Publications (8)14.81 Total impact
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Article: Developmental and age-related changes of peptidylarginine deiminase 2 in the mouse brain.
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ABSTRACT: Peptidylarginine deiminases (PADs) are a group of posttranslational modification enzymes that citrullinate (deiminate) protein arginine residues in a Ca(2+)-dependent manner. Enzymatic citrullination abolishes positive charges of native protein molecules, inevitably causing significant alterations in their structure and functions. Among the five isoforms of PADs, PAD2 and PAD4 are proved occupants of the central nervous system (CNS), and especially PAD2 is a main PAD enzyme expressed in the CNS. We previously reported that abnormal protein citrullination by PAD2 has been closely associated with the pathogenesis of neurodegenerative disorders such as Alzheimer's disease and prion disease. Protein citrullination in these patients is thought to play a role during the initiation and/or progression of disease. However, the contribution of changes in PAD2 levels, and consequent citrullination, during developmental and aging processes remained unclear. Therefore, we used quantitative real-time RT-PCR, Western blot analysis, and immunohistochemical methods to measure PAD2 expression and localization in the brain during those processes. PAD2 mRNA expression was detected in the brains of mice as early as embryonic day 15, and its expression in cerebral cortex, hippocampus, and cerebellum increased significantly as the animals aged from 3 to 30 months old. No citrullinated proteins were detected during that period. Moreover, we found here, for the first time, that PAD2 localized specifically in the neuronal cells of the cerebral cortex and Purkinje cells of the cerebellum. These findings indicate that, despite PAD2's normally inactive status, it becomes active and citrullinates cellular proteins, but only when the intracellular Ca(2+) balance is upset during neurodegenerative changes.Journal of Neuroscience Research 10/2009; 88(4):798-806. · 2.74 Impact Factor -
Article: Elevated levels of 4-hydroxynonenal-histidine Michael adduct in the hippocampi of patients with Alzheimer's disease.
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ABSTRACT: Alzheimer's disease (AD) is among the most common causes of progressive cognitive impairment in humans and is characterized by neurodegeneration in the brain. Lipid peroxidation is thought to play a role in the pathogenesis of AD. 4-hydroxynonenal (HNE) results from peroxidation of polyunsaturated fatty acids and it in turn gives evidence of lipid peroxidation in vivo. HNE reacts with protein histidine residue to form a stable HNE-histidine Michael adduct. To clarify the influence of lipid peroxidation on the pathogenesis of AD, we measured HNE-histidine Michael adduct in hippocampi from four AD patients and four age-matched controls by means of semiquantitative immunohistochemistry using a specific antibody to cyclic hemiacetal type of HNE-histidine Michael adduct. This antibody does not react with the ring-opened form of HNE-histidine Michael adduct and the pyrrole form of HNE-lysine Michael adduct. The HNE adduct was detected in the hippocampi of both AD and control donors, especially in the CA2, CA3 and CA4 sectors. Immunoreactive intensity of HNE adduct in these sectors were significantly higher in AD patients than in the controls. The HNE adduct was found in the perikarya of pyramidal cells in the hippocampus. These results show that the hippocampi of patients with AD undergo lipid peroxidation and imply that this activity underlies the production of cytotoxic products such as HNE that are responsible for the pathogenesis of AD.Biomedical Research 09/2009; 30(4):227-33. · 1.15 Impact Factor -
Article: Vitamin C depletion increases superoxide generation in brains of SMP30/GNL knockout mice.
