A ROSEN, Takafumi Noma, V WENDEL-HANSEN, J ZEUTHEN, C-Y LIN and T HONJO : In Vitro Immunization in Hybridoma Technology, --- A NEW PROGRESSION FACTOR FOR STIMULATION OF HUMAN B CELLS IN VITRO ---, Elsevier Science Publishers B.V., 1988.
28.
Hisataka Sabe, Shigeru Kondo, Takafumi Noma and Tasuku Honjo : Recent Advances in Leukemia and Lymphoma, --- MOLECULAR GENETIC STUDIES ON THE INTERLEUKIN 2 RECEPTOR ---, Alan R.Liss,Inc., 1987.
学術論文(審査論文):
1.
Intan Ruspita, Pragnya Das, Keiko Miyoshi, Takafumi Noma, L Malcolm Snead and Marianna Bei : Enam expression is regulated by Msx2., Developmental Dynamics, 2023.
(要約)
Collectively, these results illustrate that Enam gene expression is controlled by Msx2 in a spatio-temporal manner. They also suggest that Msx2 may interact with other transcription factors to control spatial and temporal expression of Enam and hence amelogenesis and enamel biomineralization.
Taigo Horiguchi, Ayako Tanimura, Keiko Miyoshi, Hiroko Hagita, Hisanori Minami and Takafumi Noma : The Effect of Heterozygous Mutation of Adenylate Kinase 2 Gene on Neutrophil Differentiation., International Journal of Molecular Sciences, Vol.23, 16089, 2022.
(要約)
Mitochondrial ATP production plays an important role in most cellular activities, including growth and differentiation. Previously we reported that Adenylate kinase 2 (AK2) is the main ADP supplier in the mitochondrial intermembrane space in hematopoietic cells, especially in the bone marrow. AK2 is crucial for the production of neutrophils and T cells, and its deficiency causes reticular dysgenesis. However, the relationship between ADP supply by AK2 and neutrophil differentiation remains unclear. In this study, we used CRISPR/Cas9 technology to establish two heterozygous AK2 knock-out HL-60 clones as models for reticular dysgenesis. Their AK2 activities were about half that in the wild-type (WT). Furthermore, neutrophil differentiation was impaired in one of the clones. In silico analysis predicted that the obtained mutations might cause a structural change in AK2. Time course microarray analysis of the WT and mutants revealed that similar gene clusters responded to all-trans retinoic acid treatment, but their expression was lower in the mutants than in WT. Application of fructose partially restored neutrophil differentiation in the heterozygous knock-out HL-60 clone after all-trans retinoic acid treatment. Collectively, our study suggests that the mutation of N-terminal region in AK2 might play a role in AK2-dependent neutrophil differentiation and fructose could be used to treat AK2 deficiency.
Susumu Tadokoro, Reiko Tokuyama-Toda, Seiko Tatehara, Shinji Ide, Hirochika Umeki, Keiko Miyoshi, Takafumi Noma and Kazuhito Satomura : A New Induction Method for the Controlled Differentiation of Human-Induced Pluripotent Stem Cells Using Frozen Sections, Cells, Vol.10(11), No.11, 2827, 2021.
(要約)
Considering that every tissue/organ has the most suitable microenvironment for its functional cells, controlling induced pluripotent stem cell (iPSC) differentiation by culture on frozen sections having a suitable microenvironment is possible. Induced PSCs were cultured on frozen sections of the liver, the brain, the spinal cord, and cover glasses (control) for 9 days. The iPSCs cultured on the sections of the liver resembled hepatocytes, whereas those on sections of the brain and the spinal cord resembled neuronal cells. The percentage of hepatocytic marker-positive cells in the iPSCs cultured on the sections of the liver was statistically higher than that of those in the iPSCs cultured on the sections of the brain and the spinal cord or on cover glasses. In contrast, the iPSCs cultured on the sections of the brain and the spinal cord revealed a high percentage of neural marker-positive cells. Thus, iPSCs can be differentiated into a specific cell lineage in response to specific factors within frozen sections of tissues/organs. Differentiation efficacy of the frozen sections markedly differed between the iPSC clones. Therefore, our induction method could be simple and effective for evaluating the iPSC quality.
Taigo Horiguchi, Yumiko Miyake, Keiko Miyoshi, Ayako Tanimura, Hiroko Hagita, Hiroshi Sakaue and Takafumi Noma : Gene-expression profile reveals the genetic and acquired phenotypes of hyperactive mutant SPORTS rad, The Journal of Medical Investigation : JMI, Vol.VOL67, No.NO1,2, 51-61, 2020.
(要約)
Spontaneously Running Tokushima Shikoku (SPORTS) rat is a hyperactive rat strain. However, the causative mutation of this phenotype has not yet been identified. To investigate the molecular basis for the unique phenotype of SPORTS rats, we examined gene-expression profiles by microarray analyses. Among adenylate kinase isozymes that maintain the homeostasis of cellular adenine nucleotide composition in the cell, only adenylate kinase 1 is highly up-regulated in both exercised and sedentary SPORTS rats compared with wild-type (WT) rats, 5.5-fold and 3.3-fold, respectively. Further comparative analyses revealed that genes involved in glucose metabolism were up-regulated in skeletal muscle tissue of exercised SPORTS rats compared with sedentary mutants, whereas genes related to extracellular matrix or region were down-regulated compared with WT rats. In brain tissue of sedentary SPORTS rats, genes associated with defense and catecholamine metabolism were highly expressed compared with WT rats. These findings suggest that genetic mutation(s) in SPORTS rat remodels metabolic demands through differentially regulating gene expression regardless of exercise. Therefore, the SPORTS rats are useful animal model not only for further examining the effects of exercise on metabolism but also for deeply studying the molecular basis how mutation affect the psychological motivation with spontaneous voluntary exercise phenotype. J. Med. Invest. 67 : 51-61, February, 2020.
Ayako Tanimura, Keiko Miyoshi, Taigo Horiguchi, Hiroko Hagita, Koichi Fujisawa and Takafumi Noma : Mitochondrial Activity and Unfolded Protein Response are Required for Neutrophil Differentiation., Cellular Physiology and Biochemistry, Vol.47, No.5, 1936-1950, 2018.
(要約)
In this study, we demonstrated that neutrophil differentiation is regulated by ER stress/UPR that is supported by mitochondrial ATP supply, in which IRE1-XBP1 activation is essential. Our findings provide the evidence that mitochondrial energy metabolism may play a critical role in neutrophil differentiation.
Yosi Dian Arinawati, Keiko Miyoshi, Ayako Tanimura, Taigo Horiguchi, Hiroko Hagita and Takafumi Noma : Deciphering defective amelogenesis using invitro culture systems., Journal of Bioscience and Bioengineering, Vol.125, No.Issue 4, 365-496, 2018.
(要約)
The conventional two-dimensional (2D) in vitro culture system is frequently used to analyze the gene expression with or without extracellular signals. However, the cells derived from primary culture and cell lines frequently deviate the gene expression profile compared to the corresponding in vivo samples, which sometimes misleads the actual gene regulation in vivo. To overcome this gap, we developed the comparative 2D and 3D in vitro culture systems and applied them to the genetic study of amelogenesis imperfecta (AI) as a model. Recently, we found specificity protein 6 (Sp6) mutation in an autosomal-recessive AI rat that was previously named AMI. We constructed 3D structure of ARE-B30 cells (AMI-derived rat dental epithelial cells) or G5 (control wild type cells) combined with RPC-C2A cells (rat pulp cell line) separated by the collagen membrane, while in 2D structure, ARE-B30 or G5 was cultured with or without the collagen membrane. Comparative analysis of amelogenesis-related gene expression in ARE-B30 and G5 using our 2D and 3D in vitro systems revealed distinct expression profiles, showing the causative outcomes. Bone morphogenetic protein 2 and follistatin were reciprocally expressed in G5, but not in ARE-B30 cells. All-or-none expression of amelotin, kallikrein-related peptidase 4, and nerve growth factor receptor was observed in both cell types. In conclusion, our in vitro culture systems detected the phenotypical differences in the expression of the stage-specific amelogenesis-related genes. Parallel analysis with 2D and 3D culture systems may provide a platform to understand the molecular basis for defective amelogenesis caused by Sp6 mutation.
Koichi Oshima, Norikazu Saiki, Michihiro Tanaka, Hiromi Imamura, Akira Niwa, Ayako Tanimura, Ayako Nagahashi, Akiyoshi Hirayama, Keisuke Okita, Akitsu Hotta, Shuichi Kitayama, Mitsujiro Osawa, Shin Kaneko, Akira Watanabe, Isao Asaka, Wataru Fujibuchi, Kohsuke Imai, Hiromasa Yabe, Yoshiro Kamachi, Junichi Hara, Seiji Kojima, Masaru Tomita, Tomoyoshi Soga, Takafumi Noma, Shigeaki Nonoyama, Tatsutoshi Nakahata and K Megumu Saito : Human AK2 links intracellular bioenergetic redistribution to the fate of hematopoietic progenitors., Biochemical and Biophysical Research Communications, Vol.497, No.2, 719-725, 2018.
(要約)
AK2 is an adenylate phosphotransferase that localizes at the intermembrane spaces of the mitochondria, and its mutations cause a severe combined immunodeficiency with neutrophil maturation arrest named reticular dysgenesis (RD). Although the dysfunction of hematopoietic stem cells (HSCs) has been implicated, earlier developmental events that affect the fate of HSCs and/or hematopoietic progenitors have not been reported. Here, we used RD-patient-derived induced pluripotent stem cells (iPSCs) as a model of AK2-deficient human cells. Hematopoietic differentiation from RD-iPSCs was profoundly impaired. RD-iPSC-derived hemoangiogenic progenitor cells (HAPCs) showed decreased ATP distribution in the nucleus and altered global transcriptional profiles. Thus, AK2 has a stage-specific role in maintaining the ATP supply to the nucleus during hematopoietic differentiation, which affects the transcriptional profiles necessary for controlling the fate of multipotential HAPCs. Our data suggest that maintaining the appropriate energy level of each organelle by the intracellular redistribution of ATP is important for controlling the fate of progenitor cells.
Koichi Fujisawa, Shuji Terai, Taro Takami, Naoki Yamamoto, Takahiro Yamasaki, Toshihiko Matsumoto, Kazuhito Yamaguch, Yuji Owada, Hiroshi Nishina, Takafumi Noma and Isao Sakaida : Modulation of anti-cancer drug sensitivity through the regulation of mitochondrial activity by adenylate kinase 4, Journal of Experimental & Clinical Cancer Research, Vol.35, No.1, 48, 2016.
(要約)
Background: Adenylate kinase is a key enzyme in the high-energy phosphoryl transfer reaction in living cells. Anisoform of this enzyme, adenylate kinase 4 (AK4), is localized in the mitochondrial matrix and is believed to beinvolved in stress, drug resistance, malignant transformation in cancer, and ATP regulation. However, the molecularbasis for the AK4 functions remained to be determined.Methods: HeLa cells were transiently transfected with an AK4 small interfering RNA (siRNA), an AK4 short hairpinRNA (shRNA) plasmid, a control shRNA plasmid, an AK4 expression vector, and a control expression vector toexamine the effect of the AK4 expression on cell proliferation, sensitivity to anti-cancer drug, metabolome, geneexpression, and mitochondrial activity.Results: AK4 knockdown cells treated with short hairpin RNA increased ATP production and showed greatersensitivity to hypoxia and anti-cancer drug, cis-diamminedichloro-platinum (II) (CDDP). Subcutaneous grafting AK4knockdown cells into nude mice revealed that the grafted cells exhibited both slower proliferation and reduced thetumor sizes in response to CDDP. AK4 knockdown cell showed a increased oxygen consumption rate with FCCPtreatment, while AK4 overexpression lowered it. Metabolome analysis showed the increased levels of thetricarboxylic acid cycle intermediates, fumarate and malate in AK4 knockdown cells, while AK4 overexpressionlowered them. Electron microscopy detected the increased mitochondrial numbers in AK4 knockdown cells.Microarray analysis detected the increased gene expression of two key enzymes in TCA cycle, succinatedehydrogenase A (SDHA) and oxoglutarate dehydrogenease L (OGDHL), which are components of SDH complexand OGDH complex, supporting the metabolomic results.Conclusions: We found that AK4 was involved in hypoxia tolerance, resistance to anti-tumor drug, and theregulation of mitochondrial activity. These findings provide a new potential target for efficient anticancer therapiesby controlling AK4 expression.
A Adiningrat, Ayako Tanimura, Keiko Miyoshi, Hiroko Hagita, RD Yanuaryska, Arinawati Yosi Dian, Taigo Horiguchi and Takafumi Noma : Isolation and characterization of dental epithelial cells derived from amelogenesis imperfecta rat., Oral Diseases, Vol.22, No.2, 132-139, 2016.
(要約)
OBJECTIVE:Disruption of the third zinc finger domain of specificity protein 6 (SP6) presents an enamel-specific defect in a rat model of amelogenesis imperfecta (AMI rats). To understand the molecular basis of amelogenesis imperfecta caused by the Sp6 mutation, we established and characterized AMI-derived rat dental epithelial (ARE) cells.MATERIALS AND METHODS:ARE cell clones were isolated from the mandibular incisors of AMI rats, and amelogenesis-related gene expression was analyzed by reverse transcription polymerase chain reaction (RT-PCR). Localization of wild-type SP6 (SP6WT) and mutant-type SP6 (SP6AMI) was analyzed by immunocytochemistry. SP6 transcriptional activity was monitored by rho-associated protein kinase 1 (Rock1) promoter activity with its specific binding to the promoter region in dental (G5 and ARE) and non-dental (COS-7) epithelial cells.RESULTS:Isolated ARE cells were varied in morphology and gene expression. Both SP6WT and SP6AMI were mainly detected in nuclei. The promoter analysis revealed that SP6WT and SP6AMI enhanced Rock1 promoter activity in G5 cells but that enhancement by SP6AMI was weaker, whereas no enhancement was observed in the ARE and COS-7 cells, even though SP6WT and SP6AMI bound to the promoter in all instances.CONCLUSION:ARE cell clones can provide a useful in vitro model to study the mechanism of SP6-mediated amelogenesis imperfecta.
(キーワード)
Sp6 mutation / amelogenesis imperfecta / in vitro disease model / tooth phenotype
Keiko Miyoshi, Taigo Horiguchi, Ayako Tanimura, Hiroko Hagita and Takafumi Noma : Gene Signature of Human Oral Mucosa Fibroblasts: Comparison with Dermal Fibroblasts and Induced Pluripotent Stem Cells., BioMed Research International, Vol.2015, 121575, 2015.
