Bianchi V, Pontis E, Reichard P
J Biol Chem. 1986 Dec 5;261(34):16037-42.
Hydroxyurea inactivates ribonucleotide reductase from mammalian cells and thereby depletes them of the deoxynucleoside triphosphates required for DNA replication. In cultures of exponentially growing 3T6 cells, with 60-70% of the cells in S-phase, 3 mM hydroxyurea rapidly stopped ribonucleotide reduction and DNA synthesis (incorporation of labeled thymidine). The pool of deoxyadenosine triphosphate (dATP) decreased in size primarily, but also the pools of the triphosphates of deoxyguanosine and deoxycytidine (dCTP) were depleted. Paradoxically, the pool of thymidine triphosphate increased. After addition of hydroxyurea this pool was fed by a net influx and phosphorylation of deoxyuridine from the medium and by deamination of intracellular dCTP. An influx of deoxycytidine from the medium contributed to the maintenance of intracellular dCTP. 10 min after addition of hydroxyurea, DNA synthesis appeared to be completely blocked even though the dATP pool was only moderately decreased. As possible explanations for this discrepancy, we discuss compartmentation of pools and/or vulnerability of newly formed DNA strands to nuclease action and pyrophosphorolysis.
羟基脲可使哺乳动物细胞中的核糖核苷酸还原酶失活，从而耗尽细胞进行DNA复制所需的脱氧核苷三磷酸。在指数生长的3T6细胞培养物中，60 - 70%的细胞处于S期，3 mM羟基脲能迅速停止核糖核苷酸还原和DNA合成（标记胸腺嘧啶核苷的掺入）。脱氧腺苷三磷酸（dATP）池的大小主要减小，脱氧鸟苷三磷酸和脱氧胞苷三磷酸（dCTP）池也被耗尽。矛盾的是，胸腺嘧啶核苷三磷酸池增加。添加羟基脲后，该池通过培养基中脱氧尿苷的净流入和磷酸化以及细胞内dCTP的脱氨基作用得到补充。培养基中脱氧胞苷的流入有助于维持细胞内dCTP。添加羟基脲10分钟后，尽管dATP池仅适度减少，但DNA合成似乎完全被阻断。作为这种差异的可能解释，我们讨论了池的区室化和/或新形成的DNA链对核酸酶作用和焦磷酸解的脆弱性。
