Dissociation of centrosome replication events from cycles of DNA synthesis and mitotic division in hydroxyurea-arrested Chinese hamster ovary cells.
Dissociation of centrosome replication events from cycles of DNA synthesis and mitotic division in hydroxyurea-arrested Chinese hamster ovary cells.
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DOI：
10.1083/jcb.130.1.105
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发表时间：
1995-07
影响因子：
7.8
通讯作者：
BRINKLEY, BR
中科院分区：
文献类型：
--
作者：
BALCZON, R;BAO, LM;ZIMMER, WE;BROWN, K;ZINKOWSKI, RP;BRINKLEY, BR
关于体细胞用来调节中心体复合体复制的机制，人们知之甚少。用电子显微镜和免疫荧光显微镜观察了CHO细胞的中心体倍增，用人自身免疫抗球体抗血清和Northern印迹法研究了中心体自身抗原PCM-1的编码部分。中心体加倍可以通过使用羟基脲或aphidiclin将细胞阻止在细胞周期的G1/S交界处而从DNA合成和有丝分裂周期中分离出来。用SPJ人自身免疫抗球体抗血清进行的免疫荧光显微镜观察表明，在没有DNA合成和有丝分裂周期的情况下，停滞的细胞能够进行多次中心体复制。Northern印迹分析表明，PCM-1mRNA的合成和降解在CHO细胞中呈细胞周期依赖性，在G1期和S期细胞中出现峰值，然后在G2和M期下降到检测不到的水平。相反，滞留在细胞周期G1/S交界处的细胞将PCM-1mRNA维持在人为升高的水平，这为解释羟基脲诱导的CHO细胞在长时间停滞期间发生多轮中心体复制提供了可能的分子机制。在羟基脲阻断的CHO细胞中，通过在透析血清中培养细胞，可以消除复制中心体的能力。然而，在透析后的血清中加入EGF可以重新启动复制中心体和合成PCM-1mRNA的能力。该实验系统可用于研究体细胞调节中心体复制的正、负分子机制，以及体细胞协调中心体复制与其他细胞周期进程事件的方法。
Relatively little is known about the mechanisms used by somatic cells to regulate the replication of the centrosome complex. Centrosome doubling was studied in CHO cells by electron microscopy and immunofluorescence microscopy using human autoimmune anticentrosome antiserum, and by Northern blotting using the cDNA encoding portion of the centrosome autoantigen pericentriolar material (PCM)-1. Centrosome doubling could be dissociated from cycles of DNA synthesis and mitotic division by arresting cells at the G1/S boundary of the cell cycle using either hydroxyurea or aphidicolin. Immunofluorescence micros-copy using SPJ human autoimmune anticentrosome antiserum demonstrated that arrested cells were able to undergo numerous rounds of centrosome replication in the absence of cycles of DNA synthesis and mitosis. Northern blot analysis demonstrated that the synthesis and degradation of the mRNA encoding PCM-1 occurred in a cell cycle-dependent fashion in CHO cells with peak levels of PCM-1 mRNA being present in G1 and S phase cells before mRNA amounts dropped to undetectable levels in G2 and M phases. Conversely, cells arrested at the G1/S boundary of the cell cycle maintained PCM-1 mRNA at artificially elevated levels, providing a possible molecular mechanism for explaining the multiple rounds of centrosome replication that occurred in CHO cells during prolonged hydroxyurea-induced arrest. The capacity to replicate centrosomes could be abolished in hydroxyurea-arrested CHO cells by culturing the cells in dialyzed serum. However, the ability to replicate centrosomes and to synthesize PCM-1 mRNA could be re- initiated by adding EGF to the dialyzed serum. This experimental system should be useful for investigating the positive and negative molecular mechanisms used by somatic cells to regulate the replication of centrosomes and for studying and the methods used by somatic cells for coordinating centrosome duplication with other cell cycle progression events.
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影响因子：
7.8
作者：
KURIYAMA, R;BORISY, GG
通讯作者：
BORISY, GG
影响因子：
--
作者：
KOVACS, AM;ZIMMER, WE
通讯作者：
ZIMMER, WE
影响因子：
7.8
作者：
ROBBINS, E;JENTZSCH, G;MICALI, A
通讯作者：
MICALI, A
影响因子：
--
作者：
BALCZON, R;WEST, K
通讯作者：
WEST, K
影响因子：
64.5
作者：
HANDELI, S;WEINTRAUB, H
通讯作者：
WEINTRAUB, H