(要約)
Oral mucosa is a useful material for regeneration therapy with the advantages of its accessibility and versatility regardless of age and gender. However, little is known about the molecular characteristics of oral mucosa. Here we report the first comparative profiles of the gene signatures of human oral mucosa fibroblasts (hOFs), human dermal fibroblasts (hDFs), and hOF-derived induced pluripotent stem cells (hOF-iPSCs), linking these with biological roles by functional annotation and pathway analyses. As a common feature of fibroblasts, both hOFs and hDFs expressed glycolipid metabolism-related genes at higher levels compared with hOF-iPSCs. Distinct characteristics of hOFs compared with hDFs included a high expression of glycoprotein genes, involved in signaling, extracellular matrix, membrane, and receptor proteins, besides a low expression of HOX genes, the hDFs-markers. The results of the pathway analyses indicated that tissue-reconstructive, proliferative, and signaling pathways are active, whereas senescence-related genes in p53 pathway are inactive in hOFs. Furthermore, more than half of hOF-specific genes were similarly expressed to those of hOF-iPSC genes and might be controlled by WNT signaling. Our findings demonstrated that hOFs have unique cellular characteristics in specificity and plasticity. These data may provide useful insight into application of oral fibroblasts for direct reprograming.
Hiroshi Kondo, Keiko Miyoshi, Shoji Sakiyama, Akira Tangoku and Takafumi Noma : Differential Regulation of Gene Expression of Alveolar Epithelial Cell Markers in Human Lung Adenocarcinoma-Derived A549 Clones, Stem Cells International, Vol.2015, 165867, 2015.
(要約)
Stem cell therapy appears to be promising for restoring damaged or irreparable lung tissue. However, establishing a simple and reproducible protocol for preparing lung progenitor populations is difficult because the molecular basis for alveolar epithelial cell differentiation is not fully understood. We investigated an in vitro system to analyze the regulatory mechanisms of alveolus-specific gene expression using a human alveolar epithelial type II (ATII) cell line, A549. After cloning A549 subpopulations, each clone was classified into five groups according to cell morphology and marker gene expression. Two clones (B7 and H12) were further analyzed. Under serum-free culture conditions, surfactant protein C (SPC), an ATII marker, was upregulated in both H12 and B7. Aquaporin 5 (AQP5), an ATI marker, was upregulated in H12 and significantly induced in B7. When the RAS/MAPK pathway was inhibited, SPC and thyroid transcription factor-1 (TTF-1) expression levels were enhanced. After treatment with dexamethasone (DEX), 8-bromoadenosine 3'5'-cyclic monophosphate (8-Br-cAMP), 3-isobutyl-1-methylxanthine (IBMX), and keratinocyte growth factor (KGF), surfactant protein B and TTF-1 expression levels were enhanced. We found that A549-derived clones have plasticity in gene expression of alveolar epithelial differentiation markers and could be useful in studying ATII maintenance and differentiation.
Ryna Dwi Yanuaryska, Keiko Miyoshi, Arya Adiningrat, Taigo Horiguchi, Ayako Tanimura, Hiroko Hagita and Takafumi Noma : Sp6 regulation of Rock1 promoter activity in dental epithelial cells, The Journal of Medical Investigation : JMI, Vol.VOL61, No.(NO 3,4), 306-317, 2014.
(要約)
Sp6 is a transcription factor of the SP/KLF family and an indispensable regulator of the morphological dynamics of ameloblast differentiation during tooth development. However, the underlying molecular mechanisms remain unclear. We have previously identified one of the Sp6 downstream genes, Rock1, which is involved in ameloblast polarization. In this study, we investigated the transcriptional regulatory mechanisms of Rock1 by Sp6. First, we identified the transcription start sites (TSS) and cloned the 5'-flanking region of Rock1. Serial deletion analyses identified a critical region for Rock1 promoter activity within the 249-bp upstream region of TSS, and chromatin immunoprecipitation assays revealed Sp6-binding to this region. Subsequent transient transfection experiments showed that Rock1 promoter activity is enhanced by Sp6, but reduced by Sp1. Treatment of dental epithelial cells with the GC-selective DNA binding inhibitor, mithramycin A, affected Rock1 promoter activity in loss of enhancement by Sp6, but not repression by Sp1. Further site-directed mutagenesis indicated that the region from -206 to -150 contains responsive elements for Sp6. Taken together, we conclude that Sp6 positively regulates Rock1 transcription by direct binding to the Rock1 promoter region from -206 to -150, which functionally distinct from Sp1.
Kosuke Higashi, Shoichi Hazama, Atsuhiro Araki, Kiyoshi Yoshimura, Norio Iizuka, Shigefumi Yoshino, Takafumi Noma and Masaaki Oka : A novel cancer vaccine strategy with combined IL-18 and HSV-TK gene therapy driven by the hTERT promoter in a murine colorectal cancer model., International Journal of Oncology, Vol.45, No.4, 1412-1420, 2014.
(要約)
A therapeutic vaccine against minimal residual cancer cells is needed for the treatment of patients with colorectal cancer. Several gene therapy studies have revealed that the combination of a suicide gene and cytokine gene might induce effective antitumor immunity. In this study, we constructed an interleukin (IL)-18 and herpes simplex virus-thymidine kinase (HSV-TK) expression vector driven by the human telomerase reverse transcriptase (hTERT) promoter to study the efficacy of combination gene therapy with IL-18 and the HSV-TK suicide gene. Low immunogenic colon 26 cells were used for transfection and inoculation into syngeneic BALB/c mice. Large established tumors of colon 26 transfectants expressing IL-18 and HSV-TK driven by the hTERT promoter were completely eradicated after GCV administration in syngeneic BALB/c mice. Immunohistochemical analysis at the tumor rejection sites revealed enormous infiltrations of CD8+ T lymphocytes as well as CD4+ T lymphocytes and CD11b+ monocytes. Moreover, established distant tumors were completely eradicated by vaccination with the IL-18 and HSV-TK transfectants in combination with GCV. These data suggest that the IL-18 and suicide gene therapy can elicit antitumor specific immunity. In conclusion, gene therapy with IL-18 and HSV-TK plasmid vector driven by the hTERT promoter may be useful for cancer vaccination.
Ayako Tanimura, Taigo Horiguchi, Keiko Miyoshi, Hiroko Hagita and Takafumi Noma : Differential expression of adenine nucleotide converting enzymes in mitochondrial intermembrane space: a potential role of adenylate kinase isozyme 2 in neutrophil differentiation., PLoS ONE, Vol.9, No.2, e89916, 2014.
(要約)
Adenine nucleotide dynamics in the mitochondrial intermembrane space (IMS) play a key role in oxidative phosphorylation. In a previous study, Drosophila adenylate kinase isozyme 2 (Dak2) knockout was reported to cause developmental lethality at the larval stage in Drosophila melanogaster. In addition, two other studies reported that AK2 is a responsible gene for reticular dysgenesis (RD), a human disease that is characterized by severe combined immunodeficiency and deafness. Therefore, mitochondrial AK2 may play an important role in hematopoietic differentiation and ontogenesis. Three additional adenine nucleotide metabolizing enzymes, including mitochondrial creatine kinases (CKMT1 and CKMT2) and nucleoside diphosphate kinase isoform D (NDPK-D), have been found in IMS. Although these kinases generate ADP for ATP synthesis, their involvement in RD remains unclear and still an open question. In this study, mRNA and protein expressions of these mitochondrial kinases were firstly examined in mouse ES cells, day 8 embryos, and 7-week-old adult mice. It was found that their expressions are spatiotemporally regulated, and Ak2 is exclusively expressed in bone marrow, which is a major hematopoietic tissue in adults. In subsequent experiments, we identified increased expression of both AK2 and CKMT1 during macrophage differentiation and exclusive production of AK2 during neutrophil differentiation using HL-60 cells as an in vitro model of hematopoietic differentiation. Furthermore, AK2 knockdown specifically inhibited neutrophil differentiation without affecting macrophage differentiation. These data suggest that AK2 is indispensable for neutrophil differentiation and indicate a possible causative link between AK2 deficiency and neutropenia in RD.
Arya Adiningrat, Ayako Tanimura, Keiko Miyoshi, Ryna Yanuaryska Dwi, Hiroko Hagita, Taigo Horiguchi and Takafumi Noma : Ctip2-mediated Sp6 transcriptional regulation in dental epithelium-derived cells., The Journal of Medical Investigation : JMI, Vol.61, No.1.2, 126-136, 2014.
(要約)
Tooth development relies on the interaction between the oral ectoderm and underlying mesenchyme, and is regulated by a complex genetic cascade. This transcriptional cascade is regulated by the spatiotemporal activation and deactivation of transcription factors. The specificity proteins 6 (Sp6) and chicken ovalbumin upstream promoter transcription factor-interacting protein 2 (Ctip2) were identified in loss-of-function studies as key transcription factors required for tooth development. Ctip2 binds to the Sp6 promoter in vivo; however, its role in Sp6 expression remains unclear. In this study, we investigated Sp6 transcriptional regulation by Ctip2. Immunohistochemical analysis revealed that Sp6 and Ctip2 colocalize in the rat incisor during tooth development. We examined whether Ctip2 regulates Sp6 promoter activity in dental epithelial cells. Cotransfection experiments using serial Sp6 promoter-luciferase constructs and Ctip2 expression plasmids showed that Ctip2 significantly suppressed the Sp6 second promoter activity, although the Sp6 first promoter activity was unaffected. Ctip2 was able to bind to the proximal region of the Sp6 first promoter, as previously demonstrated, and also to the novel distal region of the first, and second promoter regions. Our findings indicate that Ctip2 regulates Sp6 gene expression through direct binding to the Sp6 second promoter region. J. Med. Invest. 61: 126-136, February, 2014.
Taigo Horiguchi, Miyuki Fuka, Koichi Fujisawa, Ayako Tanimura, Keiko Miyoshi, Ryutaro Murakami and Takafumi Noma : Adenylate kinase 2 deficiency limits survival and regulates various genes during larval stages of Drosophila melanogaster., The Journal of Medical Investigation : JMI, Vol.61, No.1.2, 137-150, 2014.
(要約)
Adenylate kinase isozyme 2 (AK2) is located in mitochondrial intermembrane space and regulates energy metabolism by reversibly converting ATP and AMP to 2 ADPs. We previously demonstrated that disruption of the Drosophila melanogaster AK2 gene (Dak2) resulted in growth arrest during the larval stage and subsequent death. Two other groups found that human AK2 mutations cause reticular dysgenesis, a form of severe combined immunodeficiency (SCID) that is associated with severe hematopoietic defects and sensorineural deafness. However, the mechanisms underlying differential outcomes of AK2 deficiency in Drosophila and human systems remain unknown. In this study, effects of tissue-specific inactivation of the Dak2 gene on Drosophila development were analyzed using RNAi-mediated gene knockdown. In addition, to investigate the roles of AK2 in the regulation of gene expression during development, microarray analysis was performed using RNA from first and second instar larvae of Dak2-deficient mutant and wild-type D. melanogaster. Knockdown of Dak2 in all germ layers caused cessation of growth and subsequent death of flies. Microarray analysis revealed that Dak2 deficiency downregulates various genes, particularly those involved in the proteasomal function and in mitochondrial translation machinery. These data indicate that adenine nucleotide interconversion by Dak2 is crucial for developmental processes of Drosophila melanogaster. J. Med. Invest. 61: 137-150, February, 2014.
Taro Muto, Keiko Miyoshi, Taigo Horiguchi, Hiroko Hagita and Takafumi Noma : Novel genetic linkage of rat Sp6 mutation to Amelogenesis imperfecta, Orphanet Journal of Rare Diseases, Vol.7, No.1, 2012.
(要約)
ABSTRACT: BACKGROUND: Amelogenesis imperfecta (AI) is an inherited disorder characterized by incomplete formation of tooth enamel. Although several genes responsible for AI have been reported, not all causative genes for human AI have been identified to date. AMI rat has been reported as an autosomal recessive mutant with hypoplastic AI isolated from a colony of stroke-prone spontaneously hypertensive rat strain, but the causative gene has not yet been clarified. Through a genetic screen, we identified the causative gene of autosomal recessive AI in AMI and analyzed its role in amelogenesis. METHODS: cDNA sequencing of possible AI-candidate genes so far identified using total RNA of day 6 AMI rat molars identified a novel responsible mutation in specificity protein 6 (Sp6). Genetic linkage analysis was performed between Sp6 and AI phenotype in AMI. To understand a role of SP6 in AI, we generated the transgenic rats harboring Sp6 transgene in AMI (Ami/Ami + Tg). Histological analyses were performed using the thin sections of control rats, AMI, and Ami/Ami + Tg incisors in maxillae, respectively. RESULTS: We found the novel genetic linkage between a 2-bp insertional mutation of Sp6 gene and the AI phenotype in AMI rats. The position of mutation was located in the coding region of Sp6, which caused frameshift mutation and disruption of the third zinc finger domain of SP6 with 11 cryptic amino acid residues and a stop codon. Transfection studies showed that the mutant protein can be translated and localized in the nucleus in the same manner as the wild-type SP6 protein. When we introduced the CMV promoter-driven wild-type Sp6 transgene into AMI rats, the SP6 protein was ectopically expressed in the maturation stage of ameloblasts associated with the extended maturation stage and the shortened reduced stage without any other phenotypical changes. CONCLUSION: We propose the addition of Sp6 mutation as a new molecular diagnostic criterion for the autosomal recessive AI pateints. Our findings expand the spectrum of genetic causes of autosomal recessive AI and sheds light on the molecular diagnosis for the classification of AI. Furthermore, tight regulation of the temporospatial expression of SP6 may have critical roles in completing amelogenesis. .
Taro Muto, Keiko Miyoshi, Taigo Horiguchi and Takafumi Noma : Dissection of morphological and metabolic differentiation of amelobrasts via ectopic SP6 expression, The Journal of Medical Investigation : JMI, Vol.59, No.1-2, 59-68, 2012.
(要約)
Tooth enamel is the hardest organ in the body. In rodent incisor, the enamel is exclusively produced by ameloblasts with yellowish-brown pigmentation, indicating normal enamel formation. However, the molecular mechanisms of ameloblast differentiation and amelogenesis are not fully understood. Specificity protein (Sp) 6 has been reported as one of the critical factors for tooth development. To explore SP6 function, we generated Sp6 transgenic (Tg) rats. Unexpectedly, the enamel surfaces of the incisors in Tg rats were discolored, even though enamel formation and serum iron concentrations were normal. Histological analysis of incisors from 6-week-old Tg rats demonstrated that the ameloblast layer at the pigmentation stage was elongated up to the gingival margin with ectopic SP6 expression in longitudinal incisor sections. In contrast, the incisors from 10-week-old Tg rats revealed that the pigmented ameloblasts were morphologically changed to those of the reduced stage, concomitant with the sporadic disappearance of ectopic SP6 expression. Here we report that morphological differentiation and metabolism of the iron-containing pigment in ameloblasts are independently regulated during amelogenesis by means of ectopic SP6 expression.
W Trianna Utami, Keiko Miyoshi, Hiroko Hagita, Dwi Ryna Yanuaryska, Taigo Horiguchi and Takafumi Noma : Possible Linkage of SP6 Transcriptional Activity with Amelogenesis by Protein Stabilization., Journal of Biomedicine & Biotechnology, Vol.2011, 2011.