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ABSTRACT: Vitamin C (VC) has a strong antioxidant function evident as its ability to scavenge superoxide radicals in vitro. We verified that this property actually exists in vivo by using a real-time imaging system in which Lucigenin is the chemiluminescent probe for detecting superoxide in senescence marker protein-30 (SMP30)/gluconolactonase (GNL) knockout (KO) mice, which cannot synthesize VC in vivo. SMP30/GNL KO mice were given 1.5 g/L VC [VC(+)] for 2, 4, or 8 weeks or denied VC [VC(-)]. At 4 and 8 weeks, VC levels in brains from VC(-) KO mice were <6% of that in VC(+) KO mice. Accordingly, superoxide-dependent chemiluminescence levels determined by ischemia-reperfusion at the 4- and 8 weeks test intervals were 3.0-fold and 2.1-fold higher, respectively, in VC(-) KO mice than in VC(+) KO mice. However, total superoxide dismutase activity and protein levels were not altered. Thus, VC depletion specifically increased superoxide generation in a model of the living brain.Biochemical and Biophysical Research Communications 10/2008; 377(1):291-6. · 2.48 Impact Factor -
Article: Molecular evolution of nucleoside diphosphate kinase genes: conserved core structures and multiple-layered regulatory regions.
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ABSTRACT: Genomic data regarding the nucleoside diphosphate (NDP) kinase genes have been accumulated from diverged phyla. Comparison of their regulatory sequences have shed light on the multiple facets of gene regulation systems. Phylogenetic studies, including CpG island and intron-mapping, and homologous sequence comparison, have suggested that the regions of the major mammalian genes, the ortholog (rat alpha or nm23-H2) and its paralog (rat beta or nm23-H1), have been constructed by a stepwise gain and loss of alien genes resulting in "multiple-layered" regulatory systems. They contain representative cis-elements for the constitutive, stage/lineage-specific, and early response expression. These elements' binding capacities to nuclear proteins were confirmed by electrophoretic mobility shift assay. Further, these regulatory systems generate heterogeneous mRNA at the 5' untranslated region, which influences their own translation efficiencies. In terms of this process, the transcription system would control another layer of gene expression: posttranscriptional (translational) regulation.Journal of Bioenergetics 03/2003; 35(1):7-18. · 2.81 Impact Factor -
Article: Nucleoside diphosphate kinases in mammalian signal transduction systems: recent development and perspective.
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ABSTRACT: The role of nucleoside diphosphate (NDP) kinase with special reference to mammalian signal transduction systems was described. The interaction between NDP kinases and G proteins was reevaluated in view of their protein structural information and its significance was extended further on the basis of recent findings obtained with small molecular weight G proteins such as Rad, menin, and Rac. Meanwhile, observations suggesting involvement of NDP kinases in the regulation of cell growth and differentiation led to the realization that NDP kinases may play a crucial role in receptor tyrosine kinase signal transduction systems. In fact, a number of experimental results, particularly obtained with PC12 cells, implicate that NDP kinases appear to regulate differentiation marker proteins and cell-cycle-associated proteins cooperatively. Consequently, we propose a hypothesis that NDP kinases might act like a molecular switch to determine the cell fate toward proliferation or differentiation in response to environmental signals.Journal of Bioenergetics 03/2003; 35(1):41-7. · 2.81 Impact Factor -
Article: Regulation of Cellular Functions by Nucleoside Diphosphate Kinases in Mammals
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ABSTRACT: The role of nucleoside diphosphate (NDP) kinases in cell growth, differentiation, and tumormetastasis in relation to signal transduction was investigated. The essential role of NDP kinasein cell growth was validated by coupling between reduced NDP kinase levels, induced byantisense oligonucleotides, and the suppression of proliferative activity of a cultured cell line.In addition, because NDP kinase levels are often enhanced with development and differentiation,as has been demonstrated in postmitotic cells and tissues, such as the heart and brain, wefurther examined this possibility using the bone tissue (osteoblasts) and a cultured cell linePC12D. The enhanced NDP kinase accumulation was demonstrated in the matured osteoblastsin vivo and in vitro by immunohistochemistry. In PC12D cells neurite outgrowth took placein NDP kinase -transfected clones without differentiation inducers, which was accompaniedby prolongation of doubling time. Neurite outgrowth, triggered