    第18回 日本抗加齢医学会総会プログラム  2018/05
  • Role of histone demethylases in breast cancer development  [Not invited]
    Hitoshi Okada
    The 76th Annual Meeting of the Japanese Cancer Association  2017/09
  • マウスモデルを用いたDNA損傷応答制御による 治療関連白血病発症抑制の試み  [Not invited]
    岡田 斉
    第21回日本がん分子標的治療学会学術集会  2017/06
  • Attenuated DNA damage repair delays therapy-related malignancies in mouse model  [Not invited]
    Hitoshi Okada
    The 12th International Conference on Protein Phosphatase International Symposium  2016/10
  • Attenuated DNA damage repair delays therapy-related malignancies in mouse model  [Not invited]
    Hitoshi Okada
    The 75th Annual Meeting of the Japanese Cancer Association  2016/10
  • H3K9 histone demethylase, KDM4B, regulates mammary gland development  [Not invited]
    Hitoshi Okada
    2015/12
  • ヒストン脱メチル化酵素、KDM4B/JMJD2B、のがん化過程での役割  [Not invited]
    岡田 斉
    第74回日本癌学会学術総会  2015/10
  • H3K9 demethylase, KDM4B, regulates obesity in vivo.  [Not invited]
    Hitoshi Okada
    Cold Spring Harbor Laboratory Meeting, The Biology of Cancer: Microenvironment, Metastasis & Therapeutics.  2013/04
  • H3K9 histone demethylase, constitute a key component of the estrogen signaling pathway in vivo and in vitro  [Not invited]
    Hitoshi Okada
    AACR Conference: Breakthrough in Basic and Translational Cancer Research  2013/02
  • JMJD2B acts as a co-factor in estrogen receptor signaling and mammary gland development  [Not invited]
    Hitoshi Okada
    Cold Spring Harbor Laboratory Meeting, The Biology of Cancer: Microenvironment, Metastasis & Therapeutics  2011/04
  • HLA-B-associated Transcript 3, Bat3, is essential for p300-mediated acetylation of p53  [Not invited]
    Hitoshi Okada
    AACR meeting: Apoptosis  2007/03
  • Catastrophic cell death in mitosis  [Invited]
    Hitoshi Okada
    Ohio University  2005/05
  • Loss of survivin induces p53 and Bcl-2 independent catastrophic cell death in mitosis  [Invited]
    Hitoshi Okada
    Keystone meeting: Hematopoiesis  2005/03
  • Bat3 regulates male germ cell development  [Not invited]
    Hitoshi Okada
    AACR meeting: Regulation of Cell Death in Oncogenesis  2005/01
  • Apoptosis regulation in tumorigenesis and T cell development  [Invited]
    Hitoshi Okada
    Ohio State University  2005

MISC

Research Grants & Projects

  • 代謝ストレスにより誘導される新規肝がんモデルの作製と機能解析
    日本学術振興会：科学研究費助成事業 基盤研究(C)
    Date (from‐to) : 2021/04 -2024/03 
    Author : 岡田 斉
  • TGFbによるSirt1の制御と炎症性腸疾患の抑制機構の解明
    Japan Society for the Promotion of Science：Grants-in-Aid for Scientific Research Grant-in-Aid for Scientific Research (C)
    Date (from‐to) : 2019/04 -2022/03 
    Author : 天野 恭志; 岡田 斉
  • ヒト-細菌叢間 化学コミュニケーションの理解と炎症性腸疾患・がん・がん免疫
    Japan Society for the Promotion of Science：Grants-in-Aid for Scientific Research Grant-in-Aid for Scientific Research on Innovative Areas (Research in a proposed research area)
    Date (from‐to) : 2019/04 -2022/03 
    Author : 西尾 和人; 坂井 和子; 上嶋 一臣; 櫻井 俊治; 角田 郁生; 岡田 斉
     
    炎症性腸疾患（クローン病や過敏性大腸炎）および肝細胞癌患者の便中細菌叢のメタゲノム解析により、1）腸内細菌と腸上皮細胞のインタラクションが難治性炎症性腸疾患の発がんリスクに及ぼす影響の解明、2）免疫チェックポイント阻害薬による有害事象に及ぼす腸内細菌叢の影響、3)中枢神経系（CNS）炎症性疾患である多発性硬化症（MS）の発症に対する腸内細菌の影響を明らかにすることを目的とした。本年度は炎症性腸疾患4例、大腸がん110例の細菌叢のゲノム解析を行い、炎症性腸疾患症例におけるProteobacteriaの増加を検出した。潰瘍性大腸炎の病態の解明として、潰瘍性大腸炎関連大腸癌の発生に関連すると考えられる癌遺伝子ガンキリンについて大腸炎発生との関連を検討した結果、小腸粘膜でのガンキリン欠損により、大腸炎を増悪させることが明らかとなった。MSの発症と腸内細菌叢との関連について、マウス脳内にタイラーウイルスを接種するMS様病態を用いてCNSトランスクリプトーム解析と糞便を用いた腸内細菌叢解析を行った。CNS浸潤による炎症性脱髄病変の誘導に伴い、有意に腸内細菌叢の多様性増加が認められた。以上より、炎症性腸疾患ならびに中枢神経系炎症性疾患における宿主と細菌叢との化学コミュニケーションによる病態の解明は順調に進んでいる。
  • 代謝制御とがん組織微小環境を結ぶネットワーク機構の解明
    Japan Society for the Promotion of Science：Grants-in-Aid for Scientific Research Grant-in-Aid for Scientific Research (C)
    Date (from‐to) : 2018/04 -2021/03 
    Author : 岡田 斉; 天野 恭志; 古室 暁義; 上田 健; 太田 一成
     