(要約)
Ameloblasts produce enamel matrix proteins such as amelogenin, ameloblastin, and amelotin during tooth development. The molecular mechanisms of ameloblast differentiation (amelogenesis) are currently not well understood. SP6 is a transcription factor of the Sp/KLF family that was recently found to regulate cell proliferation in a cell-type-specific manner. Sp6-deficient mice demonstrate characteristic tooth anomalies such as delayed eruption of the incisors and supernumerary teeth with disorganized amelogenesis. However, it remains unclear how Sp6 controls amelogenesis. In this study, we used SP6 high producer cells to identify SP6 target genes. Based on the observations that long-term culture of SP6 high producer cells reduced SP6 protein expression but not Sp6 mRNA expression, we found that SP6 is short lived and specifically degraded through a proteasome pathway. We established an in vitro inducible SP6 expression system coupled with siRNA knockdown and found a possible linkage between SP6 and amelogenesis through the regulation of amelotin and Rock1 gene expression by microarray analysis. Our findings suggest that the regulation of SP6 protein stability is one of the crucial steps in amelogenesis.
Trianna W Utami, Keiko Miyoshi, Hiroko Hagita, Ryna D. Yanuaryska, Taigo Horiguchi and Takafumi Noma : Dynamic changes of Sp6 Transgene Expression in Dental Epithelial Cells during Long-term Culture, The Indonesian Journal of Dental Research, Vol.1, No.3, 2011.
Ivan Arie Wahyudi, Taigo Horiguchi, Keiko Miyoshi, Taro Muto, Trianna Wahyu Utami, Hiroko Hagita and Takafumi Noma : Isolation and Characterization of Mouse Specificity Protein 6 Promoter, The Indonesian Journal of Dental Research, Vol.2010, Volume 1, No.1, 21-34, 2010.
(要約)
Specificity protein 6 (SP6) is a member of the SP/Krüppel-like transcription factor family and plays key roles in tooth development. To study its biological roles, it is important to understand the spatiotemporal regulation of Sp6 gene expression. For this purpose, we first identified two separate 5' ends of the Sp6 cDNA by 5' RACE analysis using mouse mandibular RNA. Next, we isolated mouse genomic DNA fragments covering the Sp6 gene including two putative mouse Sp6 promoter regions and generated a series of luciferase reporter constructs. We confirmed the activity of both promoters by a luciferase assay and found strong second promoter activity in dental epithelial cells. Unexpectedly, we also detected potential third promoter activity in the intron 2 of the Sp6 gene. Last, we also found that bone morphogenetic protein and wingless signals could enhance Sp6 promoter activity in dental epithelial cells, suggesting the regulatory roles of two cytokines in Sp6 gene expression during tooth development. Our findings may shed new light on the regulatory mechanisms of Sp6 gene expression and provide a possible linkage between cytokine regulation of Sp6 expression and inductive epithelial and mesenchymal interactions.
Keiko Miyoshi, Daisuke Tsuji, Keiko Kudoh, Kazuhito Satomura, Taro Muto, Kouji Itou and Takafumi Noma : Generation of human induced pluripotent stem cells from oral mucosa, Journal of Bioscience and Bioengineering, Vol.110, No.3, 345-350, 2010.
(要約)
Induced pluripotent stem (iPS) cells are one of the most promising sources for cell therapy in regenerative medicine. Using a patient's own genetically identical and histocompatible cells is the ideal way to practice personalized regenerative medicine. For personalized iPS cell therapy, the prerequisites for cell source preparation are a simple and safe procedure, no aesthetic or functional damage, and quick wound healing. Oral mucosa fibroblasts (OFs) may have high potential to fulfill these requirements. In this study, biopsy was performed in a dental chair; no significant incisional damage was recognized and rapid wound healing (within a week) was observed. We generated human iPS cells from the isolated OFs via the retroviral gene transfer of OCT4, SOX2, c-MYC, and KLF4. Reprogrammed cells showed ES-like morphology and expressed undifferentiated markers such as OCT4, NANOG, SSEA4, TRA-1-60, and TRA-1-81. Subsequent in vitro and in vivo analyses confirmed the pluripotency of resultant iPS cells, which matched the criteria for iPS cells. In addition, we found that the endogenous expression levels of c-MYC and KLF4 in OFs were similar to those in dermal fibroblasts. Taken together, we propose that OFs could be a practical source for preparing iPS cells to achieve personalized regenerative medicine in the near future.
Koichi Fujisawa, Ryutaro Murakami, Taigo Horiguchi and Takafumi Noma : Adenylate kinase isozyme 2 is essential for growth and development of Drosophila melanogaster., Comparative Biochemistry and Physiology. Part B, Biochemistry & Molecular Biology, Vol.153, No.1, 29-38, 2009.
(要約)
Adenylate kinases are phylogenetically widespread, highly conserved, and involved in energy metabolism and energy transfer. Of these, adenylate kinase (AK) isozyme 2 is uniquely localized in the mitochondrial intermembrane space and its physiological role remains largely unknown. In this study, we selected Drosophila melanogaster to analyze its role in vivo. AK isozyme cDNAs were cloned and their gene expressions were characterized in D. melanogaster. The deduced amino acid sequences contain highly conserved motifs for P-loop, NMP binding, and LID domains of AKs. In addition, the effects of AK2 gene knockout on phenotype of AK2 mutants were examined using P-element technology. Although homozygous AK2 mutated embryos developed without any visible defects, their growth ceased and they died before reaching the third instar larval stage. Maternally provided AK2 mRNA was detected in fertilized eggs, and weak AK2 activity was observed in first and second instar larvae of the homozygous AK2 mutants, suggesting that maternally provided AK2 is sufficient for embryonic development. Disappearance of AK2 activity during larval stages resulted in growth arrest and eventual death. These results demonstrate that AK2 plays a critical role in adenine nucleotide metabolism in the mitochondrial intermembrane space and is essential for growth in D. melanogaster.
Keiko Miyoshi, Hideya Nagata, Taigo Horiguchi, Kaori Abe, Ivan Wahyudi Arie, Yoshinobu Baba, Hidemitsu Harada and Takafumi Noma : BMP2-induced gene profiling in dental epithelial cell line., The Journal of Medical Investigation : JMI, Vol.55, No.3-4, 216-226, 2008.
(要約)
Tooth development is regulated by epithelial-mesenchymal interactions and their reciprocal molecular signaling. Bone morphogenetic protein 2 (BMP2) is known as one of the inducers for tooth development. To analyze the molecular mechanisms of BMP2 on ameloblast differentiation (amelogenesis), we performed microarray analyses using rat dental epithelial cell line, HAT-7. After confirming that BMP2 could activate the canonical BMP-Smads signaling in HAT-7 cells, we analyzed the effects of BMP2 on 14,815 gene expressions and profiled them. Seventy-three genes were up-regulated and 28 genes were down-regulated by BMP2 treatment for 24 hours in HAT-7 cells. Functional classification revealed that 18% of up-regulated genes were ECM/adhesion molecules present in the enamel organ. Furthermore, we examined the expression of several differentiation markers in dental epithelial four cell-lineages including inner enamel epithelium (ameloblasts), stratum intermedium, stratum reticulum, and outer enamel epithelium. The results indicated that BMP2 might induce at least two different cell-lineage markers including a BMP antagonist expressed in HAT-7 cells, suggesting that BMP2 could accelerate amelogenesis via BMP signaling.
(キーワード)
Ameloblasts / Amelogenesis / Animals / Base Sequence / Bone Morphogenetic Protein 2 / Cell Line / DNA Primers / Down-Regulation / Epithelial Cells / Gene Expression Profiling / Oligonucleotide Array Sequence Analysis / Rats / Reverse Transcriptase Polymerase Chain Reaction / Signal Transduction / Smad Proteins / Up-Regulation
Two days training camp for the fourth year students of dental school was executed in 2006 and 2007.The purpose of this program is to develop good practical dental doctors through the promotion ofcommunication skills. The training camp was held at the YMCA oceanic center in Anan city of TokushimaPrefecture. It began with an oceanic program under relaxed atmosphere, and then, the programscontaining games, lectures, and workshops related to communication skills were carried out. The theme "support for career formation as a medical profession" was added to "promotion of communication skills" in 2007,and then the skills to learn the definition and the practice of the medical team treatment were installed andexecuted.The usefulness of methodology different from the regular course class was verified based on the results,and the possibility of the application in the future curriculum for the career formation was discussed in thepaper.平成18 年度と19 年度に，歯学部4年生に対して1泊2日の合宿研修を正課外授業として実施した．本事業は，コミュニケーション能力の開発を通じてより良き医療人を育成することが目的である．合宿研修場所は徳島県内の海洋センターであった．リラックスした雰囲気で海洋プログラムからスタートし，次いで，コミュニケーションに対する導入的なゲームや講演，そしてワークショップにより自分たち自身で問題点を抽出し，さらに解決するための討論を行い，その結果をプレゼンテーションをするという内容であった．平成19 年度の2回目には，医学部と薬学部の協力を得て，実施テーマとして「コミュニケーション能力の育成」に「医療専門職としてのキャリア形成支援」という新たなテーマを加え，チーム医療の定義や実践を学ぶ機会を設けて実施した．従来の正課授業と異なった教育方法の可能性について成果を基に検証し，今後のキャリア形成支援教育への応用の可能性について考察した．
Intan Ruspita, Keiko Miyoshi, Taro Muto, Kaori Abe, Taigo Horiguchi and Takafumi Noma : Sp6 downregulation of follistatin gene expression in ameloblasts., The Journal of Medical Investigation : JMI, Vol.55, No.1-2, 87-98, 2008.
(要約)
Sp6 is a member of the Sp family of transcription factors that regulate a wide range of cellular functions, such as cell growth and differentiation. Sp6, also called epiprofin, is specifically expressed in tooth germ, limb bud, and hair follicle, but there is little information on its function.To investigate the possible role of Sp6 in tooth development, first we established an Sp6-overproducing clone, CHA9, and analyzed the features of the cell, including cell proliferation and gene expression. The parental cells of CHA9 are the ameloblast-lineage G5 cells that we previously established from rat dental epithelia of lower incisor. Sp6 overproduction accelerated cell proliferation and induced the expression of ameloblastin mRNA, a marker of ameloblast differentiation. Second, we performed genome-wide screening of Sp6 target genes by microarray analysis. Out of a total 20,450 genes, 448 genes were up-regulated and 500 genes were down-regulated by Sp6. We found the expression of follistatin, a BMP antagonist, to be 22.4-fold lower in CHA9 than in control cells. Transfection of the Sp6-antisense construct into CHA9 cells restored follistatin expression back to equivalent levels seen in control cells, indicating that Sp6 regulates follistatin gene expression in ameloblasts.Our findings demonstrate that the follistatin gene is one of the Sp6 target genes in ameloblasts and suggest that Sp6 promotes amelogenesis through inhibition of follistatin gene expression.
Taro Muto, Keiko Miyoshi, Seiichi Munesue, Hiroshi Nakada, Minoru Okayama, Takashi Matsuo and Takafumi Noma : Differential expression of syndecan isoforms during mouse incisor amelogenesis., The Journal of Medical Investigation : JMI, Vol.54, No.3-4, 331-339, 2007.
(要約)
Syndecans are transmembranous heparan sulfate proteoglycans (HSPGs) with covalently attached glycosaminoglycan side-chains located on the cell surface. The mammalian syndecan family is composed of four types of syndecans (syndecan-1 to -4). Syndecans interact with the intracellular cytoskeleton through the cytoplasmic domains of their core proteins and membrane proteins, extracellular enzymes, growth factors, and matrix components, through their heparan-sulfate chains, to regulate developmental processes.Here, as a first step to assess the possible roles of syndecan proteins in amelogenesis, we examined the expression patterns of all syndecan isoforms in continuously growing mouse incisors, in which we can overview major differentiation stages of amelogenesis at a glance. Understanding the expression domain of each syndecan isoform during specific developmental stages seems useful for investigating their physiological roles in amelogenesis. Immunohistochemical analysis of syndecan core proteins in the lower incisors from postnatal day 1 mice revealed spatially and temporally specific expression patterns, with syndecan-1 expressed in undifferentiated epithelial and mesenchymal cells, and syndecan-2, -3, and -4 in more differentiated cells. These findings suggest that each syndecan isoform functions distinctly during the amelogenesis of the incisors of mice.
Kaori Abe, Keiko Miyoshi, Taro Muto, Ruspita Intan, Taigo Horiguchi, Toshihiko Nagata and Takafumi Noma : Establishment and characterization of rat dental epithelial derived ameloblast-lineage clones., Journal of Bioscience and Bioengineering, Vol.103, No.5, 479-485, 2007.
(要約)
Teeth are the hardest tissues covered with enamel produced by ameloblasts. The ameloblast differentiation is controlled by sequential epithelial-mesenchymal interactions during tooth morphogenesis. However, the molecular mechanism of ameloblast differentiation remains unclear. To address this question, we developed an in vitro assay system to evaluate the molecular mechanism of amelogenesis. First, we established dental epithelium-derived clones from 6-day-old rat incisors and established that cells of the clone SRE-G5 were the largest producers of amelogenin mRNA. Next, we analyzed the effects of several chemicals on the amelogenin expression in SRE-G5 cells. Only mitogen-activated protein kinase (MAPK) activators enhanced amelogenin mRNA expression. This finding corresponded to the immunohistochemical data showing the presence of phosphorylated forms of p38, c-Jun N-terminal kinase (JNK), and extracellular signal-regulated kinase (ERK) during ameloblast differentiation. To examine the roles of MAPK signals, we compared the effects of anisomycin and sodium salicylate on the expression of tooth-related differentiation markers. Both anisomycin and sodium salicylate induced amelogenin, Abcg2, and Bmp4 mRNA and down-regulated p75NGFR mRNA. On the other hand, ALP, ectodin, Bmp2 and Fgf8 mRNA were up-regulated only by anisomycin. These results indicate that MAPK signaling functions, at least in part, as the inducer of ameloblast differentiation.
Thrombospondin 1 (TSP1) is a multifunctional extracellular glycoprotein present mainly in the fetal and adult skeleton. Although an inhibitory effect of TSP1 against pathological mineralization in cultured vascular pericytes has been shown, its involvement in physiological mineralization by osteoblasts is still unknown. To determine the role of TSP1 in biomineralization, mouse osteoblastic MC3T3-E1 cells were cultured in the presence of antisense phosphorothioate oligodeoxynucleotides complementary to the TSP1 sequence. The 18- and 24-mer antisense oligonucleotides caused concentration-dependent increases in the number of mineralized nodules, acid-soluble calcium deposition in the cell/matrix layer, and alkaline phosphatase activity within 9 days, without affecting cell proliferation. The corresponding sense or scrambled oligonucleotides did not affect these parameters. In the antisense oligonucleotide-treated MC3T3-E1 cells, thickened extracellular matrix, well-developed cell processes, increased intracellular organelles, and collagen fibril bundles were observed. On the other hand, the addition of TSP1 to the culture decreased the production of a mineralized matrix by MC3T3-E1 cells. Furthermore, MC3T3-E1 clones overexpressing mouse TSP1 were established and assayed for TSP1 protein and their capacity to mineralize. TSP1 dose-dependently inhibited mineralization by these cells both in vitro and in vivo. These results indicate that TSP1 functions as an inhibitory regulator of bone mineralization and matrix production by osteoblasts to sustain bone homeostasis.
