||Yuichiro Koga, Daisuke Tsuchimoto, Yoshinori Hayashi, Nona Abolhassani, Yasuto Yoneshima, Kunihiko Sakumi, Hiroshi Nakanishi, Shinya Toyokuni, Yusaku Nakabeppu, Neural stem cell-specific ITPA deficiency causes neural depolarization and epilepsy, JCI Insight, in press, 2020.11, Inosine triphosphate pyrophosphatase (ITPA) hydrolyzes inosine triphosphate (ITP) and other deaminated purine nucleotides to the corresponding nucleoside monophosphates. In humans, ITPA deficiency causes severe encephalopathy with epileptic seizure, microcephaly, and developmental retardation. In this study, we established neural stem cell-specific conditional Itpa-KO mice (Itpa-cKO mice) to clarify the effects of ITPA deficiency on the neural system. The Itpa-cKO mice showed growth retardation and died within three weeks of birth. We did not observe any microcephaly in the Itpa-cKO mice although the female Itpa-cKO mice did show adrenal hypoplasia. The Itpa-cKO mice showed limb clasping upon tail suspension and spontaneous and/or audiogenic seizure. Whole-cell patch-clamp recordings from entorhinal cortex neurons in brain slices revealed a depolarized resting membrane potential, increased firing, and frequent spontaneous miniature excitatory postsynaptic current (mEPSC) and miniature inhibitory postsynaptic current (mIPSC) in the Itpa-cKO mice compared to ITPA-proficient controls. Accumulated ITP or its metabolites, such as cyclic inosine monophosphates, or RNA containing inosines may cause membrane depolarization and hyperexcitability in neurons, and induce the phenotype of ITPA-deficient mice, including seizure..
||Shinji Asada, Eiko Ohta, Yoriko Akimoto, ABOLHASSANI NONA, Daisuke Tsuchimoto, Yusaku Nakabeppu, 2-Oxoadenosine induces cytotoxicity through intracellular accumulation of 2-oxo-ATP and depletion of ATP but not via the p38 MAPK pathway, SCIENTIFIC REPORTS, 10.1038/s41598-017-06636-8, 7, 2017.07.
||M Massaad, J Zhou, Daisuke Tsuchimoto, J Chou, H Jabara, E Janssen, S Glauzy, B Olson, H Morbach, T Ohsumi, K Schmitz-Abe, M Kyriacos, J Kane, Kumiko Torisu, Yusaku Nakabeppu, LD Notarangelo, E Chouery, A Megarbane, PB Kang, Deficiency of the base excision repair enzyme NEIL3 is associated with increased lymphocyte apoptosis, autoantibodies and predisposition to autoimmunity., The Journal of Clinical Investigation, in press, 2016.10.
||Yasuto Yoneshima, Nona Abolhassani, Teruaki Iyama, Kunihiko Sakumi, Naoko Shiomi, Masahiko Mori, Tadahiro Shiomi, Tetsuo Noda, Daisuke Tsuchimoto, Yusaku Nakabeppu, Deoxyinosine triphosphate induces MLH1/PMS2- and p53-dependent cell growth arrest and DNA instability in mammalian cells., Scientific Reports, 10.1038/srep32849, 6, 32849, 2016.09, Deoxyinosine (dI) occurs in DNA either by oxidative deamination of a previously incorporated deoxyadenosine residue or by misincorporation of deoxyinosine triphosphate (dITP) from the nucleotide pool during replication. To exclude dITP from the pool, mammals possess specific hydrolysing enzymes, such as inosine triphosphatase (ITPA). Previous studies have shown that deficiency in ITPA results in cell growth suppression and DNA instability. To explore the mechanisms of these phenotypes, we analysed ITPA-deficient human and mouse cells. We found that both growth suppression and accumulation of single-strand breaks in nuclear DNA of ITPA-deficient cells depended on MLH1/PMS2. The cell growth suppression of ITPA-deficient cells also depended on p53, but not on MPG, ENDOV or MSH2. ITPA deficiency significantly increased the levels of p53 protein and p21 mRNA/ protein, a well-known target of p53, in an MLH1-dependent manner. Furthermore, MLH1 may also contribute to cell growth arrest by increasing the basal level of p53 activity..
