title: Deoxyribonucleotide metabolism in cycling and resting human fibroblasts with a missense mutation in p53R2, a subunit of ribonucleotide reductase.
abstract: Ribonucleotide reduction provides deoxynucleotides for nuclear and mitochondrial (mt) DNA replication and DNA repair. In cycling mammalian cells the reaction is catalyzed by two proteins, R1 and R2. A third protein, p53R2, with the same function as R2, occurs in minute amounts. In quiescent cells, p53R2 replaces the absent R2. In humans, genetic inactivation of p53R2 causes early death with mtDNA depletion, especially in muscle. We found that cycling fibroblasts from a patient with a lethal mutation in p53R2 contained a normal amount of mtDNA and showed normal growth, ribonucleotide reduction, and deoxynucleoside triphosphate (dNTP) pools. However, when made quiescent by prolonged serum starvation the mutant cells strongly down-regulated ribonucleotide reduction, decreased their dCTP and dGTP pools, and virtually abolished the catabolism of dCTP in substrate cycles. mtDNA was not affected. Also, nuclear DNA synthesis and the cell cycle-regulated enzymes R2 and thymidine kinase 1 decreased strongly, but the mutant cell populations retained unexpectedly larger amounts of the two enzymes than the controls. This difference was probably due to their slightly larger fraction of S phase cells and therefore not induced by the absence of p53R2 activity. We conclude that loss of p53R2 affects ribonucleotide reduction only in resting cells and leads to a decrease of dNTP catabolism by substrate cycles that counterweigh the loss of anabolic activity. We speculate that this compensatory mechanism suffices to maintain mtDNA in fibroblasts but not in muscle cells with a larger content of mtDNA necessary for their high energy requirements.
authors.affiliation: Department of Biology, University of Padova, Padova, Italy.
authors.forename: Giovanna
authors.initials: G
authors.lastname: Pontarin
authors.forename: Paola
authors.initials: P
authors.lastname: Ferraro
authors.forename: Chiara
authors.initials: C
authors.lastname: Rampazzo
authors.forename: Gittan
authors.initials: G
authors.lastname: Kollberg
authors.forename: Elisabeth
authors.initials: E
authors.lastname: Holme
authors.forename: Peter
authors.initials: P
authors.lastname: Reichard
authors.forename: Vera
authors.initials: V
authors.lastname: Bianchi
mesh_terms: D002453:Cell Cycle / Q000502:physiology*; D018797:Cell Cycle Proteins / Q000235:genetics / Q000378:metabolism*; D002478:Cells, Cultured; D004260:DNA Repair / Q000502:physiology; D004261:DNA Replication / Q000502:physiology; D004272:DNA, Mitochondrial / Q000235:genetics / Q000378:metabolism*; D003854:Deoxyribonucleotides / Q000235:genetics / Q000378:metabolism*; D005347:Fibroblasts / Q000166:cytology / Q000201:enzymology*; D006801:Humans; D020125:Mutation, Missense; D010084:Oxidation-Reduction; D012264:Ribonucleotide Reductases / Q000235:genetics / Q000378:metabolism*; D013937:Thymidine Kinase / Q000235:genetics / Q000378:metabolism
chemical_list: D018797:Cell Cycle Proteins; D004272:DNA, Mitochondrial; D003854:Deoxyribonucleotides; C405947:RRM2B protein, human; D012264:Ribonucleotide Reductases; D013937:Thymidine Kinase; C092717:thymidine kinase 1