Akemichi Ueno, Kikuji Yamashita, Keiko Miyoshi, Taigo Horiguchi, Intan Ruspita, Kaori Abe and Takafumi Noma : Soluble matrix from osteoblastic cells induces mineralization by dental pulp cells, The Journal of Medical Investigation : JMI, Vol.53, No.3-4, 297-302, 2006.
(要約)
Dental pulp cells have a capacity to differentiate into mineralization-inducing cells. To clarify the molecular mechanism, we established an in vitro mineralization-inducing system by rat clonal dental pulp cell line, RPC-C2A, and tried to purify a mineralization-inducing factor in conditioned medium (CM) from pre-osteoblastic MC3T3-E1 cells. The active factor was impermeable to an ultrafiltration membrane, and sedimented by ultracentrifugation. The sedimented factor was found as a needle-like structure about 1.3 microm in average length as observed by transmission electron microscopy. The factor contained type I collagen, suggesting not a matrix vesicle, but a soluble matrix. The mineralization-inducing activity was also detected in CM from primary culture of rat calvaria (RC) cells. These results suggested that the soluble matrices from osteoblastic cells serve, at least in part, as differentiation-inducing agents.
(キーワード)
Animals / Calcification, Physiologic / Cell Line / Collagen Type I / Dental Pulp / 細胞外マトリックス (extracellular matrix) / Extracellular Matrix Proteins / Osteoblasts / Osteogenesis / Rats / Skull
Hiroki Aoki, Peter M. Kang, James Hampe, Koichi Yoshimura, Takafumi Noma, Masunori Matsuzaki and Seigo Izumo : Direct activation of mitovhondrial apoptosis machinery by c-jun N-terminal kinase in adult cardiac myocytes, The Journal of Biological Chemistry, Vol.277, No.12, 10244-10250, 2002.
(要約)
Although oxidative stress causes activation of c-Jun N-terminal kinase (JNK) and apoptosis in many cell types, how the JNK pathway is connected to the apoptosis pathway is unclear. The molecular mechanism of JNK-mediated apoptosis was investigated in adult rat cardiac myocytes in culture as a model system that is sensitive to oxidative stress. Oxidative stress caused JNK activation, cytochrome c release, and apoptosis without new protein synthesis. Oxidative stress-induced apoptosis was abrogated by dominant negative stress-activated protein kinase/extracellular signal-regulated kinase kinase-1 (SEK1)-mediated inhibition of the JNK pathway, whereas activation of the JNK pathway by constitutively active SEK1 was sufficient to cause apoptosis. Inhibition of caspase-9, an apical caspase in the mitochondrial apoptosis pathway, suppressed oxidative stress-induced apoptosis, whereas inhibition of caspase-8 had no effect, indicating that both the JNK pathway and the mitochondrial apoptosis machinery are central to oxidative stress-induced apoptosis. Both JNK and SEK1 localized on mitochondria where JNK was activated by oxidative stress. Furthermore, active JNK caused the release of apoptogenic factors such as cytochrome c from isolated mitochondria in a cell-free assay. These findings indicate that the JNK pathway is a direct activator of mitochondrial death machinery without other cellular components and provide a molecular linkage from oxidative stress to the mitochondrial apoptosis machinery.
(キーワード)
Adenoviridae / Animals / アポトーシス (apoptosis) / Blotting, Western / Caspase 8 / Caspase 9 / Caspases / 細胞死 (cell death) / Cell-Free System / Cells, Cultured / Cytochrome c Group / Cytosol / Enzyme Activation / 過酸化水素水 (hydrogen peroxide) / In Situ Nick-End Labeling / JNK Mitogen-Activated Protein Kinases / MAP Kinase Signaling System / Microscopy, Confocal / Microscopy, Electron / ミトコンドリア (mitochondria) / Mitogen-Activated Protein Kinase 8 / Mitogen-Activated Protein Kinases / 心筋 (myocardium) / 酸化ストレス (oxidative stress) / リン酸化 (phosphorylation) / Protein Binding / Rats / Recombinant Proteins / Subcellular Fractions / Time Factors
Takafumi Noma, Koichi Fujisawa, Yasuhiro Yamashiro, Miho Shinohara, Atsushi Nakazawa, Toshikazu Gondo, Tokuhiro Ishihara and Kumiko Yoshinobu : Structure and expression of human mitochondrial adenylate kinase targeted to the mitochondrial matrix, The Biochemical Journal, Vol.358, No.1, 225-232, 2001.
(要約)
The previously isolated cDNA encoding human adenylate kinase (AK) isozyme 3 was recently renamed AK4. Consequently, human AK3 cDNA remains to be identified and we have little information about the functional relationship between human AK3 and AK4. In pursuit of the physiological roles of both the AK3 and AK4 proteins, we first isolated an authentic human AK3 cDNA and compared their expression. Nucleotide sequencing revealed that the cDNA encoded a 227-amino-acid protein, with a deduced molecular mass of 25.6 kDa, that shares greater homology with the AK3 cDNAs isolated from bovine and rat than that from human. We named the isolated cDNA AK3. Northern-blot analysis revealed that AK3 mRNA was present in all tissues examined, and was highly expressed in heart, skeletal muscle and liver, moderately expressed in pancreas and kidney, and weakly expressed in placenta, brain and lung. On the other hand, we found that human AK4 mRNA was highly expressed in kidney, moderately expressed in heart and liver and weakly expressed in brain. Western-blot analysis demonstrated expression profiles of AK3 and AK4 that were similar to their mRNA expression patterns in each tissue. Over expression of AK3, but not AK4, in both Escherichia coli CV2, a temperature-sensitive AK mutant, and a human embryonic kidney-derived cell line, HEK-293, not only produced significant GTP:AMP phosphotransferase (AK3) activity, but also complemented the CV2 cells at 42 degrees C. Subcellular and submitochondrial fractionation analysis demonstrated that both AK3 and AK4 are localized in the mitochondrial matrix.
Akimasa Yamashita, Takafumi Noma, Atsushi Nakazawa, Satoshi Saito, Kentaro Fujioka, Nobuya Zempo and Kensuke Esato : Enhanced Expression of Matrix Metalloproteinase-9 in Abdominal Aortic Aneurysms, World Journal of Surgery, Vol.25, No.3, 259-265, 2001.
(要約)
Abdominal aortic aneurysms (AAAs) are characterized by structural alterations of the aortic wall resulting from degradation of collagen and elastin. Matrix metalloproteinases (MMPs), particularly MMP-2 and MMP-9, show strong elastinolytic activity. We examined the levels of mRNA for MMP-2, MMP-9, membrane type (MT)-MMP-1, tissue inhibitor of metalloproteinase-1 (TIMP-1), and TIMP-2 in AAAs (n = 8), atherosclerotic occlusive diseases (AOD) (n = 8), and normal subjects (n = 8) using the reverse transcription-polymerase chain reaction (RT-PCR). We also analyzed the gelatinolytic activity of these metalloproteinases using gelatin zymography. The levels of MMP-2 and MMP-9 mRNA were increased in the AAA group compared with those in the AOD group and normal subjects. The levels for TIMP-1 and TIMP-2 mRNA in the AAA group were also higher than those in the AOD and normal groups. Only in the case of MT-MMP-1 was the difference between AAA and AOD not statistically significant. By gelatin zymography with the same samples used for RT-PCR, gelatinolytic activity of MMP-9 was elevated in all AAA tissues. The 62-kDa form of MMP-2 was elevated in both the AAA and AOD groups and did not differ significantly between them. Linear regression analysis demonstrated a significant positive correlation between mRNA levels of MMPs and those of TIMPs. These observations suggest that aneurysm formation in patients with atherosclerosis is related to the degree of MMP-9 expression.
Takahiko Hara, Takafumi Noma, Yasuhiro Yamashiro, Katsusuke Naito and Atushi Nakazawa : Quantitative analysis of telomerase activity and telomerase reverse transcriptase expression in renal cell carcinoma, Urological Research, Vol.29, No.1, 1-6, 2001.
(要約)
To investigate the relationship between the telomerase activity levels and clinicopathological features of tumors, we quantified the telomerase activities of 23 renal cell carcinomas (RCCs) and four non-cancerous tissues, using a modified telomeric repeat amplification protocol assay, and assessed the hTERT mRNA levels of these samples by reverse transcription-polymerase chain reaction analysis. Elevated levels of telomerase activity had correlation with tumor stages as well as the degree of nuclear grades. Our findings suggested that telomerase activity is a useful indicator for tumor aggressiveness in RCCs. However, hTERT mRNA levels in RCCs had no correlation with nuclear grades and tumor stages. The telomerase activities and the hTERT mRNA levels in cancer cells were not always in parallel. These results suggested that telomerase activity is regulated in a posttranscriptional manner as well as a post-translational manner in tumor cells.
Kiyoshi Yoshimura, Shoichi Hazama, Norio Iizuka, Shigefumi Yoshino, Koutaro Yamamoto, Masahiro Muraguchi, Yasuichi Ohmoto, Takafumi Noma and Masaaki Oka : Successful immunogene therapy using colon cancer cells (colon 26) transfected with plasmid vector containing mature interleukin-18 cDNA and the Ig leader sequence, Cancer Gene Therapy, Vol.8, No.1, 9-16, 2001.
(要約)
IL-18 is a novel cytokine that induces interferon (IFN)-gamma secretion and plays an important role in antitumor immunity. In the present study, we constructed plasmid vectors encoding the murine mature IL-18 cDNA linked with the Igkappa leader sequence and the pro-IL-18 cDNA to estimate the efficacy of the mature IL- 18 vector and to evaluate IL-18--producing tumor cells as a tumor vaccine. Colon 26 cells were transfected with the abovementioned vectors or with vector alone (mock). Reverse transcription-polymerase chain reaction analysis showed increased expression of murine IL-18 cDNA in both mature IL-18 and pro-IL-18 transfectants in comparison to that in mock transfected cells. The ability of the culture supernatants of mature IL-18 transfectants to induce IFN-gamma secretion was extremely high (40-140 pg/10(6) cells) in comparison to that of pro-IL-18 transfectants (4-18 pg/10(6) cells). When injected into syngeneic BALB/c mice, the growth of mature IL-18 transfectants, but not pro-IL-18 transfectants, was significantly less than that in mock transfected cells ( P< .01, by ANOVA and analysis of covariance). In addition, injection of colon 26 or Meth-A cells into mice immunized with a mature IL-18 transfectant revealed acquired immunity. Depletion of natural killer cells did not affect the growth of transfectants. However, the growth inhibitory effects were partially abrogated following treatment with anti-CD4+ and anti-CD8+ antibodies. These data suggest that the rejection of mature IL-18/colon 26 cells was mediated through T-cell activation. Gene therapy using mature IL-18 transfectants containing a plasmid vector and the Igkappa leader sequence may be a useful tumor vaccine.
N Mori, M Oka, S Hazama, N Iizuka, K Yamamoto, S Yoshino, Akira Tangoku, Takafumi Noma and K Hirose : Detection of telomerase activity in peritoneal lavage fluid from patients with gastric cancer using immunomagnetic beads, British Journal of Cancer, Vol.83, No.8, 1026-1032, 2000.
(要約)
Cytologic examination of peritoneal lavage fluid is a useful predictor of peritoneal recurrence in gastric cancer. However, this technique is not overly sensitive and requires special abilities in the cytologist. In this study, telomerase activity was used to detect free cancer cells in peritoneal lavage fluid from patients with gastric cancer. In the first part, 12 lavage-fluid samples obtained from 12 patients with gastric cancer were analysed using the conventional telomeric repeat amplification protocol (TRAP) assay. Three of five patients with early gastric cancer had positive telomerase activity. These false-positive results may have been due to lymphocyte contamination. Furthermore, polymerase chain reaction inhibitors were also detected in the lavage-fluid samples. Therefore, we developed a novel method for elimination of haematopoietic cell and Taq polymerase inhibitors to increase the accuracy of the TRAP assay using immunomagnetic beads, which bind to most normal and neoplastic human epithelial cells. Telomerase activity was found in 10 of 20 (50%) lavage-fluid samples from patients with serosal or subserosal invasion. Cytologic examination was positive in nine of 20 (45%) samples. Both the telomerase activity and cytology were negative in all 14 patients without serosal or subserosal invasion. These results suggest that the TRAP assay combined with immunomagnetic beads might be useful for detection of free cancer cells in the peritoneal space in gastric cancer without the aid of an experienced cytologist.
Takafumi Noma, Ryutaro Murakami, Ysuhiro Yamashiro, Koichi Fujisawa, Sachie Inouye and Atsushi Nakazawa : cDNA cloning and chromosomal mapping of the gene encoding adenylate kinase 2 from Drosophila melanogaster, Biochimica et Biophysica Acta (BBA) - Gene Structure and Expression, Vol.1490, No.1-2, 109-114, 2000.
(要約)
As a step toward understanding of the role of adenylate kinase (AK) in energy metabolism, we analyzed this enzyme in Drosophila melanogaster. The enzyme activities of all three AK isozymes were determined in cell-free extracts of flies, and their proteins were detected by Western blot analysis using polyclonal antibodies against the mammalian isozymes. A cDNA encoding adenylate kinase was isolated from D. melanogaster cDNA library. The cDNA encodes a 240-amino acid protein, which shows high similarity to bovine, human and rat AK2, and hence was named DAK2. Preliminary subcellular fractionation analysis indicated that DAK2 is localized in both cytoplasm and mitochondria. In situ hybridization to salivary gland polytene chromosomes revealed that the Dak2 gene is located at 60B on the right arm of the second chromosome.
Takafumi Noma, Naoto Adachi, Haruhide Ito and Atsushi Nakazawa : Characterization of the 5'-flanking region of the gene encoding bovine adenylate kinase isozyme 3, Biochimica et Biophysica Acta (BBA) - Gene Structure and Expression, Vol.1489, No.2-3, 383-388, 1999.
(要約)
We have characterized the 5'-flanking region of the gene encoding bovine adenylate kinase isozyme 3 (AK3). S1 mapping analysis revealed multiple transcription start points in the bovine AK3 gene. The promoter activities were tested in HeLa cells using the chloramphenicol acetyltransferase (CAT) gene as a reporter. The CAT analysis showed that the basal promoter sequence was located within the region from -189 to +228. In the presence of short DNA fragments of the 5'-flanking region as competitors, the transcriptional activity of the bovine AK3 promoter changed depending on the fragments used. The results identified the basal regulatory elements in the proximal promoter region.
Takafumi Noma, Naoto Adachi and Atsushi Nakazawa : Cloning and Functional Characterization of the Promoter Region of the Gene Encoding Human Adenylate Kinase Isozyme 3, Biochemical and Biophysical Research Communications, Vol.264, No.3, 990-997, 1999.
(要約)
The 5'-flanking region of the gene encoding human adenylate kinase isozyme 3 was isolated and compared with that of the bovine AK3 gene previously characterized. Four conserved DNA sequences (elements-a, -b, -c, and -d) were found in both the regions. The promoter activities were analyzed in HeLa cells using promoter-CAT reporter constructs. The proximal promoter region (-217 to +261), which contains three of four conserved elements, gave a maximum promoter activity. In a series of electrophoretic mobility-shift assays, DNA fragments and double-stranded oligodeoxyribonucleotides containing sequences of the four conserved elements interacted with nuclear extracts of HeLa cells. The a-element contained the W-element, while the d-element, which had a high G + C content, was a novel regulatory cis-element distinct from the GC box. The b- and c-elements were homologous to each other and had a motif resembling downstream promoter element. Mutations of the c- and d-elements significantly reduced the promoter activity, indicating that the c- and d-elements in the AK3 promoter are crucial. These elements may also be involved in the transcriptional regulation of other TATA-less genes.