||Jeroen E. J. Guikema, Erin K. Linehan, Nada Esa, Daisuke Tsuchimoto, Yusaku Nakabeppu, Robert T. Woodland, Carol E. Schrader, Apurinic/Apyrimidinic Endonuclease 2 Regulates the Expansion of Germinal Centers by Protecting against Activation-Induced Cytidine Deaminase-Independent DNA Damage in B Cells., The Journal of Immunology, 2014 Jun 16. pii: 1400002. [Epub ahead of print], 2014.06, Activation-induced cytidine deaminase (AID) initiates a process generating DNA mutations and breaks in germinal center (GC) B cells that are necessary for somatic hypermutation and class-switch recombination. GC B cells can "tolerate" DNA damage while rapidly proliferating because of partial suppression of the DNA damage response by BCL6. In this study, we develop a model to study the response of mouse GC B cells to endogenous DNA damage. We show that the base excision repair protein apurinic/apyrimidinic endonuclease (APE) 2 protects activated B cells from oxidative damage in vitro. APE2-deficient mice have smaller GCs and reduced Ab responses compared with wild-type mice. DNA double-strand breaks are increased in the rapidly dividing GC centroblasts of APE2-deficient mice, which activate a p53-independent cell cycle checkpoint and a p53-dependent apoptotic response. Proliferative and/or oxidative damage and AID-dependent damage are additive stresses that correlate inversely with GC size in wild-type, AID-, and APE2-deficient mice. Excessive double-strand breaks lead to decreased expression of BCL6, which would enable DNA repair pathways but limit GC cell numbers. These results describe a nonredundant role for APE2 in the protection of GC cells from AID-independent damage, and although GC cells uniquely tolerate DNA damage, we find that the DNA damage response can still regulate GC size through pathways that involve p53 and BCL6..
||Janet Stavnezer, Erin K. Linehan, Mikayla R. Thompson, Ghaith Habboub, Anna J. Ucher, Tatenda Kadungure, Daisuke Tsuchimoto, Yusaku Nakabeppu, Carol E. Schrader, Differential expression of APE1 and APE2 in germinal centers promotes error-prone repair and A:T mutations during somatic hypermutation, Proceedings of the National Academy of Sciences of the United States of America , Jun 9. pii: 201405590. [Epub ahead of print], 2014.06, Somatic hypermutation (SHM) of antibody variable region genes is
initiated in germinal center B cells during an immune response by
activation-induced cytidine deaminase (AID), which converts cytosines
to uracils. During accurate repair in nonmutating cells, uracil
is excised by uracil DNA glycosylase (UNG), leaving abasic sites
that are incised by AP endonuclease (APE) to create single-strand
breaks, and the correct nucleotide is reinserted by DNA polymerase
β. During SHM, for unknown reasons, repair is error prone. There
are two APE homologs in mammals and, surprisingly, APE1, in contrast
to its high expression in both resting and in vitro-activated
splenic B cells, is expressed at very low levels in mouse germinal
center B cells where SHM occurs, and APE1 haploinsufficiency has
very little effect on SHM. In contrast, the less efficient homolog,
APE2, is highly expressed and contributes not only to the frequency
of mutations, but also to the generation of mutations at A:T base
pair (bp), insertions, and deletions. In the absence of both UNG and
APE2, mutations at A:T bp are dramatically reduced. Single-strand
breaks generated by APE2 could provide entry points for exonuclease
recruited by the mismatch repair proteins Msh2–Msh6, and the
known association of APE2 with proliferating cell nuclear antigen
could recruit translesion polymerases to create mutations at AIDinduced
lesions and also at A:T bp. Our data provide new insight
into error-prone repair of AID-induced lesions, which we propose is
facilitated by down-regulation of APE1 and up-regulation of APE2
expression in germinal center B cells..
