Mimosine arrests proliferating human cells before onset of DNA replication in a dose-dependent manner

被引:156
作者
Krude, T
机构
[1] Univ Cambridge, Wellcome CRC Inst, Cambridge CB2 1QR, England
[2] Univ Cambridge, Dept Zool, Cambridge CB2 1TN, England
关键词
D O I
10.1006/excr.1998.4342
中图分类号
R73 [肿瘤学];
学科分类号
100214 ;
摘要
The synchronization effects of the plant amino acid mimosine on proliferating higher eukaryotic cells are still controversial. Here, I show that 0.5 mM mimosine can induce a cell cycle arrest of human somatic cells in late G1 phase, before establishment of active DNA replication forks. The DNA content of nuclei isolated from mimosine-treated cells was determined by flow cytometry. The presence or absence of DNA replication forks in these isolated nuclei was then detected by DNA replication run-on assays in vitro. Treatment of asynchronously proliferating HeLa or EJ30 cells for 24 h with 0.5 mM mimosine resulted in a population synchronized in late G1 phase. S phase entry was inhibited by 0.5 mM mimosine in cells released from a block in mitosis or from quiescence. When added to early S phase cells, 0.5 mM mimosine did not prevent S phase transit, but delayed progression through late stages of S phase after a lag of 4 h, eventually resulting in a G1 phase population by preventing entry into the subsequent S phase. In contrast, lower concentrations of mimosine (0.1-0.2 mM) failed to prevent S phase entry, resulting in cells containing active DNA replication foci. The G1 phase arrest by 0.5 mM mimosine was reversible upon mimosine withdrawal. This synchronization protocol using 0.5 mM mimosine can be exploited for studying the initiation of human DNA replication in vitro. (C) 1999 Academic Press.
引用
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页码:148 / 159
页数:12
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