N Iizuka, K Hirose, Takafumi Noma, S Hazama, Akira Tangoku, H Hayashi, T Abe, K Yamamoto and M Oka : The nm23-H1 gene as a predictor of sensitivity to chemotherapeutic agents in oesophageal squamous cell carcinoma, British Journal of Cancer, Vol.81, No.3, 469-475, 1999.
(要約)
Recently, nm23-H1, an anti-metastasis gene, has been reported to correlate with sensitivity to chemotherapeutic agents including cisplatin in human breast and ovarian carcinoma cells. The aim of this study was to evaluate a role for nm23-H1 in responsiveness to cisplatin-based chemotherapy in patients with oesophageal squamous cell carcinoma (OSCC). The expression of nm23-H1 protein was examined immunohistochemically in 32 eligible patients with OSCC who underwent adjuvant chemotherapy with cisplatin, etoposide, and 5-fluorouracil after tumour resection. Fifteen (46.9%) of 32 patients were positive for nm23-H1 staining and 17 (53.1%) were negative. Both disease-free survival and overall survival rates of nm23-H1-negative patients were significantly shorter than in nm23-H1-positive patients (P < 0.01 for both). There was no significant difference in clinicopathologic characteristics between nm23-H1-positive and nm23-H1-negative groups. Multivariate analysis also showed that nm23-H1 expression was the most significant factor for overall survival of OSCC patients included in this study (P = 0.0007). To further study the role of nm23-H1, a human OSCC cell line (YES-2) was transfected with a plasmid containing a fragment of the nm23-H1 cDNA in an antisense orientation. Reduced expression of nm23-H1 protein in the antisense-transfected (AS) clones was found by Western blot analysis as compared to wild-type YES-2 and YES-2/Neo (clone transfected with the neomycin resistance gene alone). MTT (3-(4,5-dimethyl-2-thiazol)-2,5-diphenyl-2H tetrazolium bromide) assay showed that reduced expression of the nm23-H1 protein in AS clones was consistent with the degree of increased resistance to cisplatin but not etoposide or 5-fluorouracil. These data support the conclusion that reduced expression of nm23-H1 may be associated with resistance to cisplatin, suggesting the value of nm23-H1 expression as a prognostic marker for OSCC patients who are to undergo cisplatin-based chemotherapy.
Takafumi Noma, Yong-Shik Yoon, Yasuhiro Yamashiro, Koichi Fujisawa and Atsushi Nakazawa : Regulation of NeuroD expression by activation of the protein kinase-C pathway in Y79 human retinoblastoma cells, Neuroscience Letters, Vol.272, No.1, 45-48, 1999.
(要約)
To examine the signals that regulate NeuroD expression, we analyzed the effects of activation of two major signal pathways, the protein kinase A (PKA) pathway and the protein kinase C (PKC) pathway, on the expression of NeuroD in Y79human retinoblastoma cells. Activation of PKC resulted in marked induction of NeuroD mRNA and NeuroD protein. NeuroD mRNA induction was inhibited by calphostin C, an inhibitor of PKC. On the other hand, stimulation of PKA by forskolin had a weak suppressive effect on NeuroD mRNA expression. Induction of NeuroD expression was followed by enhancement of expression of the AK1 gene, one of the target genes of NeuroD, which encodes adenylate kinase isozyme 1, an important enzyme in the cellular adenine nucleotide homeostasis. Our results indicate that NeuroD expression is regulated, at least in part, by the PKC pathway and not by the PKA pathway.
(キーワード)
Adenylate kinase isozyme 1 / Basic helix-loop-helix / NeuroD / Protein kinase A / Protein kinase C / Reverse transcription-polymerase chain reaction
S Hazama, Takafumi Noma, F Wang, N Iizuka, Y Ogura, K Yoshimura, E Inoguchi, M Hakozaki, K Hirose, T Suzuki and M Oka : Tumour cells engineered to secrete interleukin-15 augment anti-tumour immune responses in vivo, British Journal of Cancer, Vol.80, No.9, 1420-1426, 1999.
(要約)
We examined the effect of interleukin-15 (IL-15) gene transfer into tumour cells on the host's anti-tumour response. In BALB/c mice IL-15 producing Meth-A cells (Meth-A/IL-15) underwent complete rejection, in a response characterized by massive infiltration of CD4+ T-cells and neutrophils. In contrast, Meth-A cells transfected with vector alone (Meth-A/Neo) grew rapidly. Moreover, rechallenged parental cells also were rejected in association with CD8* T-cell infiltration. However, in nude mice there was no drastic difference between Meth-A/IL-15 and Meth-A/Neo cells. These results demonstrate that IL-15-secreting tumour cells can stimulate local and systemic T-cell-dependent immunity and therefore may have a potential role in cancer therapy.
(キーワード)
IL-15 / tumour immunity / vaccination / in vivo animal models / 遺伝子治療 (gene therapy)
Takafumi Noma, Yong-Shik Yoon and Atsushi Nakazawa : Overexpression of NeuroD in PC12 cells alters morphology and enhances expression of the adenylate kinase isozyme 1 gene, Brain Research. Molecular Brain Research, Vol.67, No.1, 53-63, 1999.
(要約)
NeuroD, a basic helix-loop-helix transcription factor, plays an important role in neuronal differentiation. A rat NeuroD cDNA was obtained by the aid of reverse transcription-polymerase chain reaction (RT-PCR) and ligated to an expression vector having a CMV promoter. Transfection of the NeuroD-expression plasmid into PC12 cells, a rat pheochromocytoma cell line, induced morphological changes featured by neurite-like processes and synapse-like structures without a differentiation-inducing reagent such as NGF. In the transfected cells, the overproduced NeuroD was detected by Western blot analysis, and the expression of the gene encoding mid-sized neurofilaments, a neuron-specific marker, was demonstrated by RT-PCR. Adenylate kinase isozyme 1 (AK1) is an enzyme involved in the homeostasis of energy metabolism and appears specifically in neuronal cells during differentiation. The CAT reporter assay of the 5'-flanking region of the AK1 gene suggests that NeuroD activates the AK1 expression through E-boxes in the promoter region. RT-PCR analysis indicated the enhanced level of AK1 mRNA in NeuroD-producing PC12 cells. Electrophoretic mobility shift assays demonstrated that NeuroD was able to interact with a proximal E-box element of the AK1 promoter. The results indicated that NeuroD promoted the PC12 cells to differentiate into neuron-like cells with concomitant activation of the target genes including the AK1 and the neurofilament genes.
Camilla Köhler, Annie Gahm, Takafumi Noma, Atsushi Nakazawa, Sten Orrenius and Boris Zhivotovsky : Release of adenylate kinase 2 from the mitochondrial intermembrane space during apoptosis, FEBS Letters, Vol.447, No.1, 10-12, 1999.
(要約)
The release of two mitochondrial proteins, cytochrome c and apoptosis-inducing factor (AIF), into the soluble cytoplasm of cells undergoing apoptosis is well established. Using spectrophotometric determination of enzyme activity, the accumulation of adenylate kinase (AK) activity in the cytosolic fraction of apoptotic cells has also been observed recently. However, three isozymes, AK1, AK2 and AK3, have been characterized in mammalian cells and shown to be localized in the cytosol, mitochondrial intermembrane space and mitochondrial matrix, respectively, and it is unknown which one of these isozymes accumulates in the cytosol during apoptosis. We now demonstrate that in apoptotic cells only AK2 was translocated into the cytosol concomitantly with cytochrome c. The amount of AK1 in cytosol, as well as the amount of matrix-associated AK3, remained unchanged during the apoptotic process. Thus, our data suggest that only intermembrane proteins are released from mitochondria during the early phase of the apoptotic process.
Yong-Shik Yoon, Takafumi Noma, Yasuhiro Yamashiro, Haruhide Ito and Atsushi Nakazawa : Molecular cloning and characterization of the gene encoding human NeuroD, Neuroscience Letters, Vol.251, No.1, 17-20, 1998.
(要約)
NeuroD is a basic helix-loop-helix transcription factor that binds to an E-box sequence, CANNTG, in the target gene promoter region. The gene encoding NeuroD is expressed specifically in brain, intestine, and pancreatic cells and plays a pivotal role in tissue-specific differentiation. To investigate the regulation of human NeuroD gene expression, we isolated and sequenced a genomic DNA containing two exons and flanking regions of the NeuroD gene. The sizes of exons 1 and 2 were 168 and 2404 bp, respectively. CAT reporter analysis showed that the elements responsible for basal promoter activity lay in the proximal 408-bp region. Cotransfection of the CAT reporter plasmids with a NeuroD expression plasmid resulted in 5-fold enhancement of the CAT activity, indicating an autoregulatory mechanism is involved in human NeuroD gene expression.
Takafumi Noma, Shaochuen Song, Yong-Shik Yoon, Shinichi Tanaka and Atsushi Nakazawa : cDNA cloning and tissue-specific expression of the gene encoding human adenylate kinase isozyme 2, Biochimica et Biophysica Acta (BBA) - Gene Structure and Expression, Vol.1395, No.1, 34-39, 1998.
(要約)
We isolated two kinds of cDNAs encoding human adenylate kinase (AK) isozyme 2 from a HeLa cell cDNA library using bovine AK2 cDNA as a probe. Nucleotide sequencing revealed that the cDNAs encoded 239- and 232-amino acid proteins with deduced molecular mass of 26.5 (AK2A) and 25.6kDa (AK2B), respectively. Northern blot analysis demonstrated that AK2 mRNA is strongly expressed in liver, heart, skeletal muscle and pancreas, and moderately in kidney, placenta and brain, and weakly in lung. However, Western blot analysis showed that AK2 protein was present in large amounts in liver, heart, kidney, and in a small amount in lung, and undetectable in brain and skeletal muscle. These results suggested the presence of the tissue-specific gene-expression including post-transcriptional regulation in expression of the AK2 gene.
(キーワード)
adenylate kinase isozyme 2 / cDNA cloning / Nucleotide sequencing / Northern blot / Western blot / tissue distribution
Shuji Terai, Takafumi Noma, Teruaki Kimura, Atsushi Nakazawa, Fumie Kurokawa and Kiwamu Okita : Wild-type p53 gene-induced morphological changes and growth suppression in hepatoma cells, Journal of Gastroenterology, Vol.32, No.3, 330-337, 1997.
(要約)
The human hepatocellular carcinoma (HCC) cell line, HLF, expresses only mutant-type p53 (mt-p53), which has an amino acid substitution at the 244th residue from glycine to alanine. HLF cells were transfected with wild-type p53 (wt-p53) cDNA construct pC53-SN3, mt-p53 cDNA construct pC53-SCX [which differs by a single nucleotide, resulting in alanine instead of valine at the 143rd residue in p53 (p53-143)], or pCMV-Neo-Bam, as a control, by a liposome method. After G418 selection, three wt-p53 stable transformants (WT), four mt-p53 transformants (MT), and three control vector transformants (VT) were obtained. We analyzed the cell growth and morphological changes of these transformants under different culture conditions [fetal calf serum (FCS), 10%, 1%, and 0%]. Whereas no difference from control in the growth rate and morphology was observed under the 10% FCS conditions, serum starvation induced remarkable phenotypical changes in all three WTs, but not in the other transformant. Corresponding to these phenotypical changes, the transcriptional activity of wt-p53 was increased more than nine fold. These results indicated that serum starvation would induce wt-p53 biological function, which is tightly linked to morphological changes and growth suppression. To induce these changes, the introduction of the wt-p53 gene itself was not sufficient, and additional triggering, i.e., serum starvation, was indispensable.
Tomoko Ohksa, Takafumi Noma, Takeshi Ueyama, Yuji Hisamatsu, Masafumi Yano, Kensuke Esato, Atsushi Nakazawa and Masunori Matsuzaki : Differences in sarcoplasmic reticulum gene expression in myocardium from patients undergoing cardiac surgery. Quantification of steady-state levels of messenger RNA using the reverse transcription-polymerase chain reaction, Heart and Vessels, Vol.12, No.1, 1-9, 1997.
(要約)
Little is known about any alterations in sarcoplasmic reticulum (SR) gene expression associated with cardiac diseases of varying degrees of severity. We assessed, using the reverse transcription-polymerase chain reaction (RT-PCR) technique, SR Ca2+ transport protein gene expression in small tissue samples from failing hearts in patients undergoing cardiac surgery. Total RNA was extracted from 30- to 50-mg samples from the hearts of 13 patients with coronary artery disease, congenital heart disease, or valvular heart disease. We used RT-PCR to synthesize and amplify cDNA encoding cardiac SR Ca(2+)-ATPase, ryanodine receptor (RYR), and glyceraldehyde-3-phosphate dehydrogenase (GAPDH). The amount of each mRNA in the sample was expressed relative to the amount of GAPDH mRNA. The expression level of each mRNA was correlated with the cardiac functional index. The mRNA levels for Ca(2+)-ATPase and RYR varied between heart samples, but showed a positive correlation with left ventricular ejection fraction. Ca(2+)-ATPase mRNA levels showed in inverse relationship with plasma brain natriuretic peptide. In addition, we isolated partial cDNA encoding a human cardiac RYR. The cDNA consisted of 487 nucleotides, and the nucleotide and deduced amino acid sequences showed 93% and 99% homology, respectively, to those of rabbit cardiac RYR. These results suggest that decreased levels of mRNA for SR Ca2+ transport protein could be related to abnormal cardiac function, regardless of the etiology of the heart disease. RT-PCR provides a rapid and economical way of quantifying the expression of multiple genes in small specimens and may, therefore, aid understanding of the pathophysiology and treatment of heart disease.
Hiroyoshi Ogasa, Takafumi Noma, Hidenori Murata, Shinya Kawai and Atsushi Nakazawa : Cloning of a cDNA encoding the human transforming growth factor-beta type II receptor: heterogeneity of the mRNA, Gene, Vol.181, No.1-2, 185-190, 1996.
(要約)
We have isolated two novel cDNAs encoding the transforming growth factor-beta (TGF-beta) type II receptor (TGF-beta IIR), termed TGF-beta IIR alpha and TGF-beta IIR beta 1 from a human fetal liver library. They have unique nucleotide (nt) sequences, compared with the reported TGF-beta IIR sequence, at the 5' end. Southern blot analysis using probes from each clone detected the specific genomic DNA fragments. RT-PCR analysis revealed a distinct pattern of expression for each isoform. These results indicated that TGF-beta IIR has heterogeneity in the structure, and the expression of TGF-beta IIR isoforms is differentially regulated. The heterogeneity of TGF-beta IIR molecules could be derived from alternative splicing and might elicit specific TGF-beta receptor functions.