by nerve growth factor and acyclic AMP analog, was down-regulated upon forced expression of inactive mutant NDPkinase by virtue of a dominant negative effect. NDP kinase -transfected rat mammaryadenocarcinoma cells (MTLn3) and nm23-H2-transfected human oral squamous cell carcinomacells (LMF4) manifested reduced metastatic potential and were associated with an alteredsensitivity to environmental factors, such as motility and growth factors. NDP kinase ,compared to NDP kinase , was involved in a wide variety of the cellular phenomena examined.Taken together, NDP kinase isoforms appear to elicit both their own respective and commoneffects. They may have an ability to lead cells to both proliferative and differentiated statesby modulating responsiveness to environmental factors, but their fate seems to depend on theirsurrounding milieu.Journal of Bioenergetics 05/2000; 32(3):309-315. · 2.81 Impact Factor -
Article: Recombinant rat nucleoside diphosphate kinase isoforms (α and β): purification, properties and application to immunological detection of native isoforms in rat tissues
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ABSTRACT: We previously demonstrated that at least two isoforms of nucleoside diphosphate (NDP) kinase, the products of two different tandemly arrayed genes, are present in rat. To understand the physiological role of each isoform, some biochemical properties of recombinant rat NDP kinase α- and β-isoforms, produced in large amount, were studied, cDNAs of the two isoforms were inserted in an expression vector pET3b and recombinant enzymes were overproduced in Escherichia coli. Their primary structures were different from the native enzymes in that the latter suffer from modification of the NH2-terminal end. The two recombinant isoforms were purified from the cell lysate to apparent homogeneity by ammonium sulfate fractionation followed by three successive column chromatographies. Despite their extreme similarity in the amino-acid sequences, the two showed somewhat different enzymic properties in terms of di- and triphosphate nucleotide substrate specificity. They showed similar mobilities on SDS-PAGE as expected from their calculated molecular weight (α-isoform, 17 283 versus β-isoform, 17 192) but differed in isoelectric point (α-isoform, pI 6.7; β-isoform, pI 7.8) and heat stability. Polyclonal antibody which reacted with both isoforms and α-isoform-specific monoclonal antibodies differentially recognized native enzymes from rat tissues after the tissue extracts were separated by isoelectric focusing gel electrophoresis under a denaturation condition. The results showed that the α-isoform, though its amount varied from one tissue to another, was the major form in rat tissues examined compared with the β-isoform which was detectable in brain and testis. There was no preference in their subcellular localization when examined with myelin, synaptosomal supernatant and total homogenate fractions from the rat cerebrum and cerebellum.Biochimica et Biophysica Acta (BBA) - Protein Structure and Molecular Enzymology. -
Article: Estradiol up-regulates the stimulatory GTP-binding protein expression in the MCF-7 human mammary carcinoma cell line
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ABSTRACT: The effect of estradiol treatment of the human mammary carcinoma cell MCF-7 on the adenylyl cyclase system was examined. Treatment with 10 nM estradiol for 72 h increased the basal level ofcAMP, and isoproterenol-, PGE2- or calcitonin-stimulated cAMP production. Estradiol also increased the response to cholera toxin but did not alter the response to forskolin. No significant change in growth rate was observed during the 72 h of estradiol treatment. In MCF-7 cell membranes the responsiveness to isoproterenol, PGE2, or cholera toxin was also enhanced by estradiol treatment. The cholera toxin-catalyzed ADP-ribosylation of gsoe in MCF-7 cell membranes was significantly increased by 72 h of treatment with estradiol. Consistent with this observation, the level of Gsα immunoreactivity was increased in the estradiol-treated cell membranes. On the other hand, pertussis toxin did not change the responsiveness to isoproterenol, PGE2 or calcitonin in either control or estradiol-treated cells. In addition, ADP-ribosylation with pertussis toxin also did not reveal any change in Gi. These results clearly indicate that Gs expression is under the control of estradiol, and that this effect may contribute to the increased sensitivity of hormone-stimulated adenylyl cyclase activities in MCF-7 cells.FEBS Letters.
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Institutions
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2008–2009
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Tokyo Metropolitan Institute of Gerontology
Tokyo, Tokyo-to, Japan
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