Evidence for activities inhibiting in trans initiation of DNA replication in extract prepared from irradiated cells.
Evidence for activities inhibiting in trans initiation of DNA replication in extract prepared from irradiated cells.
复制标题
从受辐射细胞制备的提取物中抑制 DNA 复制反式起始的活性证据。
DOI：
10.2307/3579062
复制
发表时间：
1996
影响因子：
3.4
通讯作者：
G. Iliakis
中科院分区：
文献类型：
--
作者：
Y. Wang;M. Huq;G. Iliakis
我们先前已经表明，当使用辐射细胞的提取物时，含有猿猴病毒40（SV 40）复制起点的质粒的体外复制减少（Wang等人，Radiat. Res. 142，169-175，1995）。我们提出，观察到的总体复制活性的降低是由于起始事件效率的降低，并且它是由电离辐射诱导或激活抑制DNA反式复制的因子引起的。我们扩展了这些研究，并提供了证据表明，从辐射细胞制备的提取物的复制活性降低不是非特异性的结果。蛋白质失活或对SV 40大肿瘤抗原（TAg）的需求增加，后者是体外DNA复制所需的唯一非细胞蛋白质。混合实验表明，在辐照细胞提取物中存在显性抑制活性，可有效阻止使用未辐照细胞提取物组装的反应中的复制。抑制活性是一种稳定的、不可透析的分子。动力学研究表明，抑制活性影响DNA复制的起始步骤，并且至少部分地通过修饰SV 40 ori DNA复制的关键起始蛋白TAg而起作用。很可能相同的抑制活性通过修饰TAg的细胞同源物来调节细胞DNA复制。这种抑制活性的纯化和表征将有助于我们理解电离辐射暴露后细胞调节DNA复制的机制，并将定义在S期操作的检查点。在酵母中，已经报道了S期检查点不同于G1和G2期检查点的遗传证据。
We have previously shown that replication in vitro of plasmids containing the Simian virus 40 (SV40) origin of replication is reduced when an extract of irradiated cells is used (Wang et al., Radiat. Res. 142, 169-175, 1995). We proposed that the observed reduction in the overall replication activity is due to a reduction in the efficiency of initiation events, and that it is caused by the induction or activation by ionizing radiation of a factor(s) that inhibits DNA replication in trans. Here, we extend these studies and provide evidence that the reduced replication activity of an extract prepared from irradiated cells is not the result of a nonspecific inactivation of proteins or of an increase in the requirement for SV40 large tumor antigen (TAg), the only noncellular protein required for in vitro DNA replication. Mixing experiments demonstrate the presence of a dominant inhibitory activity(ies) in the extract of irradiated cells that efficiently stalls replication in reactions assembled using extract of nonirradiated cells. The inhibitory activity is a stable, nondialyzable molecule. Studies of kinetics suggest that the inhibitory activity(ies) affects the initiation steps of DNA replication and acts, at least partly, by modifying TAg, the key initiation protein of SV40 ori DNA replication. It is likely that the same inhibitory activity(ies) regulates cellular DNA replication by modifying the cellular homologues of TAg. Purification and characterization of this inhibitory activity(ies) will contribute to our understanding of the mechanism developed by the cell to regulate DNA replication after exposure to ionizing radiation and will define a checkpoint operating in S phase. Genetic evidence for a checkpoint in S phase distinct from the checkpoints operating in G1 and G2 phase has been reported in yeast.
登录
查看更多内容
X 射线对用癌基因 H-ras 和 v-myc 转化原代大鼠胚胎成纤维细胞获得的细胞中 DNA 合成的长期抑制。
影响因子：
11.2
作者：
Wang,Y;Iliakis,G
通讯作者：
Iliakis,G
DOI：
--
发表时间：
1994-04
期刊：
The Journal of biological chemistry
影响因子：
--
作者：
S. Waga;Glenn BauerS;B. Stillman
通讯作者：
S. Waga;Glenn BauerS;B. Stillman
真核生物中染色体 DNA 复制的起始。
DOI：
10.1016/0168-9525(90)90305-p
发表时间：
1990
期刊：
Trends in genetics : TIG
影响因子：
--
作者：
Diffley,JF;Stillman,B
通讯作者：
Stillman,B
DOI：
10.1073/pnas.91.18.8655
发表时间：
1994-08-30
影响因子：
11.1
作者：
FLORESROZAS, H;KELMAN, Z;HURWITZ, J
通讯作者：
HURWITZ, J
反式作用因子对受辐射细胞中 DNA 复制的调节。
DOI：
--
发表时间：
1995
期刊：
Radiation research.
影响因子：
--
作者：
Wang,Y;Huq,MS;Cheng,X;Iliakis,G
通讯作者：
Iliakis,G