    組織微小環境は、がん、生活習慣病をはじめとするヒト疾患の発症・進展・治療反応性に重要である。近年、タンパク質恒常性維持ネットワーク（PN）が代謝制御、炎症・免疫応答、小胞体ストレス応答シグナル経路と相互作用し、組織微小環境の構築と維持に寄与することが報告されている。BAT3は分子シャペロンとアポトーシス制御、プロテオソーム機能、免疫応答に関与するという一連の報告が我々と他のグループからなされている。しかしながら現在まで、BAT3の個体レベルでの機能は十分に解明されていない。 独自に作成したコンベンショナルノックアウトマウスを作成し解析した結果、Bat3のコンベンショナルノックアウトマウスは胎児性致死を引き起こすことが明らかとなった。そこで我々は、組織特異的BAT3ノックアウト（Bat3KO）マウスを独自に作成した。当該年度はC57BL/6J遺伝背景を有する肝特異的Bat3KOマウス（Bat3LKO: Bat3flox/flox;Albumin-Cre）と膵臓β細胞特異的Bat3KOマウス（Bat3BKO: Bat3flox/flox;Ins-Cre）の樹立を試みたが、順調にマウスの交配が進行し、それぞれのラインを樹立することができた。 Bat3の欠損が肝臓あるいは膵臓β細胞の機能にどのような影響を与えるかを明らかにする目的で、正常食、高グルコース食、高脂肪食で摂餌し、継続的な体重、随時血糖測定を行い、肥満と耐糖能異常の有無を比較した。また、それぞれの摂餌条件から得られるサンプルから抽出したRNAを用いて、遺伝子発現の網羅的解析を行い、Bat3欠損特異的に上昇あるいは低下している遺伝子群の候補を同定する予定である。さらに例数を増やし、これまで得られているデータと合わせて解析し、Bat3特異的な信号伝達系を同定することを予定している。
  • Elucidation of epigenetics-mediated glucose and lipid metabolism
    Japan Society for the Promotion of Science：Grants-in-Aid for Scientific Research Grant-in-Aid for Scientific Research (C)
    Date (from‐to) : 2018/04 -2019/03 
    Author : 太田 一成; 岡田 斉
     
    DNAのメチル化やヒストン翻訳後修飾といったエピジェネティクスによる脂質代謝疾患の制御機構は不明な点が多い。KDM6A/Utxはジメチル化・トリメチル化ヒストンH3K27 (H3K27me2/me3) を基質とするヒストン脱メチル化酵素であり、脂肪分化段階依存的に脂肪分化を制御することが報告されている。われわれは成熟脂肪細胞のUtxの機能に着目し、脂肪細胞特異的Utx欠損マウス（Utx cKOマウス）の解析を行った。 Utx cKOマウスは高脂肪食負荷で対照群に比べて体重が減少し、脂肪組織が縮小することを見出した。脂肪肝やインスリン抵抗性の進行が抑制されていた。網羅的遺伝子発現解析により、Utx cKOマウスの白色脂肪組織ではコレステロール・エストロゲン合成に関連する遺伝子発現が亢進していることが分かった。トリアシルグリセロール合成に関連する遺伝子発現は低下していた。 そこで卵巣由来のエストロゲンを取り除くために卵巣摘出を行ったところ、Utx cKOマウスでは同様に高脂肪食負荷による肥満が抑制された。Utx cKOマウスの内臓脂肪におけるエストロゲンの濃度は対照群に比べて上昇していた。Utx欠失白色脂肪細胞における局所的なエストロゲン産生亢進が、高脂肪食による肥満の抑制に寄与していることが示唆された。 以上の結果より、白色脂肪細胞におけるUtxが脂質代謝を制御し、肥満およびメタボリック症候群に関連することを明らかにした。Utxを介したエピゲノム制御が代謝疾患の治療標的となる可能性を示し、新規予防法・治療法開発への分子基盤を確立することができた。
  • 増殖シグナル制御機構の解明と革新的がん治療法の開発
    文部科学省：私立大学戦力的基盤形成支援事業
    Date (from‐to) : 2016 -2019 
    Author : 岡田 斉
  • Histone demethylase regulates mammary gland development and breast cancer development
    Japan Sciety for the Promotion of Science：Grant in Aid for Scientific Research
    Date (from‐to) : 2014 -2015 
    Author : Hitoshi Okada
  • トリプルネガティブ乳がん治療耐性獲得機序に関する知的基盤の確立
    金沢大学がん進展制御研究所：平成27 年度 金沢大学がん進展制御研究所 共同研
    Date (from‐to) : 2015 
    Author : 岡田 斉
  • To investigate role of Bat3 in nuclear oncogenic signaling networks
    Canadian Institute of Health Research：Operating grant
    Date (from‐to) : 2011 -2014 
    Author : Hitoshi Okada
  • エピジェネティクスによる老化制御機構の解明
    花王健康科学研究会：花王健康科学研究会助成金
    Date (from‐to) : 2014 
    Author : 岡田 斉
  • Establishment of in vivo and in vitro models for systematic tissue-specific epigenomic mapping
    Canadian Institute of Health Research：Epigenetics, Environment and Health (CEEHR) Catalyst Grant
    Date (from‐to) : 2012 -2013 
    Author : Hitoshi Okada
  • To investigate the roles of histone demethylases in estrogen receptor signaling Network
    Canadian Institute of Health Research：Operating grant
    Date (from‐to) : 2011 -2012 
    Author : Hitoshi Okada
  • Roles of heat-shock 70 family of proteins in breast cancer
    Canadian Breast Cancer Foundation：Operating grant
    Date (from‐to) : 2009 -2012 
    Author : Hitoshi Okada
  • Role of post-translational modifications of p53 in cancer pathogenesis
    Canadian Institute of Health Research：Operating grant
    Date (from‐to) : 2007 -2010 
    Author : Hitoshi Okada
  • Role of apoptosis signaling pathway in tumorigenesis
    University Health Network：Operating grant
    Date (from‐to) : 2005 -2007 
    Author : Hitoshi Okada
  • 哺乳類中枢神経系の発生におけるPOU転写因子の機能
    Japan Society for the Promotion of Science：Grants-in-Aid for Scientific Research Grant-in-Aid for Scientific Research on Priority Areas
    Date (from‐to) : 1996 -1996 
    Author : 美野輪 治; 岡田 斉; 高野 洋志
     