||Hiroko Nomaru, SAKUMI Kunihiko, Atsuhisa Katogi, Yoshinori N. Ohnishi, Kosuke Kajitani, Daisuke Tsuchimoto, Eric J. Nestler, Yusaku Nakabeppu, Fosb gene products contribute to excitotoxic microglial activation by regulating the expression of complement C5a receptors in microglia., Glia, 10.1002/glia.22680. , 62, 8, 1284-1298, Epub 2014 Apr 25., 2014.08, The Fosb gene encodes subunits of the activator protein-1 transcription factor complex. Two mature mRNAs, Fosb and ΔFosb, encoding full-length FOSB and ΔFOSB proteins respectively, are formed by alternative splicing of Fosb mRNA. Fosb products are expressed in several brain regions. Moreover, Fosb-null mice exhibit depressive-like behaviors and adult-onset spontaneous epilepsy, demonstrating important roles in neurological and psychiatric disorders. Study of Fosb products has focused almost exclusively on neurons; their function in glial cells remains to be explored. In this study, we found that microglia express equivalent levels of Fosb and ΔFosb mRNAs to hippocampal neurons and, using microarray analysis, we identified six microglial genes whose expression is dependent on Fosb products. Of these genes, we focused on C5ar1 and C5ar2, which encode receptors for complement C5a. In isolated Fosb-null microglia, chemotactic responsiveness toward the truncated form of C5a was significantly lower than that in wild-type cells. Fosb-null mice were significantly resistant to kainate-induced seizures compared with wild-type mice. C5ar1 mRNA levels and C5aR1 immunoreactivity were increased in wild-type hippocampus 24 hours after kainate administration; however, such induction was significantly reduced in Fosb-null hippocampus. Furthermore, microglial activation after kainate administration was significantly diminished in Fosb-null hippocampus, as shown by significant reductions in CD68 immunoreactivity, morphological change and reduced levels of Il6 and Tnf mRNAs, although no change in the number of Iba-1-positive cells was observed. These findings demonstrate that, under excitotoxicity, Fosb products contribute to a neuroinflammatory response in the hippocampus through regulation of microglial C5ar1 and C5ar2 expression..
||Zijing Sheng, Sugako Oka, Daisuke Tsuchimoto, Nona Abolhassani, Hiroko Nomaru, SAKUMI Kunihiko, Hideaki Yamada, Yusaku Nakabeppu, 8-Oxoguanine causes neurodegeneration during MUTYH-mediated DNA base excision repair, The Journal of Clinical Investigation, 10.1172/JCI65053, 122, 12, 4344-4361, 2012.12.
||Iwama E., Tsuchimoto D., Iyama T., Sakumi K., Nakagawara A., Takayama K., Nakanishi Y., and Nakabeppu Y., Cancer-related PRUNE2 protein is associated with nucleotides and is highly expressed in mature nerve tissues., Journal of Molecular Neuroscience
, Epub ahead of print, 2011.01.
||Jeroen E.J. Guikema, Rachel M. Gerstein, Erin K. Linehan, Erin K. Clohert, Eric Evan-Browning, Daisuke Tsuchimoto, Yusaku Nakabeppu, and Carol E. Schrader, AP-Endonuclease 2 is necessary for normal B cell development and recovery of lymphoid progenitors after chemotherapeutic challenge., Journal of Immunology, 186(4), 1943-50, 2011.02.
||Iyama T, Abolhassani N, Tsuchimoto D, Nonaka M, Nakabeppu Y., NUDT16 is a (deoxy)inosine diphosphatase, and its deficiency induces accumulation of single-strand breaks in nuclear DNA and growth arrest., Nucleic Acids Research, doi:10.1093/nar/gkq249 , 38(14), 4834-43, 2010.08.
