title: Age-Related Changes in D-Aspartate Oxidase Promoter Methylation Control Extracellular D-Aspartate Levels and Prevent Precocious Cell Death during Brain Aging.
abstract: The endogenous NMDA receptor (NMDAR) agonist D-aspartate occurs transiently in the mammalian brain because it is abundant during embryonic and perinatal phases before drastically decreasing during adulthood. It is well established that postnatal reduction of cerebral D-aspartate levels is due to the concomitant onset of D-aspartate oxidase (DDO) activity, a flavoenzyme that selectively degrades bicarboxylic D-amino acids. In the present work, we show that d-aspartate content in the mouse brain drastically decreases after birth, whereas Ddo mRNA levels concomitantly increase. Interestingly, postnatal Ddo gene expression is paralleled by progressive demethylation within its putative promoter region. Consistent with an epigenetic control on Ddo expression, treatment with the DNA-demethylating agent, azacitidine, causes increased mRNA levels in embryonic cortical neurons. To indirectly evaluate the effect of a putative persistent Ddo gene hypermethylation in the brain, we used Ddo knock-out mice (Ddo(-/-)), which show constitutively suppressed Ddo expression. In these mice, we found for the first time substantially increased extracellular content of d-aspartate in the brain. In line with detrimental effects produced by NMDAR overstimulation, persistent elevation of D-aspartate levels in Ddo(-/-) brains is associated with appearance of dystrophic microglia, precocious caspase-3 activation, and cell death in cortical pyramidal neurons and dopaminergic neurons of the substantia nigra pars compacta. This evidence, along with the early accumulation of lipufuscin granules in Ddo(-/-) brains, highlights an unexpected importance of Ddo demethylation in preventing neurodegenerative processes produced by nonphysiological extracellular levels of free D-aspartate.
authors.affiliation: Istituto di Endocrinologia ed Oncologia Sperimentale, Consiglio Nazionale delle Ricerche (CNR), 80131 Naples, Italy, Department of Environmental, Biological and Pharmaceutical Sciences and Technologies, Second University of Naples, 81100 Caserta, Italy.
authors.forename: Daniela
authors.initials: D
authors.lastname: Punzo
authors.affiliation: Laboratory of Behavioural Neuroscience, Ceinge Biotecnologie Avanzate, 80145 Naples, Italy, Department of Molecular Medicine and Medical Biotechnology, University of Naples Federico II, 80131 Naples, Italy.
authors.forename: Francesco
authors.identifier: http://orcid.org/0000-0002-3636-7395
authors.initials: F
authors.lastname: Errico
authors.affiliation: Institute of Biomolecular Chemistry, CNR, 80078 Pozzuoli, Italy.
authors.forename: Luigia
authors.initials: L
authors.lastname: Cristino
authors.affiliation: Department of Biotechnology and Life Sciences, University of Insubria, 21100 Varese, Italy, Protein Factory Research Center, Politecnico di Milano and University of Insubria, 20133 Milan, Italy.
authors.forename: Silvia
authors.identifier: http://orcid.org/0000-0002-2338-2561
authors.initials: S
authors.lastname: Sacchi
authors.affiliation: Istituto di Endocrinologia ed Oncologia Sperimentale, Consiglio Nazionale delle Ricerche (CNR), 80131 Naples, Italy, Department of Molecular Medicine and Medical Biotechnology, University of Naples Federico II, 80131 Naples, Italy.
authors.forename: Simona
authors.initials: S
authors.lastname: Keller
authors.affiliation: Department of Experimental Medicine, Section of Pharmacology, Second University of Naples, 80138 Naples, Italy.
authors.forename: Carmela
authors.initials: C
authors.lastname: Belardo
authors.affiliation: Department of Experimental Medicine, Section of Pharmacology, Second University of Naples, 80138 Naples, Italy.
authors.forename: Livio
authors.initials: L
authors.lastname: Luongo
authors.affiliation: Department of Environmental, Biological and Pharmaceutical Sciences and Technologies, Second University of Naples, 81100 Caserta, Italy, Laboratory of Behavioural Neuroscience, Ceinge Biotecnologie Avanzate, 80145 Naples, Italy.
authors.forename: Tommaso
authors.identifier: http://orcid.org/0000-0002-2397-3417
authors.initials: T
authors.lastname: Nuzzo
authors.affiliation: Institute of Biomolecular Chemistry, CNR, 80078 Pozzuoli, Italy.
authors.forename: Roberta
authors.initials: R
authors.lastname: Imperatore
authors.affiliation: Istituto di Endocrinologia ed Oncologia Sperimentale, Consiglio Nazionale delle Ricerche (CNR), 80131 Naples, Italy, Department of Molecular Medicine and Medical Biotechnology, University of Naples Federico II, 80131 Naples, Italy.