(キーワード)
nucleotide sequence / isoform / Southern blot analysis / alternative splicing / Rt-pcr
Norio Iizuka, Masaaki Oka, Takafumi Noma, Atsushi Nakazawa, Kunitaka Hirose and Takashi Suzuki : NM23-H1 and NM23-H2 messenger RNA abundance in human hepatocellular carcinoma, Cancer Research, Vol.55, No.3, 652-657, 1995.
53.
Toshifumi Tomoda, Toshiyuki Kudoh, Takafumi Noma, Atsushi Nakazawa, Masa-aki Muramatsu and Ken-ichi Arai : Molecular cloning of a mouse counterpart for human TGF-beta type I receptor, Biochemical and Biophysical Research Communications, Vol.198, No.3, 1054-1062, 1994.
54.
Tsuyoshi Tanabe, Mamoru Yamada, Takafumi Noma, Tadashi Kajii and Atsushi Nakazawa : Tissue-specific and developmentally regulated expression of the genes encoding adenylate kinase isozymes, The Journal of Biochemistry, Vol.113, No.2, 200-207, 1993.
(要約)
Adenylate kinase (AK) is known to play an important role in homeostasis of adenine nucleotide metabolism. We isolated cDNAs for rat AK isozymes (AK1, AK2, and AK3), determined their mRNAs in rat tissues by Northern blot analysis, and measured the isozyme activities. Tissue-dependent activities of AK1 and AK2 paralleled the contents of mRNAs. Tissues with high AK1 levels showed low AK2 levels and vice versa, suggesting that tissue-specific expressions of the AK1 and AK2 genes are inversely regulated. AK3 mRNA was detected in most tissues examined, suggesting that AK3 gene expression is constitutive. We further examined developmental changes in mRNAs and enzyme activities of AK isozymes in rat skeletal muscle and liver. In the skeletal muscle, AK1 and AK3 activities started to increase at around the weaning period. AK1 mRNA accumulated at the prenatal stage and further increased during development, while AK3 mRNA was at high levels during the fetal stage and remained fairly constant during development. In the liver, AK2 and AK3 activities started to increase after birth and were further elevated during growth, whereas their mRNAs were present at relatively high levels throughout development. The physiological meanings of the tissue-specific expression of the AK isozyme genes are discussed.
(キーワード)
Adenylate Kinase / Amino Acid Sequence / Animals / Base Sequence / DNA (DNA) / Embryonic and Fetal Development / 女性 (female) / Gene Expression Regulation, Enzymologic / Isoenzymes / 男性 (male) / Molecular Sequence Data / RNA, Messenger / Rats / Rats, Wistar / Sequence Homology, Amino Acid / Tissue Distribution
Takafumi Noma, B Adam Glick, Geiser G Andrew, O`reilly A Michael, Miller Jeanne, Roberts B Anita and Sporn B Michael : Molecular cloning and structure of the human transforming growth factor-beta 2 gene promoter, Growth factors, Vol.4, No.4, 247-255, 1991.
(要約)
Genomic DNA extending over 10 kb 5' of the transforming growth factor-beta 2 (TGF-beta 2) coding region was isolated from a human lung fibroblast lambda phage library. A 5.6 kb Hind III fragment containing the 5'-untranslated region and flanking sequences was subcloned and sequenced. S1 nuclease protection analysis identified a transcriptional initiation site 1357 nucleotides 5' of the methionine initiation codon (ATG). A "TATA box" consensus sequence was identified 30 bp from this transcriptional start site; however, consensus "CAT box" sequences were not observed. Approximately 50 nucleotides of homopurine-pyrimidine [d(GA.CT)50] sequence were identified in the 5'-untranslated region, as well as two short open reading frames of 5 and 45 amino acids. Several AP-1, AP-2, CRE and SP1-like DNA consensus sequence elements were also identified surrounding the transcription initiation site. 5'-deletion mutants of the promoter region were fused to the chloramphenicol acetyl transferase (CAT) gene and promoter activity of the isolated genomic DNA was demonstrated in several cell lines. DNA constructs containing nucleotides between -508 to +63 demonstrated high levels of promoter activity. However, sequences between -778 and -508 nucleotides modulated this promoter activity in a manner which was dependent upon the cell line utilized, suggesting that regulation of TGF-beta 2 gene transcription may be dependent upon the cellular background. The TGF-beta 2 promoter is markedly different from the promoters that have been recently characterized for TGF-beta 1 and TGF-beta 3.
Toshihiro Tanaka, Kenzo Takahashi, Shin Ideyama, Sadao Imamura and Takafumi Noma : Demonstration of clonal proliferation of T lymphocytes in early neoplastic disease. Studies with probes for the beta-chain of the T cell receptor and human T cell lymphotropic virus type I, Journal of the American Academy of Dermatology, Vol.21, No.2 Pt 1, 218-223, 1989.
(要約)
This study demonstrates that the detection of rearrangement of the T-cell receptor gene and adult T cell leukemia virus or human T cell lymphotropic virus type I (ATLV/HTLV-I) integration in the genome are sensitive and practical methods for the diagnosis and characterization of cutaneous T cell neoplasms. Biopsy specimens obtained from small skin nodules containing a dense infiltration of lymphocytes were analyzed by Southern blotting with the use of probes for both the beta-chain of the T-cell receptor gene and the ATLV/HTLV-I genome. DNA samples from one patient with early, chronic adult T cell lymphoma or leukemia, revealed the monoclonal proliferation of lymphocytes. For another patient with early cutaneous T cell lymphoma analysis by Southern blotting with the beta-chain probe of DNA from three separate lesions revealed identical rearranged bands, indicating not only the monoclonal proliferation of T cells in each lesion but also the clonal origin of each lesion. In contrast, analysis by Southern blotting of DNA samples extracted from three nodules from a patient with lymphomatoid papulosis showed no rearranged band and therefore no clonal proliferation. DNA extracted from blood lymphocytes of these patients failed to hybridize with the probes described above as a rearranged band. These results indicate that analysis of DNA extracted from skin lesions may have diagnostic value in determining monoclonal proliferation of lymphocytes in patients with early stage cutaneous lymphomas.
Takafumi Noma, Hiroshi Nakakubo, Kazuaki Hama and Tasuku Honjo : Multiple effects of human recombinant interleukin 4 on human myeloid monocyte cell lines, Immunology Letters, Vol.21, No.4, 323-328, 1989.
(要約)
The effects of recombinant human interleukin 4 (rIL-4) on proliferation and differentiation on human myeloid/monocytic leukemia cell lines were examined. At high concentrations, rIL-4 had a slight enhancing effect on [3H]thymidine incorporation by U937 cells. rIL-4 markedly induced expression of the Fc epsilon receptor (CD23) and the Leu-M3 antigen (CD14) on U937 cells. HL60 and THP-1 cells treated with rIL-4 also showed increased CD23 expression, but little change of CD14 antigen expression. CD23 induction required lower amounts of IL-4 than needed for T cell growth, indicating that CD23 induction on U937 will serve as a sensitive assay for human IL-4. rIL-4 reduced the steady state level of IL-1 beta mRNA in U937.
Hideo Enokihara, Shimpei Furusawa, Hiroshi Nakakubo, Hiroshi Kajitani, Shigeki Nagashima, Kenji Saito, Hideo Shishido, Yasumichi Hitoshi, Kiyoshi Takatsu, Takafumi Noma, Akira Shimizu and Tasuku Honjo : T cells from eosinophilic patients produce interleukin-5 with interleukin-2 stimulation, Blood, Vol.73, No.7, 1809-1813, 1989.
(要約)
Anti-murine (m) interleukin-5 (IL-5) antibody was found to inhibit eosinophil (Eo) colony formation stimulated by recombinant human (rh) IL-5, but did not inhibit the production of Eo stimulated by rh IL-3 or granulocyte-macrophage colony-stimulating factor (GM-CSF). Conditioned medium (CM) prepared from eosinophilic patients' T cells with interleukin-2 (IL-2) stimulation (T-IL-2-CM), was found to contain CFU-Eo growth-stimulating factor. Using anti-mIL-5 antibody, we demonstrated that T-IL-2-CM from patients with eosinophilia contained a significant amount of IL-5. We also detected IL-5 mRNA in T cells from eosinophilic patients with IL-2 stimulation. These results suggest that IL-5 plays an important role in the induction of selective eosinophilia in humans and that IL-5 is produced from T cells with IL-2 stimulation.
Takafumi Noma, Hiroshi Nakakubo, Masahiko Sugita, Shunichi Kumagai, Michiyuki Maeda, Akira Shimizu and Tasuku Honjo : Expression of different combinations of interleukins by human T cell leukemic cell lines that are clonally related, The Journal of Experimental Medicine, Vol.169, No.5, 1853-1858, 1989.
Michiyuki Maeda, Takafumi Noma, Kazuaki Hama and Tasuku Honjo : Application of a human T cell line derived from a Sezary syndrome patient for human interleukin 4 assay, Immunology Letters, Vol.18, No.4, 247-253, 1988.
(キーワード)
Interleukin 4 / B cell stimulatory factor 1 / assay system for human IL-4 / 人間 (human)
62.
Hideo Enokihara, Shigeki Nagasima, Takafumi Noma, Hiroshi Kajitani, Hiroyuki Hamaguchi, Kenji Saito, Shimpei Furusawa, Hideo Shishido and Tasuku Honjo : Effect of human recombinant interleukin 5 and G-CSF on eosinophil colony formation, Immunology Letters, Vol.18, No.1, 73-76, 1988.
(要約)
Human recombinant (r) IL-5 was shown to have the activity to stimulate eosinophil (Eo) colony formation from human non-T, non-adherent bone marrow cells. The majority of these colonies were found to contain a small number of basophils, macrophages or neutrophils. Human rG-CSF, which alone did not stimulate Eo colony formation, showed an enhancing effect on Eo colony formation when added with IL-5. IL-5 seems to stimulate the proliferation and differentiation of CFU-Eo, while G-CSF acts on the early stage of eosinophilopoiesis.
Kenji Nakanishi, Tomohiro Yoshimoto, Yoshiya Katoh, Shiro Ono, Kiyoshi Matsui, Keisai Hiroishi, Takafumi Noma, Tasuku Honjo, Kiyoshi Takatsu, Kazuya Higashino and Toshiyuki Hamaoka : Both B151-T cell replacing factor 1 and IL-5 regulate Ig secretion and IL-2 receptor expression on a cloned B lymphoma line, The Journal of Immunology, Vol.140, No.4, 1168-1174, 1988.
(要約)
In the present study, we have demonstrated that both B151-T cell-replacing factor 1 and rIL-5 are responsible for the activity to partially induce CL-3 cells into IgM-synthesizing cells and also to synergize with IL-2 to augment IL-2R expression on and IgM synthesis in CL-3 cells. These actions of rIL-5 on a homogeneous cloned line (BCL1-CL-3 cells) allow us to identify and characterize the two alternated B cell developmental pathways. One is an IL-2-independent, IL-5-driven differentiation pathway without preceding up-regulated IL-2R expression, and the other is an IL-5 plus IL-2-dependent augmented differentiation pathway with preceding up-regulated IL-2R expression. We have also demonstrated the functional difference of two distinct B cell growth-promoting factors, B cell-stimulating factor 1 (rIL-4) and rIL-5. CL-3 cells are equally stimulated to grow by rIL-4 and rIL-5, whereas only rIL-5 can render CL-3 cells responsive to rIL-2, indicating that these two lymphokines affect B cells in a strikingly different manner.
Tatsunobu-Ryushin Mizuta, Toshizumi Tanabe, Hiroshi Nakakubo, Takafumi Noma and Tasuku Honjo : Molecular cloning and structure of the mouse interleukin-5 gene, Growth factors, Vol.1, No.1, 51-57, 1988.
(キーワード)
Nucleotide sequence / conserved sequences in 5`-flanking region / transcription initiation site
65.
Toshizumi Tanabe, Mikio Konishi, Tatsunobu Mizuta, Takafumi Noma and Tasuku Honjo : Molecular cloning and structure of the human interleukin-5 gene, The Journal of Biological Chemistry, Vol.262, No.34, 16580-16584, 1987.
(要約)
We isolated the chromosomal gene for human interleukin-5 (IL-5) from human genomic libraries using a cloned human IL-5 cDNA as probe. Nucleotide sequence determination of the IL-5 gene and its flanking regions showed that the gene consisted of four exons and three introns. TATA-like and CAAT-like sequences reside 26 and 73 base pairs, respectively, upstream of the transcription initiation site identified by S1 mapping analysis. The 5'-flanking region of the IL-5 gene has sequences homologous with those of the corresponding regions of the genes for human granulocyte/macrophage colony-stimulating factor, murine IL-4, human interferon-gamma, and human IL-2. The intron-exon organization and location of the cysteine residue suggest that the IL-5 gene is phylogenetically related to the genes for the granulocyte/macrophage colony-stimulating factor, IL-4, and IL-2, although their amino acid sequences are not significantly conserved.
(キーワード)
Amino Acid Sequence / Base Sequence / Cloning, Molecular / Colony-Stimulating Factors / Cysteine / Endonucleases / Humans / Interferon-gamma / Interleukin-2 / Interleukin-5 / Interleukins / Introns / Molecular Sequence Data / RNA, Messenger / Single-Strand Specific DNA and RNA Endonucleases
(文献検索サイトへのリンク)
● PubMed @ National Institutes of Health, US National Library of Medicine (PMID): 2824500
Takafumi Noma, Tatsunobu Mizuta, Anders Rosen, Toshio Hirano, Tadamitsu Kishimoto and Tasuku Honjo : Enhancement of the interleukin 2 receptor expression on T cells by multiple B-lymphotropic lymphokines, Immunology Letters, Vol.15, No.3, 249-253, 1987.
(要約)
Three new human lymphokines, interleukin-5, BSF-2 and BSF-MP6, were shown to be active in the enhancement of the IL-2 receptor expression on T cells, although they do not stimulate growth of the T cells.
Chihiro Azuma, Toshizumi Tanabe, Mikio Konishi, Tatsuo Kinashi, Takafumi Noma, Fumihiko Matsuda, Yoshio Yaoita, Kiyoshi Takatsu, Lennart Hammarstrom, C.I.Edvard Smith, Eva Severinson and Tasuku Honjo : Cloning of cDNA for human T-cell replacing factor(interleukin-5)and comparison with the murine homologue, Nucleic Acids Research, Vol.14, No.22, 9149-9158, 1986.
(要約)
We have cloned cDNA for T-cell replacing factor (interleukin-5), which replaces T-cell helper function for normal B cells which secrete immunoglobulin, from human T cell leukemia line, ATL-2, using mouse interleukin-5 cDNA as probe. Total nucleotide sequence of the cDNA (816 base pairs) was determined and compared with that of mouse interleukin-5 cDNA. The cloned cDNA encoded the interleukin-5 precursor of 134 amino acids containing an N-terminal signal sequence. Although the human interleukin-5 precursor is one amino acid longer than the murine homologue, the sizes of the mature proteins appear similar. The nucleotide and amino acid sequence homologies of the coding regions of human and murine interleukin-5 are 77% and 70%, respectively. Human interleukin-5 synthesized by the direction of the cloned cDNA induced immunoglobulin synthesis in human B cells stimulated by Staphylococcus aureus mitogen.