||Abolhassani N, Iyama T, Tsuchimoto D, Sakumi K, Ohno M, Behmanesh M, Nakabeppu Y., NUDT16 and ITPA play a dual protective role in maintaining chromosome stability and cell growth by eliminating dIDP/IDP and dITP/ITP from nucleotide pools in mammals., Nucleic Acids Res., 2010 May;38(9):2891-903., 2010.05.
||Behmanesh M, Sakumi K, Abolhassani N, Toyokuni S, Oka S, Ohnishi YN, Tsuchimoto D, Nakabeppu Y, ITPase-deficient mice show growth retardation and die before weaning, Cell Death and Differentiation , 2009 Jun 5. [Epub ahead of print], 2009.06.
||Nonaka M, Tsuchimoto D, Sakumi K, Nakabeppu Y. , Mouse RS21-C6 is a mammalian 2'-deoxycytidine 5'-triphosphate pyrophosphohydrolase that prefers 5-iodocytosine. , FEBS J. , 276(6):1654-1666, 2009.03.
||Kajitani K, Nomaru H, Ifuku M, Yutsudo N, Dan Y, Miura T, Tsuchimoto D, Sakumi K, Kadoya T, Horie H, Poirier F, Noda M, Nakabeppu Y., Galectin-1 promotes basal and kainate-induced proliferation of neural progenitors in the dentate gyrus of adult mouse hippocampus., Cell Death Differ., 16(3):417-427, 2009.03.
||Dan Y, Ohta Y, Tsuchimoto D, Ohno M, Ide Y, Sami M, Kanda T, Sakumi K, Nakabeppu Y., Altered gene expression profiles and higher frequency of spontaneous DNA strand breaks in APEX2-null thymus., DNA Repair (Amst), 7(9):1437-1454., 2008.09.
||Oka S, Ohno M, Tsuchimoto D, Sakumi K, Furuichi M, Nakabeppu Y, Two distinct pathways of cell death triggered by oxidative damage to nuclear and mitochondrial DNAs., The EMBO Journal, 27(2):421-432, 2008.01.
||Guikema JEJ, Kinehan EK, Tsuchimoto D, Nakabeppu Y, Strauss PR, Stavnezer J, and Schrader CE, APE1- and APE2-dependent DNA breaks in immunoglobulin class switch recombination, The Journal of Experimental Medicine, 204(12):3017-3026, 2007.11.
||Ohno M., Miura T., Furuichi M., Tominaga Y., Tsuchimoto D., Sakumi K. and Nakabeppu Y., A genome-wide distribution of 8-oxoguanine correlates with the preferred regions for recombination and single-nucleotide polymorphism in the human genome., Genome Research, 16:567-575, 2006.05.
||Torisu K., Tsuchimoto D., Ohnishi Y. and Nakabeppu Y., Hematopoietic tissue-specific expression of mouse Neil3 for Endonuclease VIII-like protein., The Journal of Biochemistry, 10.1093/jb/mvi168, 138, 6, 763-772, 138(6):763-772, 2005.12.
||Ushijima Y., Tominaga Y, Miura T, Tsuchimoto D, Sakumi K and Nakabeppu Y., A functional analysis of the DNA glycosylase activity of mouse MUTYH protein excising 2-hydroxyadenine opposite guanine in DNA., Nucleic Acids Research, 10.1093/nar/gki214, 33, 2, 672-682, 33(2):672-682, 2005.01.
||Behmanesh B., Sakumi K., Tsuchimoto D., Torisu K., Ohnishi-Honda Y., Rancourt D.E. and Nakabeppu Y., Characterization of the structure and expression of mouse Itpa gene and its related sequences in the mouse genome., DNA Research, 10.1093/dnares/12.1.39, 12, 1, 39-51, 12:29-41, 2005.02.
