Isotope-dilution analysis of the effects of deoxyguanosine and deoxyadenosine on the incorporation of thymidine and deoxycytidine by hydroxyurea-treated thymus cells.
Isotope-dilution analysis of the effects of deoxyguanosine and deoxyadenosine on the incorporation of thymidine and deoxycytidine by hydroxyurea-treated thymus cells.
复制标题
同位素稀释分析脱氧鸟苷和脱氧腺苷对羟基脲处理的胸腺细胞掺入胸苷和脱氧胞苷的影响。
DOI：
10.1042/bj1900721
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发表时间：
1980
期刊：
影响因子：
--
通讯作者：
D. Forsdyke
中科院分区：
文献类型：
--
作者：
F. Scott;D. Forsdyke
据推测，复制DNA合成所需的dGTP和dATP可以通过补救途径或从头生物合成的方式形成。通过向大鼠胸腺细胞培养物中加入羟基脲以抑制核糖核苷二磷酸还原酶（从头途径的关键酶）来检查这一点。低浓度的羟基脲（100- 500 μ m）对[Me-（3）H]胸苷和脱氧[5-（3）H]胞苷掺入的大部分抑制作用可被补救途径的底物（400 μ m-脱氧鸟苷和较小程度的200 μ m-脱氧腺苷）阻止。然而，同位素稀释研究表明，嘌呤脱氧核糖核苷通过减少嘧啶脱氧核糖核苷酸竞争池来防止抑制。有证据表明，在胸苷竞争池（可能是dTTP）的羟基脲诱导的增加被阻止在同等程度上由脱氧鸟苷和胸苷酸合成酶，脱氧-5-氟尿苷的抑制剂。这些化合物对羟基脲剂量-反应曲线和胸苷同位素稀释图的影响几乎相同。有证据表明，外源性嘌呤脱氧核糖核苷不能阻止由羟基脲胸腺细胞DNA合成的抑制。这可能意味着，就嘌呤脱氧核糖核苷酸的代谢而言，核糖核苷二磷酸还原酶与DNA聚合酶在多酶复合物中紧密偶联。该复合物不允许外源性代谢中间体进入"从头“途径，但仍会受到这些中间体的调节作用。因此，dGTP和dATP形成的外源性嘌呤脱氧核糖核苷的补救途径可能会耗尽嘧啶脱氧核糖核苷酸竞争池抑制相对羟基脲不敏感的活性的核糖核苷二磷酸还原酶。
It is presumed that the dGTP and dATP needed for replicative DNA synthesis can be formed by way of either ;salvage' pathways or biosynthesis de novo. This was examined by adding hydroxyurea to cultures of rat thymus cells to inhibit ribonucleoside diphosphate reductase, a key enzyme of the ;de novo' pathway. Most of the inhibition of the incorporation of [Me-(3)H]thymidine and deoxy[5-(3)H]cytidine by low concentrations of hydroxyurea (100-500mum) was prevented by substrates of the salvage pathway (400mum-deoxyguanosine and, to a lesser extent, 200mum-deoxyadenosine). However, isotope-dilution studies indicated that the purine deoxyribonucleosides prevented inhibition by decreasing pyrimidine deoxyribonucleotide competitor pools. Evidence was obtained that a hydroxyurea-induced increase in the thymidine-competitor pool (probably dTTP) was prevented to an equal extent by deoxyguanosine and by the inhibitor of thymidylate synthase, deoxy-5-fluorouridine. These compounds had almost identical effects on hydroxyurea dose-response curves and on thymidine isotope-dilution plots. The evidence suggests that exogenous purine deoxyribonucleosides cannot prevent the inhibition by hydroxyurea of thymus-cell DNA synthesis. This could mean that, with respect to the metabolism of purine deoxyribonucleotides, ribonucleoside diphosphate reductase is tightly coupled to DNA polymerase in a multienzyme complex. The complex would not permit entry of exogenous metabolic intermediates into the ;de novo' pathway, but would still be subject to the regulatory effects of these intermediates. Thus dGTP and dATP formed from exogenous purine deoxyribonucleosides by salvage pathways might deplete pyrimidine deoxyribonucleotide competitor pools by inhibiting relatively hydroxyurea-insensitive activities of ribonucleoside diphosphate reductase.