FUNCTIONAL COMPARTMENTATION OF DCTP POOLS - PREFERENTIAL UTILIZATION OF SALVAGED DEOXYCYTIDINE FOR DNA-REPAIR IN HUMAN LYMPHOBLASTS

The utilization of dCTP derived from de nora synthesis through ribonuc leotide reductase in exponentially growing CCRF-CEM cells was compared with the metabolic fate of dCTP produced by the salvage pathway, Exog enous dCyd was not effectively incorporated into replicating DNA; inst ead, dCTP derived from 1 ribonucleotide reductase (labeled by [5-H-3]C yd) was the main precursor for that purpose, apparently because of fun ctional compartmentation of the dCTP pool in these cells, Studies of t he metabolic route of incorporation of exogenous [5-H-3]dCyd into DNA of growing CCRF-CEM cells demonstrated that it was mainly incorporated through the DNA repair pathway. Incorporation of [5-H-3]dCyd into DNA of synchronized cell populations was maximal in G(1) cells, whereas [ H-3]dThd incorporation occurred predominantly in S phase cells, When c ellular DNA was density labeled by incubation with BrdUrd, repaired DN A, which was less dense than replicated DNA, was preferentially labele d by [5-H-3]dCyd. In contrast, replicated DNA was labeled by both [H-3 ]dThd and [5-H-3]Cyd, The DNA-damaging agents methylmethanesulfonate, ultraviolet irradiation, and gamma-irradiation inhibited [H-3]dThd inc orporation, whereas they stimulated the accumulation of [5-H-3]dCyd in DNA. Based on these results, we propose that the dCTP pool is functio nally compartmentalized in growing CCRF-CEM cells, dCTP derived from t he salvage pathway is utilized predominantly for DNA repair, whereas t he de nova pathway supplies dCTP far DNA replication.