title: The pivotal role of the mitochondrial amidoxime reducing component 2 in protecting human cells against apoptotic effects of the base analog N6-hydroxylaminopurine.
abstract: N-Hydroxylated nucleobases and nucleosides as N-hydroxylaminopurine (HAP) or N-hydroxyadenosine (HAPR) may be generated endogenously in the course of cell metabolism by cytochrome P450, by oxidative stress or by a deviating nucleotide biosynthesis. These compounds have shown to be toxic and mutagenic for procaryotic and eucaryotic cells. For DNA replication fidelity it is therefore of great importance that organisms exhibit effective mechanisms to remove such non-canonical base analogs from DNA precursor pools. In vitro, the molybdoenzymes mitochondrial amidoxime reducing component 1 and 2 (mARC1 and mARC2) have shown to be capable of reducing N-hydroxylated base analogs and nucleoside analogs to the corresponding canonical nucleobases and nucleosides upon reconstitution with the electron transport proteins cytochrome b5 and NADH-cytochrome b5 reductase. By RNAi-mediated down-regulation of mARC in human cell lines the mARC-dependent N-reductive detoxication of HAP in cell metabolism could be demonstrated. For HAPR, on the other hand, the reduction to adenosine seems to be of less significance in the detoxication pathway of human cells as HAPR is primarily metabolized to inosine by direct dehydroxylamination catalyzed by adenosine deaminase. Furthermore, the effect of mARC knockdown on sensitivity of human cells to HAP was examined by flow cytometric quantification of apoptotic cell death and detection of poly (ADP-ribose) polymerase (PARP) cleavage. mARC2 was shown to protect HeLa cells against the apoptotic effects of the base analog, whereas the involvement of mARC1 in reductive detoxication of HAP does not seem to be pivotal.
authors.affiliation: From the Department of Pharmaceutical and Medicinal Chemistry, Pharmaceutical Institute, Christian-Albrechts-University of Kiel, 24118 Kiel, Germany.
authors.forename: Birte
authors.initials: B
authors.lastname: Plitzko
authors.affiliation: From the Department of Pharmaceutical and Medicinal Chemistry, Pharmaceutical Institute, Christian-Albrechts-University of Kiel, 24118 Kiel, Germany.
authors.forename: Antje
authors.initials: A
authors.lastname: Havemeyer
authors.affiliation: From the Department of Pharmaceutical and Medicinal Chemistry, Pharmaceutical Institute, Christian-Albrechts-University of Kiel, 24118 Kiel, Germany.
authors.forename: Thomas
authors.initials: T
authors.lastname: Kunze
authors.affiliation: From the Department of Pharmaceutical and Medicinal Chemistry, Pharmaceutical Institute, Christian-Albrechts-University of Kiel, 24118 Kiel, Germany bclement@pharmazie.uni-kiel.de.
authors.forename: Bernd
authors.initials: B
authors.lastname: Clement
mesh_terms: D000225:Adenine / Q000031:analogs & derivatives* / Q000378:metabolism; D000241:Adenosine / Q000031:analogs & derivatives* / Q000378:metabolism; D017209:Apoptosis / Q000235:genetics; D042966:Cytochrome-B(5) Reductase / Q000235:genetics / Q000378:metabolism; D015786:Cytochromes b5 / Q000235:genetics / Q000378:metabolism; D015870:Gene Expression; D057809:HEK293 Cells; D006367:HeLa Cells; D006801:Humans; D008565:Membrane Proteins / Q000037:antagonists & inhibitors / Q000235:genetics / Q000378:metabolism*; D050216:Metabolic Detoxication, Phase I; D008928:Mitochondria / Q000201:enzymology; D024101:Mitochondrial Proteins / Q000037:antagonists & inhibitors / Q000235:genetics / Q000378:metabolism*; D010088:Oxidoreductases / Q000037:antagonists & inhibitors / Q000235:genetics / Q000378:metabolism*; D011065:Poly(ADP-ribose) Polymerases / Q000235:genetics / Q000378:metabolism; D059748:Proteolysis; D034741:RNA, Small Interfering / Q000235:genetics / Q000378:metabolism
chemical_list: D008565:Membrane Proteins; D024101:Mitochondrial Proteins; D034741:RNA, Small Interfering; C032610:6-N-hydroxylaminopurine; D015786:Cytochromes b5; C570001:MTARC2 protein, human; D010088:Oxidoreductases; C534493:amidoxime reducing component, human; C583417:CYB5R3 protein, human; D042966:Cytochrome-B(5) Reductase; D011065:Poly(ADP-ribose) Polymerases; D000225:Adenine; D000241:Adenosine; C009890:6-hydroxyadenosine
keywords: MOSC; N-reduction; N6-hydroxyadenosine; N6-hydroxylaminopurine; RNA interference (RNAi); apoptosis; cell metabolism; flow cytometry; mARC; nucleoside/nucleotide analogue