title: Reduction of 8-oxodGTP in the nucleotide pool by hMTH1 leads to reduction in mutations in the human lymphoblastoid cell line TK6 exposed to UVA.
abstract: UVA has been suggested to play an important role in UV-induced mutagenesis. The mechanisms by which UVA induces mutations are still a matter of debate. Our aim was to investigate the protective capacity of hMTH1, a nucleotide pool sanitization enzyme with 8-oxodGTPase activity. Human B lymphoblastoid cells were stably transfected with shRNA directed against hMTH1. Clonogenic survival, mutations, intracellular and extracellular levels of 8-oxodG (8-oxo-7, 8-dihydro-2'-deoxyguanosine) and dG in the nucleotide pool of UVA-irradiated transfected and non-transfected cells were investigated. Mutations were determined in the thymidine kinase locus. Intracellular 8-oxodG and dG were measured using a modified ELISA and HPLC, respectively, after extraction of the nucleotide pool and conversion of nucleotides to their corresponding nucleosides. 8-oxodG in the medium was measured using ELISA. UVA-induced mutations were significantly higher while the survival was slightly lower in transfected compared to non-transfected cells. The increased mutation rate in transfected cells at increased exposure correlated with enhanced levels of 8-oxodG in the nucleotide pool, and a somewhat reduced level of 8-oxodG in the medium. The results indicate that the nucleotide pool is a significant target for UVA-induced mutations and implicates that hMTH1 plays an important role in protecting cells from UVA-induced oxidative stress.
authors.affiliation: Centre for Radiation Protection Research, Department of Genetics, Microbiology and Toxicology, Stockholm University, SE-10691 Stockholm, Sweden.
authors.forename: Asal
authors.initials: A
authors.lastname: Fotouhi
authors.forename: Sara
authors.initials: S
authors.lastname: Skiöld
authors.forename: Sara
authors.initials: S
authors.lastname: Shakeri-Manesh
authors.forename: Siv
authors.initials: S
authors.lastname: Osterman-Golkar
authors.forename: Andrzej
authors.initials: A
authors.lastname: Wojcik
authors.forename: Dag
authors.initials: D
authors.lastname: Jenssen
authors.forename: Mats
authors.initials: M
authors.lastname: Harms-Ringdahl
authors.forename: Siamak
authors.initials: S
authors.lastname: Haghdoost
mesh_terms: D000080242:8-Hydroxy-2'-Deoxyguanosine; D001402:B-Lymphocytes / Q000378:metabolism*; D002460:Cell Line; D045643:DNA Repair Enzymes / Q000494:pharmacology* / Q000502:physiology*; D003848:Deoxyguanine Nucleotides / Q000378:metabolism*; D003849:Deoxyguanosine / Q000031:analogs & derivatives* / Q000378:metabolism; D006801:Humans; D009154:Mutation / Q000187:drug effects*; D018384:Oxidative Stress; D010744:Phosphoric Monoester Hydrolases / Q000494:pharmacology* / Q000502:physiology*; D014162:Transfection; D014466:Ultraviolet Rays / Q000009:adverse effects*
chemical_list: D003848:Deoxyguanine Nucleotides; C078206:8-oxodeoxyguanosine triphosphate; D000080242:8-Hydroxy-2'-Deoxyguanosine; D010744:Phosphoric Monoester Hydrolases; C078207:8-oxodGTPase; D045643:DNA Repair Enzymes; D003849:Deoxyguanosine