Regulation of 1-beta-D-arabinofuranosylcytosine 5'-triphosphate accumulation in human leukemia cells by deoxycytidine 5'-triphosphate.
Cell cycle-specific fluctuations in the ability of human leukemic cells to phosphorylate 1-beta-D-arabinofuranosylcytosine (ara-C) to the toxic metabolite 1-beta-D-arabinofuranosylcytosine 5'-triphosphate (ara-CTP) was investigated in whole cells and in cell extracts. Exponentially growing CCRF-CEM cells were fractionated into populations enriched for G1 phase cells and S phase cells by centrifugal elutriation. The accumulation of ara-CTP by S phase-enriched cells was 50% greater than in G1-enriched cells. However, the ability of extracts of S phase-enriched cells to phosphorylate ara-C was twice that of G1 phase-enriched cell extracts. As cells passed from G1 to S phase, this disproportionality was significant. As demonstrated in other cell types, deoxycytidine 5'-triphosphate (dCTP) also potently inhibited ara-C phosphorylation in CCRF-CEM cell extracts (Ki = 5.9 microM). Deoxynucleotide pool levels determined by high pressure liquid chromatography showed a 5 microM dCTP concentration in G1-enriched cells, whereas S phase-enriched cells contained 15 microM dCTP. These findings suggest that the lack of proportionality between the accumulation of ara-CTP in whole cells and the increase of ara-C phosphorylation in extracts during the G1 to S phase transition may be caused by more stringent regulation of ara-C phosphorylation in whole cells by the concomitant increase in cellular dCTP concentrations. Because such regulation is unlikely to be observed in cell extracts, these results indicated that assays of ara-C phosphorylating activity in cell extracts represent upper limits for that function in whole cells. Such determinations may not reflect the regulated nature of the metabolic pathway.
D001085:Arabinofuranosylcytosine Triphosphate / Q000096:biosynthesis*; D001088:Arabinonucleotides / Q000096:biosynthesis*; D002453:Cell Cycle; D002460:Cell Line; D003561:Cytarabine / Q000378:metabolism; D003845:Deoxycytosine Nucleotides / Q000494:pharmacology; D006801:Humans; D007399:Interphase; D007938:Leukemia / Q000378:metabolism*; D009711:Nucleotides / Q000378:metabolism; D010766:Phosphorylation; D013601:T-Lymphocytes / Q000378:metabolism
D001088:Arabinonucleotides; D003845:Deoxycytosine Nucleotides; D009711:Nucleotides; D003561:Cytarabine; D001085:Arabinofuranosylcytosine Triphosphate; C024107:2'-deoxycytidine 5'-triphosphate