Regulation of 1-beta-D-arabinofuranosylcytosine 5'-triphosphate accumulation in human leukemia cells by deoxycytidine 5'-triphosphate.
Regulation of 1-beta-D-arabinofuranosylcytosine 5'-triphosphate accumulation in human leukemia cells by deoxycytidine 5'-triphosphate.
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发表时间：
1986-03
期刊：
影响因子：
11.2
通讯作者：
J. Liliemark;W. Plunkett
中科院分区：
文献类型：
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作者：
J. Liliemark;W. Plunkett
研究了人白血病细胞对毒性代谢物1-β-D-阿拉伯呋喃胞嘧啶(Ara-C)与毒性代谢物1-β-D-阿拉伯呋喃胞苷5‘-三磷酸(Ara-CTP)磷酸化能力的细胞周期特异性波动。通过离心洗脱，将指数生长期的CCRF-CEM细胞分为G1期细胞和S期细胞。S期细胞对Ara-CTP的积累量比G1期细胞高50%。然而，S期细胞提取物对Ara-C的磷酸化能力是G1期细胞提取物的两倍。随着细胞从G1期进入S期，这种不成比例现象非常明显。与其他类型的细胞一样，脱氧胞苷5‘-三磷酸(DCTP)也能有效地抑制CCRF-CEM细胞提取物中Ara-C的磷酸化(Ki=5.9微米)。高压液相色谱检测脱氧核酸池水平显示，G1期浓缩细胞中dCTP含量为5微米，而S时相浓缩细胞中dCTP含量为15微米。这些结果表明，在G1-S相变期间，细胞内Ara-CTP的积累与提取物中Ara-C磷酸化的增加之间缺乏正比，这可能是由于细胞内dCTP浓度的增加对整个细胞中Ara-C的磷酸化进行了更严格的调控。由于这种调节不太可能在细胞提取物中观察到，这些结果表明，细胞提取物中Ara-C磷酸化活性的测定代表了整个细胞中这一功能的上限。这种测定可能不能反映代谢途径的调节性质。
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.