Guanine for DNA synthesis. A compulsory route through ribonucleotide reductase.
Two alternative pathways for the synthesis of dGTP and its incorporation into DNA were studied: guanine (Gua)----GMP----GDP----dGDP----dGTP----DNA and dG----dGMP----dGDP----dGTP----DNA. To determine the contribution of each pathway to DNA synthesis independently of each other, [14C]Gua and [3H]dG tracer experiments were performed in a double-mutant S-49 mouse T-lymphoma cell line, dGuo-L, with purine nucleoside phosphorylase (EC 2.4.2.1)-deficiency and dGTP-feedback-resistant ribonucleotide reductase (RR, EC 1.17.4.1). In this cell line, dGTP pools can be selectively elevated by exogenous dG without affect RR and DNA synthesis. Although [3H]dG, but not [14C]Gua (up to 200 microM), readily expanded the cellular dGTP pool in a dose-dependent fashion in asynchronous cells, only a small fraction of the Gua flux into DNA was derived from [3H]dG, with the major fraction coming from [14C]Gua. H.p.l.c. analysis of G1- and partially enriched S-phase cells revealed that [3H]dGTP only accumulates in G1- but not in S-phase cells because of a rapid turnover of the dGTP pool during DNA synthesis. These results fail to provide evidence for cellular dGTP compartmentation and suggest that the pathway dG----dGMP----dGDP----dGTP alone has insufficient capacity to maintain DNA synthesis.
Duan
D S
DS
Department of Pharmaceutical Chemistry, School of Pharmacy, University of California, San Francisco 94143.
Sadee
W
W
D000818:Animals; D002250:Carbon Radioisotopes; D004247:DNA / Q000096:biosynthesis*; D003848:Deoxyguanine Nucleotides / Q000378:metabolism; D003849:Deoxyguanosine / Q000378:metabolism; D006147:Guanine / Q000378:metabolism*; D006160:Guanosine Triphosphate / Q000378:metabolism; D051379:Mice; D012264:Ribonucleotide Reductases / Q000378:metabolism*; D014316:Tritium; D014407:Tumor Cells, Cultured / Q000378:metabolism
D002250:Carbon Radioisotopes; D003848:Deoxyguanine Nucleotides; D014316:Tritium; D006147:Guanine; D006160:Guanosine Triphosphate; C029603:deoxyguanosine triphosphate; D004247:DNA; D012264:Ribonucleotide Reductases; D003849:Deoxyguanosine