    本研究では、Class III POU転写調節因子群(Brn-1,Brn-2,Brn-4)の哺乳類の神経発生、特に中枢神経系の神経系譜の決定と制御における機能の解明を目的として、これらの遺伝子が欠損したマウスを樹立し、その解析を行っている。 Brn-1欠損マウスの解析からは中枢神経系の組織学的解析から海馬の層構造に異常があることが判明し、Brn-1が海馬の形成に重要な機能を有することが明らかとなったが、中枢神経系他の多くの領域では異常は明らかでなかった。また、生後48時間以内に死亡する原因が腎不全であることを突き止めた。腎臓でも、Brn-1が機能していることは興味深く、現在免疫組織化学等によりこれらの詳細な解析を行っている。 Brn-4欠損マウスは高度難聴となることが、生理学的解析(ABR,AP等)から明らかとなった。現在、聴覚系を中心に組織学的解析を行っている。 我々はさらにBrn-1欠損マウスとBrn-2欠損マウスの交配により両遺伝子を欠損したマウスを作成した。Brn-1,2二重欠損マウスは生直後に死亡し、中枢神経系の組織学的解析から、各遺伝子の単独欠損マウスでは認められない新たな異常を発見した。これはBrn-1,Brn-2遺伝子の機能的代償性を示すものでありその詳細な組織学的解析により、その詳細な機構が解明されることが期待される。 今後はBrn-1,Brn-2,Brn-4遺伝子の二重、三重欠損マウスを作製し解析することにより、これらの遺伝子の機能の全貌が明らかになるであろう。また、これらの各変異マウスから抽出したRNAを用いたDifferntial Display法により、クラスIII POU転写調節因子群の標的遺伝子の同定を試みる予定である。

Social Contribution

  • Date (from-to) : 2005/09-2013/09
    Role : Advisor
    Category : Others
    Sponser, Organizer, Publisher  : Campbell Family Institue for Breast Cancer Research, Ontario Cancer Institute
    Event, Program, Title : Weekend to End Women's Cancers Walks

Others

  • 2017/04 -2020/03  核酸医薬による変異K-RAS遺伝子発現制御と革新的抗癌薬の開発 
    近畿大学学内助成金 研究種目： 21世紀研究開発奨励金（共同研究助成） 課題番号： KD1704 研究内容： 本研究では核酸医薬を用いて変異KRAS遺伝子の発現を選択的に抑制することにより、あるいは、抗変異KRAS核酸医薬とセツキシマブ等のモノクロナール抗体医薬やエルロチニブ等の分子標的薬との併用により革新的な変異K-RAS依存性がんの治療薬開発を目指す。1塩基変異を敏感に認識できる核酸医薬を化学合成し、大腸がん由来細胞SW48及びHCT116中に発現する変異KRAS遺伝子の発現制御効果を評価する。次に、3次元培養法により癌細胞の増殖抑制効果を評価し、さらに、LSL-KRASマウスモデルを用いてin vivo試験を行う。

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