Takafumi Noma, Tomoyuki Nakamura, Michiyuki Maeda, Masafumi Okada, Yoshihisa Taniguchi, Yutaka Tagaya, Yoshio Yaoita, Junji Yodoi and Tasuku Honjo : Interleukin 1 alpha mRNA in virus-transformed T and B cells, Biochemical and Biophysical Research Communications, Vol.139, No.1, 353-360, 1986.
(要約)
IL-1 alpha cDNA clone was isolated from a T cell line infected by the human T lymphotropic retrovirus type-I (HTLV-I/ATLV). We found significant amounts of mRNA hybridizing to IL-1 alpha cDNA not only in HTLV-I-transformed T cells but also in Epstein-Barr Virus-transformed B cells. A part of IL-2 receptor inducing activity in Adult T cell leukemia (ATL) cell line seems to be due to IL-1 alpha.
Mieko Kodaira, Kinashi Tatsuo, Umemura Isao, Matsuda Fumihiko, Takafumi Noma, Yasushi Ono and Tasuku Honjo : Organization and evolution of variable region genes of the human immunoglobulin heavy chain, Journal of Molecular Biology, Vol.190, No.4, 529-541, 1986.
(要約)
We have isolated 23 different cosmid clones of the heavy-chain variable region genes (VH) of human immunoglobulin. These clones encompass about 1000 X 10(3) base-pairs of DNA containing 61 VH genes. Characterization of the 23 clones by Southern blot hybridization showed that VH genes belonging to different families were physically linked in many regions. Cluster 71, which was analyzed in detail, comprised seven VH segments arranged in the same orientation with different intervals. This clone contained internal homology regions, each carrying two VH segments of different families. Comparison of the nucleotide sequences of VH segments within each family showed that profiles of accumulation of mutations in framework (FR) and complementarity-determining (CDR) regions were different. CDR had more mutations at amino-acid-substituting positions than at silent positions, whereas FR had the reverse distribution of mutations. Five out of seven VH segments of this cluster were pseudogenes containing various mutations. VH pseudogenes were classified into two distinct groups; one with a few replacement mutations (conserved pseudogenes), and the other with rather extensive mutations (diverged pseudogenes). The possibility that conserved pseudogenes serve as a reservoir of VH segments is discussed.
(キーワード)
Base Sequence / Biological Evolution / Chromosome Mapping / Cloning, Molecular / DNA / Genes / Humans / Immunoglobulin Heavy Chains / Multigene Family / Nucleic Acid Hybridization / Sequence Homology, Nucleic Acid
Shun-ichi Takeda, Takayuki Naito, Kazuaki Hama, Takafumi Noma and Tasuku Honjo : Construction of chimaeric processed immunoglobulin genes containing mouse variable and human constant region sequences, Nature, Vol.314, No.6010, 452-454, 1985.
(要約)
The specificity of monoclonal antibodies provides a powerful diagnostic and therapeutic tool in investigating human neoplasia. Radiological scanning and immunotherapy with mouse tumour-specific monoclonal antibodies have been applied to patients with some success, but a major problem is the neutralization of the mouse antibody induced by repeated administration of heterologous antibodies. To avoid or reduce such immune reactions, chimaeric immunoglobulins consisting of mouse variable (V) and human constant (C) regions can be synthesized. We have constructed a recombinant retrovirus DNA carrying genomic heavy-chain (H) variable-diversity joining (VH-D-JH) and C gamma 1 genes from different species and show here that the chimaeric intervening sequences are spliced out precisely. This procedure provides a useful method to construct the chimaeric mouse-human immunoglobulin gene to be expressed in Escherichia coli, yeast and animal cells. Unexpectedly, a hidden splice donor site in the 5'-flanking region of a human VH gene is used in place of the donor site of the leader sequence exon, resulting in the formation of the V region without the leader sequence.
(キーワード)
Animals / Base Sequence / Chimera / DNA, Recombinant / Genes / 遺伝子工学 (genetic engineering) / Humans / Immunoglobulin Constant Regions / Immunoglobulin Heavy Chains / Immunoglobulin Variable Region / Immunoglobulin kappa-Chains / Immunoglobulins / Mice / Plasmids
Junji Yodoi, Keisuke Teshigawara, Toshio Nikaido, Kiyoshi Fukui, Takafumi Noma, Tasuku Honjo, Masahiro Takigawa, Masao Sasaki, Nagahiro Minato, Mitsuru Tsudo, Takashi Uchiyama and Michiyuki Maeda : TCGF (IL-2)-Receptor Inducing Factor(s)., --- I. Regulation of IL 2 Receptor on a Natural Killer-like Cell Line (YT Cells). ---, The Journal of Immunology, Vol.134, No.3, 1623-1630, 1985.
73.
Hisataka Sabe, Shigeru Kondo, Akira Shimizu, Yutaka Tagaya, Junji Yodoi, Nobuyuki Kobayashi, Masakazu Hatanaka, Norisada Matsunami, Michiyuki Maeda, Takafumi Noma and Tasuku Honjo : Properties of human interleukin-2 receptors expressed on non-lymphoid cells by cDNA transfection, Molecular Biology & Medicine, Vol.2, No.6, 379-396, 1984.
(要約)
We have established non-lymphoid cell lines HeLa, Ltk and NIH3T3 expressing the human interleukin-2 (IL-2) receptor by transfection of human IL-2 receptor complementary DNA. While IL-2 receptors on T cells are classified into the high and low affinity species, the receptors expressed on the cDNA-transfected non-lymphoid cells belong to the low affinity species. These IL-2 receptors could not transmit the growth signal although they were similar in size to those expressed on T cells. Phorbol ester-induced phosphorylation of the IL-2 receptors on HeLa cells did not affect the affinity of the receptor. We have also constructed a cDNA encoding a mutant IL-2 receptor that replaced the major phosphorylation site, the serine residue at position 247 with the glycine residue. This mutant IL-2 receptor expressed on non-lymphoid cells also had the low affinity for IL-2. The results indicate that the high and low affinity states of the IL-2 receptor are not solely determined by phosphorylation of the receptor. The IL-2 receptors expressed on these non-lymphoid cells were internalized four to eight times more slowly than those on T cells. Possible defects of the IL-2 receptors expressed on non-lymphoid cells are discussed.
Naoki Takahashi, Takafumi Noma and Tasuku Honjo : Rearranged immunoglobulin heavy chain variable region (VH) pseudogene that deletes the second complementarity-determining region, Proceedings of the National Academy of Sciences of the United States of America, Vol.81, No.16, 5194-5198, 1984.
Nobuyuki Kobayashi, Hiroshi Konishi, Hisataka Sabe, Katsuya Shigesada, Takafumi Noma, Tasuku Honjo and Masakazu Hatanaka : Genomic structure of HTLV (human T-cell leukemia virus): detection of defective genome and its amplification in MT-2 cells, The EMBO Journal, Vol.3, No.6, 1339-1343, 1984.
(要約)
We studied the genomic structure of human T-cell leukemia virus (HTLV) in the HTLV producer cell line MT-2. Southern blotting revealed that at least eight HTLV proviruses were integrated in the chromosomes of MT-2 cells. The genomic structure of these proviruses was analyzed using fragments of cloned HTLV that were specific to gag, pol, env, pXs and U3R genes as probes. We have identified a complete genome of HTLV in MT-2 (non-defective type). However, seven of the eight proviruses had defective genomes. Provirus T2-a contains only the U3R (LTR) of HTLV and T2-b corresponds to the non-defective genome. T2-c possesses only a portion of env, and pXs and U3R. T2-d consists of gag, pol, part of env and U3R. On the other hand, T2-e, f, g and h consist of gag, pXs and U3R. Northern blotting experiments with mRNA from MT-2 cells supported the evidence of amplification of the gag-pXs gene of HTLV. 26S mRNA is considered to be a subgenomic species of 35S RNA. 32S mRNA may represent the T2-d provirus which lacks a portion of env and pXs, while 20S mRNA was a subgenomic species. The gag-pXs gene may correspond to 24S mRNA, the amount which was amplified in MT-2 cells.
We prepared career advancement program "Experience learning in medical team treatment" towards the second，third and fouth year students of dental school. This program consits of prior learningconcerned with 'Team treatment and Dental profession' and experience learning in medical team treatment at Tokushima University Hospital.The purpose of this program was to rouse career conciousness as a dentalprofession. This program provided students with career image which they want to be in the future. Besides，they were motivated to study more widely to approach their own image. In this paper，we evaluated theeffects of the program"The experience learning in medical team treatment" on career advancement ofdental school students. As a result，it is suggested that this program might be fruitful as career advancement on dental students.我々は歯学部学生のキャリア形成支援のための教育プログラム「チーム医療体験学習」を開発し，歯学部の2，3，4年次の学生を対象として実施した．本プログラムは，グループワークによる事前学習とチーム医療の現場体験を2つの柱とし，学生の歯科医療職としてのキャリア意識を強化することを目的とした．プログラム終了時のアンケートから，学生には歯科専門職としての将来の姿勢や目指す方向性が具体化すると同時に幅広い知識を身につけたいといった学習に対する動機付が強化されるなど明らかな教育成果が認められた．プログラム実施前後のアンケートの分析結果から， 「チーム医療体験学習」が歯学部学生のキャリア形成支援教育の手法として有効なプログラムである可能性が示唆された．
Takafumi Noma : Dynamics of nucleotide metabolism as a supporter of life phenomena, The Journal of Medical Investigation : JMI, Vol.52, No.3,4, 127-136, Aug. 2005.
(要約)
Adenylate kinase (hereinafter referred to as AK) catalyzes a reversible high-energy phosphoryl transfer reaction between adenine nucleotides. The enzyme contributes to the homeostasis of cellular adenine nucleotide composition in addition to the nucleotide biosynthesis. So far, six AK isozymes, AK1, AK2, AK3, AK4, AK5, and AK6, were identified. AK1 is localized in neuronal processes, sperm tail and on the cytoskeleton in cardiac cells at high concentrations, suggesting its regulatory function as a high-energy beta-phosphoryl transfer chain from ATP-synthesizing sites to the ATP-utilizing sites in the cell. AK2, AK3 and AK4 are mitochondrial proteins. AK2 is expressed in the intermembrane space, while AK3 and AK4 are localized in the mitochondrial matrix. AK3 is expressed in all tissues except for red blood cells indicating that AK3 gene is a housekeeping-type gene. On the other hand, AK4 is tissue-specifically expressed mainly in kidney, brain, heart, and liver although its enzymatic activity is not yet detected. AK5 is solely expressed in a limited area of brain. AK6 is recently identified in nucleus, suggesting its role in nuclear nucleotide metabolism. All data, so far reported, indicated the function of AK is associated with the mechanism of efficient transfer of high-energy phosphate in micro-compartment within the cell.
Roberts B Anita, Seong-Jin Kim, Takafumi Noma, Adam B Glick, Robert Lafyatis, Robert Lechieider, Sonia B Jakowlew, Andrew Geiser, Michael A O`Reilly, David Danielpour and Michael B Sporn : Multiple forms of TGF-β:distinct promoters and differential expression, Ciba Foundation Symposium, Vol.157, 7-28, 1991.
(要約)
There are now five known distinct isoforms of TGF-beta with 64-82% identity. Of these, only TGF-beta 1, 2 and 3 thus far have been demonstrated to be expressed in mammalian tissues; TGF-beta 4 has been described only in chicken and TGF-beta 5 only in frog. Although the biological activities of these five isoforms of TGF-beta are indistinguishable in most in vitro assays their sites of synthesis and localization in vivo are often distinct. Expression of the various isoforms is differentially controlled both in vivo, as in development, and in vitro after treatment of cells with steroids, such as oestrogen or tamoxifen, or with retinoids. To investigate the basis of these observations we have cloned and characterized the promoters for the human TGF-beta 1, 2 and 3 genes. Significant differences have been found: whereas the TGF-beta 1 promoter has no TATAA box and is regulated principally by AP-1 sites, both the TGF-beta 2 and 3 promoters have TATAA boxes as well as AP-2 sites and cAMP-responsive elements. Accordingly, TGF-beta 1 gene expression is induced strongly by phorbol esters whereas that of TGF-beta 2 and 3 is induced by forskolin, an activator of adenylate cyclase. Expression of TGF-beta 2 and 3 is often coordinately regulated in vivo in a pattern distinct from that of TGF-beta 1.
ANITA B. ROBERTS, SEONG-JIN KIM, PATURU KONDAIAH, SONIA B. JAKOWLEW, FABIENNE DENHEZ, ADAM B. GLICK, ANDREW G. GEISER, SHINICHI WATANABE, Takafumi Noma, ROBERT LECHLEIDER and SPORN B MICHAEL : Transcriptional Control of Expression of the TGF-βs, Annals of the New York Academy of Sciences, Vol.593, 43-50, Jan. 1990.
野間 隆文 : IL-5の遺伝子の構造と機能, Medical Immunology, Vol.15, No.1, 27-33, 1988年1月.
(キーワード)
IL-5 / TRF / BCGF- / EDF
18.
野間 隆文, 本庶 佑 : IL-4, 代謝, Vol.25, 449-455, 1988年.
(キーワード)
B細胞増殖因子 / B細胞刺激因子 / IgG1誘導因子 / インターロイキン4
国際会議:
1.
Keiko Miyoshi, Arinawati Yosi Dian, Hagita Hiroko, Taigo Horiguchi and Takafumi Noma : Possible roles of Sp6 in ameloblast differentiation, The 6th International Conference on Biology and Pathobiology of KLF/Sp Transcription Factors, Oct. 2018.
2.
Ayako Tanimura, Keiko Miyoshi, Taigo Horiguchi, Hagita Hiroko, Koichi Fujisawa and Takafumi Noma : A role of ER stress and UPR on hematopoietic differentiation, 51st Miami Winter Symposium, Stem Cells Todays Research Tomorrows Therapies, Miami, Jan. 2018.
3.
Arinawati Yosi Dian, Keiko Miyoshi, Ayako Tanimura, Taigo Horiguchi and Takafumi Noma : Comparative study of gene profiling using 2D and 3D culture as an in vitro amelogenesis imperfecta model, Internationl Joint meeting 4th ASEAN plus Tokushima Joint International Conference, Dec. 2017.
4.
Ayako Tanimura, Keiko Miyoshi, Taigo Horiguchi, Hagita Hiroko, Koichi Fujisawa and Takafumi Noma : Impact of mitochondrial ATP production on neutrophil differentiation, Gordon Research Conference(From Molecular Structures and Mechanisms to Cellular Bioenergetics in Health and Disease), Jun. 2017.
5.
Keiko Miyoshi, Hagita Hiroko, Arinawati Yosi Dian, Ayako Tanimura, Taigo Horiguchi and Takafumi Noma : Regulation of SP6 protein expression in stably transformed dental epithelial cells, 12th International Conference on Tooth Morphogenesis and Differentiation, Porvoo,Finland, Jun. 2016.
6.
Takafumi Noma : Potential and application of oral mucosa in regenerative medicine, National Colloquium on Stem Cell Research 2016, Malaysia, Mar. 2016.
7.