||Ide Y, Tsuchimoto D, Tominaga Y, Nakashima M, Watanabe T, Sakumi K, Ohno M, Nakabeppu Y., Growth retardation and dyslymphopoiesis accompanied by G2/M arrest in APEX2-null mice., Blood, 10.1182/blood-2004-04-1476, 104, 13, 4097-4103, 104(13):4097-4103, 2004.12.
||Tsuchimoto D, Tojo A, Asano S, A Mechanism of Transcriptional Regulation of the CSF-1 Gene by Interferon-gamma., Immunological Investigations, 10.1081/IMM-200038662, 33, 4, 397-405, 33(4):397-405, 2004.12.
||Tominaga Y, Ushijima Y, Tsuchimoto D, Mishima M, Shirakawa M, Hirano S, Sakumi K, Nakabeppu Y., MUTYH prevents OGG1 or APEX1 from inappropriately processing its substrate or reaction product with its C-terminal domain, Nucleic Acids Research, 10.1093/nar/gkh642, 32, 10, 3198-3211, 32(10):3198-3211, 2004.06.
||Miura T, Takahashi M, Horie H, Kurushima H, Tsuchimoto D, Sakumi K, Nakabeppu Y., Galectin-1beta, a natural monomeric form of galectin-1 lacking its six amino-terminal residues promotes axonal regeneration but not cell death, Cell Death and Differentiation, 10.1038/sj.cdd.4401462, 11, 10, 1076-1083, 11(10):1076-1083, 2004.10.
||Nakabeppu Y, Tsuchimoto D, Ichinoe A, Ohno M, Ide Y, Hirano S, Yoshimura D, Tominaga Y, Furuichi M, Sakumi K., Biological significance of the defense mechanisms against oxidative damage in nucleic acids caused by reactive oxygen species: from mitochondria to nuclei, Ann N Y Acad Sci., 10.1196/annals.1293.011, 1011, 101-111, 1011:101-111, 2004.04.
||Ichinoe A, Behmanesh M, Tominaga Y, Ushijima Y, Hirano S, Sakai Y, Tsuchimoto D, Sakumi K, Wake N, Nakabeppu Y., Identification and characterization of two forms of mouse MUTYH proteins encoded by alternatively spliced transcripts, Nucleic Acids Research, 10.1093/nar/gkh214, 32, 2, 477-487, 32(2):477-487, 2004.01.
||Hirano S, Tominaga Y, Ichinoe A, Ushijima Y, Tsuchimoto D, Honda-Ohnishi Y,, Mutator phenotype of MUTYH-null mouse embryonic stem cells., J Biol Chem., 10.1074/jbc.C300316200, 278, 40, 38121-38124, 278(40):38121-38124, 2003.10.
||Tahara K, Tsuchimoto D, Tominaga Y, Asoh S, Ohta S, Kitagawa M, Horie H, Nakabeppu Y, DeltaFosB, but not FosB, induces delayed apoptosis independent of cell
proliferation in the Rat1a embryo cell line., Cell Death Differ., 10.1038/sj.cdd.4401173, 10, 5, 496-507, 10(5):496-507., 2003.05.
||Ide Y, Tsuchimoto D, Tominaga Y, Iwamoto Y, Nakabeppu Y., Characterization of the genomic structure and expression of the mouse Apex2
gene., Genomics, 10.1016/S0888-7543(02)00009-5, 81, 1, 47-57, 81(1):47-57., 2003.01.
||Tsuchimoto D, Sakai Y, Sakumi K, Nishioka K, Sasaki M, Fujiwara T, Nakabeppu Y, Human APE2 protein is mostly localized in the nuclei and to some extent in the
mitochondria, while nuclear APE2 is partly associated with proliferating cell
nuclear antigen., Nucleic Acids Res., 10.1093/nar/29.11.2349, 29, 11, 2349-2360, 29(11):2349-2360, 2001.06.