Functional compartmentation of dCTP pools. Preferential utilization of salvaged deoxycytidine for DNA repair in human lymphoblasts.
The utilization of dCTP derived from de novo synthesis through ribonucleotide reductase in exponentially growing CCRF-CEM cells was compared with the metabolic fate of dCTP produced by the salvage pathway. Exogenous dCyd was not effectively incorporated into replicating DNA; instead, dCTP derived from ribonucleotide reductase (labeled by [5-3H]Cyd) was the main precursor for that purpose, apparently because of functional compartmentation of the dCTP pool in these cells. Studies of the metabolic route of incorporation of exogenous [5-3H]dCyd into DNA of growing CCRF-CEM cells demonstrated that it was mainly incorporated through the DNA repair pathway. Incorporation of [5-3H]dCyd into DNA of synchronized cell populations was maximal in G1 cells, whereas [3H]dThd incorporation occurred predominantly in S phase cells. When cellular DNA was density labeled by incubation with BrdUrd, repaired DNA, which was less dense than replicated DNA, was preferentially labeled by [5-3H]dCyd. In contrast, replicated DNA was labeled by both [3H]dThd and [5-3H]Cyd. The DNA-damaging agents methylmethanesulfonate, ultraviolet irradiation, and gamma-irradiation inhibited [3H]dThd incorporation, whereas they stimulated the accumulation of [5-3H]dCyd in DNA. Based on these results, we propose that the dCTP pool is functionally compartmentalized in growing CCRF-CEM cells. dCTP derived from the salvage pathway is utilized predominantly for DNA repair, whereas the de novo pathway supplies dCTP for DNA replication.
Xu
Y Z
YZ
Department of Clinical Investigation, University of Texas M. D. Anderson Cancer Center, Houston 77030.
Huang
P
P
Plunkett
W
W
D001973:Bromodeoxyuridine; D002451:Cell Compartmentation; D002478:Cells, Cultured; D003562:Cytidine / Q000378:metabolism / Q000494:pharmacology; D004247:DNA / Q000187:drug effects; D004249:DNA Damage; D004260:DNA Repair; D003845:Deoxycytosine Nucleotides / Q000378:metabolism*; D006801:Humans; D008214:Lymphocytes; D013936:Thymidine / Q000378:metabolism
D003845:Deoxycytosine Nucleotides; C024107:2'-deoxycytidine 5'-triphosphate; D003562:Cytidine; D004247:DNA; D001973:Bromodeoxyuridine; D013936:Thymidine