Keiko Miyoshi, Adiningrat Arya, Ayako Tanimura, Yanuaryska Dwi Ryna, Arinawati Yosi Dian, Taigo Horiguchi and Takafumi Noma : Establishment of an in Vitro amelogenesis imperfecta model, Challenge to Intractable Oral Diseases International Symposium 2015 which takes place at Yumikura Hall,Osaka University Graduate School of Dentistry in Japan from 10-11 December 2015, Dec. 2015.
(キーワード)
amelogenesis imperfecta / ARE / in vitro model / SP6 / structure-activity relationship
8.
Taigo Horiguchi, Keiko Miyoshi, Ayako Tanimura, H. Hagita, Y. Miyatake, Hiroshi Sakaue and Takafumi Noma : Gene expression analysis of hyperactive mutant SPORTS rat, Cell Symposia:Exercise and Metabolism which takes place at NH Grand Krasnapolsky Hotel Amsterdam from 12-14 July 2015, Amsterdam, Jul. 2015.
9.
Hiroko Hagita, Keiko Miyoshi, Taigo Horiguchi, Ayako Tanimura, Arya Adiningrat, Dian Yosi Arinawati and Takafumi Noma : Analysis of GBA1 gene structure and expression, The 11th International workshop on Advanced Genomics, Tokyo, May 2015.
10.
Keiko Miyoshi, Taigo Horiguchi, Ayako Tanimura, Hiroko Hagita, Yoshihiro Touda, Shoji Kagami, Kenji Mori, Daisuke Tsuji, Kouji Itou and Takafumi Noma : Gaucher disease caused by possible atypical mechanism, Gordon Research Conference, USA,Texas,Galveston(Hotel Galvez), Mar. 2015.
11.
Ayako Tanimura, Taigo Horiguchi, Keiko Miyoshi, Hiroko Hagita and Takafumi Noma : Adenylate Kinase 2 Regulates Neutrophil Differentiation Via Mitochondrial, The 3rd ASEAN Plus and Tokushima Joint International Conference, Makassar,Indonesia, Dec. 2014.
12.
Arya Adinigrat, Ayako Tanimura, Keiko Miyoshi, Ryna Dwi Yanuaryska, Hiroko Hagita, Taigo Horiguchi and Takafumi Noma : Ctip2 Regulation Of Tooth Development Via Sp6 Gene Expression, The 3rd ASEAN Plus and Tokushima Joint International Conference, Makassar,Indonesia, Dec. 2014.
13.
Susumu Tadokoro, Reiko Tokuyama, Seiko Tatehara, Shinji Ide, Hirochika Umeki, Tatsuhiro Fukushima, Keiko Miyoshi, Takafumi Noma and Kazuhito Satomura : A WEW INDUCTION METHOD FOR THE CONTROLLED DIFFERENTIATION OF HUMAN IPS CELLS USING FROZEN SECTIONS, INTERNATIONAL SOCIETY FOR STEM CELL RESEARCH, Vancouver, Jun. 2014.
14.
Ayako Tanimura, Taigo Horiguchi, Keiko Miyoshi, Hiroko Hagita and Takafumi Noma : A role of adenine nucleotide converting enzymes in the mitochondrial intermembrane space on the hematopoietic cell differentiation, Keystone meeting Mitochondrial Dynamics and Physiology, SantaFe, New Mexico, USA, Feb. 2014.
15.
Taigo Horiguchi, Miyuki Fuka, Koichi Fujisawa, Ayako Tanimura, Keiko Miyoshi, Ryutaro Murakami and Takafumi Noma : A Role of AK2 during Development of Drosohila melanogaster, The 4th International Symposium on Dynamics of Mitochondria, Oct. 2013.
16.
Arya Adiningrat, Keiko Miyoshi, Hiroko Hagita, Taigo Horiguchi, Ayako Tanimura, Ryna Dwi Yanuaryska and Takafumi Noma : Role of Bcl11b/Ctip2 on Sp6 Gene Expression in Dental Epithelial Cell, ASEAN PLUS and TOKUSHIMA JOINT INTERNATIONAL CONFERENCE, Dec. 2012.
17.
Ryna Dwi Yanuaryska, Keiko Miyoshi, Taigo Horiguchi, Ayako Tanimura, Hiroko Hagita, Arya Adiningrat and Takafumi Noma : SP6 Regulation of Rock1 Expression in Dental Epithelial Cells, ASEAN PLUS and TOKUSHIMA JOINT INTERNATIONAL CONFERENCE, Tokushima, Dec. 2012.
18.
Keiko Miyoshi, Taro Muto, Taigo Horiguchi, Hiroko Hagita and Takafumi Noma : A frameshift mutation in Sp6 linked to Amelogenesis imperfecta, ASEAN PLUS and TOKUSHIMA JOINT INTERNATIONAL CONFERENCE, Tokushima, Dec. 2012.
19.
Trianna W Utami, Keiko Miyoshi, Hiroko Hagita, Ryna D Yanuaryska, Taigo Horiguchi and Takafumi Noma : REGULATION OF SP6 GENE EXPRESSION AND CELL TYPE SPECIFIC FUNCTION OF SP6 IN DENTAL EPITHELIAL CELLS, The 2nd International Joint Sypmosium on Oral and Dental Sciences In Conjuction with Dental Specialists Seminar, Mar. 2012.
20.
Takafumi Noma : Perspective for the future regeneration therapy, Internationla Joint Symposium on Oral Science, Dec. 2010.
21.
Utami Wahyu Trianna, Keiko Miyoshi, Taigo Horiguchi, Taro Muto, Wahyudi Arie Ivan, Hiroko Hagita and Takafumi Noma : Differential regulation of Sp6 silencing in dental epithelial cells, Inter national Joint Symposium on Oral Science, Dec. 2010.
22.
Ivan Arie Wahyudi, Taigo Horiguchi, Keiko Miyoshi, Taro Muto, Trianna Wahyu Utami, Hagita Hiroko and Takafumi Noma : Caracterization of Specificity Protein 6 promoter Activity, International Joint Symposium on Oral Science, Dec. 2010.
23.
Takafumi Noma : Introduction of Research and Study in Tokushima University, ムハマディア大学 招待講演, Dec. 2010.
24.
Takafumi Noma : Basic Aspect of Regeneration Therapy Generation of iPS Cells from Oral Mucosa and Its Application, Recent Advance in Dentistry, Oct. 2010.
25.
Taro Muto, Keiko Miyoshi and Takafumi Noma : Differentiation of the pigmentation ameloblasts into the pigment release stage is disturbed in incisors of Sp6 transgenic rats, Gordon Resarch Conferences, Apr. 2010.
26.
Takafumi Noma, Taro Muto, Keiko Miyoshi, Taigo Horiguchi, Hiroko Hagita, Wahyudi Arie Ivan and Utami Wahyu Trianna : From a study of tooth morphology and development to a perspective for the future regeneration therapy, ガジャマダ大学 創立62年 記念講演会, Feb. 2010.
27.
Ruspita Intan, Keiko Miyoshi, Taro Muto, Kaori Abe, Taigo Horiguchi and Takafumi Noma : Identification of Sp6 target gene in dental epithelial cells, Gordon Research Conferences, Feb. 2008.
28.
Keiko Miyoshi, Taigo Horiguchi, Kazuki Abe, Inoue Hideo and Takafumi Noma : Effects of glycyrrhizin on the gene expression in CC14-induced hepatitis, 8th World Congress on Inflammation, Jun. 2007.
29.
Takafumi Noma, Keiko Miyoshi, Yuki Akazawa and Taigo Horiguchi : Tissue-ditribution and possible functional roles of AK4, Mitchondrial Medicine 2007: Riding the Wave of the Future, Jun. 2007.
30.
Takafumi Noma, Keiko Miyoshi, Yuki Akazawa and Taigo Horiguchi : Tissue-distribution and possible functional roles of AK4, Mitochondrial Medicine Meeting, San Diego, Jun. 2007.
Kaori Abe, Keiko Miyoshi, Taro Muto, Intan Ruspita, Taigo Horiguchi, Toshihiko Nagata and Takafumi Noma : Establishment and charactarization of rat ameloblast-lineage clones, The 19th Annual and International Meeting of the Japanese Association for Animal Cell Technology, Kyoto, Sep. 2006.
32.
Koichi Fujisawa, Ryutaro Murakami and Takafumi Noma : Adenine nucleotide metabolism in the mitochondrial intermembrane space is essential for growth and development in Drosopila melanogaster, 20th IUBMB international Congress of Biochemistry and Molecular biology and 11th FAOBMB Congress, Kyoto, Jun. 2006.
33.
Taro Muto, Keiko Miyoshi, Hiroshi Nakata, Seiichi Munesue, Minoru Okayama, Takashi Matsuo and Takafumi Noma : Comparative analysis of syndecan expression during mouse incisor development, Extracellular Glycomatrix in health and Disease symposium, Awaji, Jun. 2006.
34.
Intan Ruspita, Taigo Horiguchi, Keiko Miyoshi, Akemichi Ueno, Hidemitsu Harada and Takafumi Noma : Regulation of Gene Expression in Dental Epithelial Cells, Eighth International Conference on Tooth Morphogenesis and Differentiation, York, UK, Jul. 2004.
35.
Keiko Miyoshi, Taigo Horiguchi, Kaori Abe, Akemichi Ueno, Hideya Nagata, Yoshinobu Baba, Hidemitsu Harada and Takafumi Noma : Effects of BMP2 on Gene Expression in Dental Epithelial Cell Line, Eight International Conference on Tooth Morphogenesis and Differentiation, York, UK, Jul. 2004.
36.
Takafumi Noma : Control of Inflammation by a Medicinal Herb, The UAE-Japan Joint Workshop Program for Traditional & Herbal Medicine and Recent Topics in Life Science, Hiroshima, Feb. 2004.
37.
Takafumi Noma : Structure and expression of hAK4 targeted to the mitochondrial matrix, 11th International Congress on Genes,Gene Families and Isozymes, Stockholm, Jun. 2001.
38.
S Inooue, Takafumi Noma and A Nakazawa : Tissue-specific expression and physiological significance of adenylate kinase isozymes, 9th international Congress on Genes,Gene families,and Isozymes, San Antonio,Texas,USA, 1997.
39.
S Terai, K Uchida, T Kimura, I Sakaida, K Okita and Takafumi Noma : JNK/MEKK cascade induced the enhancement of transcriptional activation of p53, AACR Special Conference in Cancer Research:Inducible Genomic Responses, Stevenson,WA,USA, 1996.
40.
S Terai, T Kimura, K Okita and Takafumi Noma : Serum starvation induced morphological change and growth suppression in hepatocellular carcinoma cell line after introduction of p53 gene, AACR Special Conference in Cancer Research:The Molecular Basis of Gene Transcription, San Diego, 1995.
41.
A Nakazawa, M Nobumoto, S Song, S Inoue and Takafumi Noma : Compartmentation of adenylate kinase isozymes, 8th International Congress on Isozymes, Brisbane,Australia, 1995.
42.
Takafumi Noma, H Ogasa and A Nakazawa : Heterogeneity of TGF-b type 2 receptor cDNAs, The Eight International Conference of the International Society of differentiation, Hiroshima, 1994.
43.
S Terai, K Okita and Takafumi Noma : MDM2 and p53 interaction during hepatocyte growth, The Eight international Conference of the International Society of differentiation, Hiroshima, 1994.
44.
Takafumi Noma, H Ogasa and A Nakazawa : Heterogeneity of TGF-b type 2 receptor, Fogarty International Center Conference:TGF-bs:Biological mechanisms and clinical Applications, Bethesda,Maryland,USA, 1994.
45.
M Nobumoto, M Yamada, S Inoue, Takafumi Noma and A Nakazawa : In vitro import of adenylate kinase isozymes into mitochondria, International Symposium "Dynamics in Biological Functions of Adenylate Metabolism", Ube, 1994.
46.
N Adachi, Md Shahjahan, Takafumi Noma, H Itoh and A Nakazawa : Molecular cloning and characterization of the gene encoding bovine and human mitochondrial adenylate kinase isozyme 3, First IUBMB Conference,Biochemistry of Diseases, Nagoya, 1994.
47.
Takafumi Noma, A Glick, A Geiser, E J Miller, B A Roberts and B M Sporn : Molecular cloning and characterization of the promoter of the transforming growth factor-beta 2 gene, UCLA SYMPOSIA ON MOLECULAR&CELLULAR BIOLOGY, Taos,NewMexico,USA, 1990.
Arinawati Yosi Dian, Keiko Miyoshi, Hagita Hiroko, Taigo Horiguchi and Takafumi Noma : Perturbation of gene regulateion in an in vitro amelogenesis imperfecta model, The 91st Annual Meeting of the Japanese Biochemical Society, Sep. 2018.
Arinawati Yosi Dian, Keiko Miyoshi, Hagita Hiroko, Taigo Horiguchi and Takafumi Noma : Demonstration of defective amelogenesis using an in vitro amelogenesis imperfecta model, 第257回徳島医学会学術集会, Aug. 2018.
Utami Wahyu Trianna, Keiko Miyoshi, Hiroko Hagita, Yanuaryska D Ryna, Ayako Tanimura and Takafumi Noma : Regulation of Sp6 expression and function in dental epithelial cells, 第5回 日本エピジェネティクス研究会, May 2011.
37.
Utami Wahyu Triannna, Keiko Miyoshi, Taigo Horiguchi, Wahyudi A Ivan, Hiroko Hagita, Yanuaryska D Ryna and Takafumi Noma : Regulation of Sp6 expression and function in C9 cells, 第52回 日本生化学会中国 四国支部例会, May 2011.
38.
Utami Wahyu Trianna, Keiko Miyoshi, Taigo Horiguchi, Taro Muto, Wahyudi Arie Ivan, Hiroko Hagita and Takafumi Noma : Regulation of Sp6 expression in CHA9 cells, 第33回 日本分子生物学会 第83回日本生化学会, Dec. 2010.
39.
Wahyudi Arie Ivan, Taigo Horiguchi, Keiko Miyoshi, Taro Muto, Utami Wahyu Trianna, Hiroko Hagita and Takafumi Noma : Isolation and Characterization of Mouse Specificity Protein 6 Promoter, 第33回 日本分子生物学会 第83回 日本生化学会, Dec. 2010.
阿部 佳織, 三好 圭子, 武藤 太郎, 堀口 大吾, INTAN RUSPITA, 野間 隆文, 永田 俊彦 : MAPK activation via ROS induces amelognin expression in the ameloblast-lineage cells, 「魅力ある大学院教育」イニシアティブ 「21世紀の口腔科学が目指すべき方向性」, 2007年.
52.
Akemichi Ueno, Kikuji Yamashita, Keiko Miyoshi, Taigo Horiguchi, Ruspita Intan, Kaori Abe and Takafumi Noma : Overexpression of thrombospondin1(TSP1) inhibits mineralization by MC3T3 cells in vivo, 77th mass Meeting of the Japanese Biochemical Society, Oct. 2004.
Takafumi Noma, A Glick, A Geiser, J Miller, M O`Reilly, A Roberts and M Sporn : Molecular cloning and characterization of the promoter of the transforming growth factor-beta 2 gene, 19th Annual Meetings UCLA SYMPOSIA ON MOLECULAR&CELLULAR BIOLOGY, 307, Feb. 1990.