title: Checkpoint deficient rad53-11 yeast cannot accumulate dNTPs in response to DNA damage.
abstract: Deoxyribonucleotide pools are maintained at levels that support efficient and yet accurate DNA replication and repair. Rad53 is part of a protein kinase regulatory cascade that, conceptually, promotes dNTP accumulation in four ways: (1) it activates the transcription of ribonucleotide reductase subunits by inhibiting the Crt1 repressor; (2) it plays a role in relocalization of ribonucleotide reductase subunits RNR2 and RNR4 from nucleus to cytoplasm; (3) it antagonizes the action of Sml1, a protein that binds and inhibits ribonucleotide reductase; and (4) it blocks cell-cycle progression in response to DNA damage, thus preventing dNTP consumption through replication forks. Although several lines of evidence support the above modes of Rad53 action, an effect of a rad53 mutation on dNTP levels has not been directly demonstrated. In fact, in a previous study, a rad53-11 mutation did not result in lower dNTP levels in asynchronous cells or in synchronized cells that entered the S-phase in the presence of the RNR inhibitor hydroxyurea. These anomalies prompted us to investigate whether the rad53-11 mutation affected dNTP levels in cells exposed to a DNA-damaging dose of ethylmethyl sulfonate (EMS). Although dNTP levels increased by 2- to 3-fold in EMS treated wild-type cells, rad53-11 cells showed no such change. Thus, the results indicate that Rad53 checkpoint function is not required for dNTP pool maintenance in normally growing cells, but is required for dNTP pool expansion in cells exposed to DNA-damaging agents.
authors.affiliation: Izmir Institute of Technology, Department of Molecular Biology and Genetics, 35430 Urla, Izmir, Turkey. ahmetkoc@iyte.edu.tr
authors.forename: Ahmet
authors.initials: A
authors.lastname: Koc
authors.forename: Gary F
authors.initials: GF
authors.lastname: Merrill
mesh_terms: D002453:Cell Cycle / Q000502:physiology*; D018797:Cell Cycle Proteins / Q000378:metabolism; D064447:Checkpoint Kinase 2; D004249:DNA Damage / Q000502:physiology*; D004260:DNA Repair / Q000502:physiology*; D003854:Deoxyribonucleotides / Q000378:metabolism*; D018816:Genes, cdc / Q000502:physiology; D017346:Protein Serine-Threonine Kinases / Q000378:metabolism*; D012441:Saccharomyces cerevisiae / Q000166:cytology* / Q000235:genetics*
chemical_list: D018797:Cell Cycle Proteins; D003854:Deoxyribonucleotides; D064447:Checkpoint Kinase 2; D017346:Protein Serine-Threonine Kinases