authors.forename: Ermanno
authors.initials: E
authors.lastname: Florio
authors.affiliation: Department of Experimental Medicine, Section of Pharmacology, Second University of Naples, 80138 Naples, Italy.
authors.forename: Vito
authors.initials: V
authors.lastname: De Novellis
authors.affiliation: Istituto di Endocrinologia ed Oncologia Sperimentale, Consiglio Nazionale delle Ricerche (CNR), 80131 Naples, Italy, Department of Molecular Medicine and Medical Biotechnology, University of Naples Federico II, 80131 Naples, Italy.
authors.forename: Ornella
authors.identifier: http://orcid.org/0000-0002-8355-9509
authors.initials: O
authors.lastname: Affinito
authors.affiliation: Department of Biology, Unit of Cell and Developmental Biology, University of Pisa, 56126 Pisa, Italy, and.
authors.forename: Sara
authors.identifier: http://orcid.org/0000-0003-2741-269X
authors.initials: S
authors.lastname: Migliarini
authors.affiliation: Department of Biology, Unit of Cell and Developmental Biology, University of Pisa, 56126 Pisa, Italy, and.
authors.forename: Giacomo
authors.identifier: http://orcid.org/0000-0002-0485-8544
authors.initials: G
authors.lastname: Maddaloni
authors.affiliation: Istituto di Endocrinologia ed Oncologia Sperimentale, Consiglio Nazionale delle Ricerche (CNR), 80131 Naples, Italy.
authors.forename: Maria Josè
authors.initials: MJ
authors.lastname: Sisalli
authors.affiliation: Department of Biology, Unit of Cell and Developmental Biology, University of Pisa, 56126 Pisa, Italy, and Center for Neuroscience and Cognitive Systems, Istituto Italiano di Tecnologia, 38068 Rovereto, Italy.
authors.forename: Massimo
authors.identifier: http://orcid.org/0000-0002-0844-8139
authors.initials: M
authors.lastname: Pasqualetti
authors.affiliation: Department of Biotechnology and Life Sciences, University of Insubria, 21100 Varese, Italy, Protein Factory Research Center, Politecnico di Milano and University of Insubria, 20133 Milan, Italy.
authors.forename: Loredano
authors.identifier: http://orcid.org/0000-0003-1733-7243
authors.initials: L
authors.lastname: Pollegioni
authors.affiliation: Department of Experimental Medicine, Section of Pharmacology, Second University of Naples, 80138 Naples, Italy.
authors.forename: Sabatino
authors.initials: S
authors.lastname: Maione
authors.affiliation: Istituto di Endocrinologia ed Oncologia Sperimentale, Consiglio Nazionale delle Ricerche (CNR), 80131 Naples, Italy, Department of Molecular Medicine and Medical Biotechnology, University of Naples Federico II, 80131 Naples, Italy, chiariot@unina.it usiello@ceinge.unina.it.
authors.forename: Lorenzo
authors.initials: L
authors.lastname: Chiariotti
authors.affiliation: Department of Environmental, Biological and Pharmaceutical Sciences and Technologies, Second University of Naples, 81100 Caserta, Italy, Laboratory of Behavioural Neuroscience, Ceinge Biotecnologie Avanzate, 80145 Naples, Italy, chiariot@unina.it usiello@ceinge.unina.it.
authors.forename: Alessandro
authors.initials: A
authors.lastname: Usiello
mesh_terms: D000367:Age Factors; D000375:Aging; D000818:Animals; D000831:Animals, Newborn; D001374:Azacitidine / Q000031:analogs & derivatives / Q000494:pharmacology; D001921:Brain / Q000166:cytology / Q000378:metabolism*; D016923:Cell Death / Q000235:genetics; D050804:D-Aspartate Oxidase / Q000235:genetics / Q000378:metabolism*; D026603:D-Aspartic Acid / Q000378:metabolism*; D000077209:Decitabine; D004622:Embryo, Mammalian; D004791:Enzyme Inhibitors / Q000494:pharmacology; D008297:Male; D008745:Methylation; D051379:Mice; D008810:Mice, Inbred C57BL; D008822:Mice, Transgenic; D009474:Neurons / Q000187:drug effects / Q000502:physiology*; D011401:Promoter Regions, Genetic / Q000235:genetics*; D012333:RNA, Messenger / Q000378:metabolism; D016194:Receptors, N-Methyl-D-Aspartate / Q000378:metabolism
chemical_list: D004791:Enzyme Inhibitors; D012333:RNA, Messenger; D016194:Receptors, N-Methyl-D-Aspartate; D026603:D-Aspartic Acid; D000077209:Decitabine; D050804:D-Aspartate Oxidase; D001374:Azacitidine
keywords: DNA methylation; NMDA receptor; aging; d-amino acids; neurodegeneration