Bioluminescence imaging of DNA synthetic phase of cell cycle in living animals.
Bioluminescence imaging of DNA synthetic phase of cell cycle in living animals.
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活体动物细胞周期 DNA 合成阶段的生物发光成像
DOI：
10.1371/journal.pone.0053291
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发表时间：
2013
期刊：
影响因子：
3.7
通讯作者：
Zhang GJ
中科院分区：
文献类型：
--
作者：
Chen ZH;Zhao RJ;Li RH;Guo CP;Zhang GJ
生物发光报告蛋白已广泛应用于开发监测活细胞生物事件的工具。目前，流式细胞术等检测方法可用于体外研究DNA合成相（s相）靶向抗癌药物的活性；然而，体内模型的成像技术仍然有限。已知Cyclin A2促进哺乳动物进入s期。其表达水平在g1期较低，但在s期开始时升高。细胞周期蛋白A2在前期通过泛素依赖、蛋白酶体介导的蛋白水解降解。在这项研究中，我们开发了一种细胞周期蛋白a2 -荧光素酶（CYCA-Luc）融合蛋白，用于泛素-蛋白酶体依赖性降解，并评估了其在筛选s期靶向抗癌药物中的应用。与内源性cyclin A2类似，CYCA-Luc在s期积累，在G2/ m期降解。利用Cdc20 siRNA，我们已经证明了Cdc20可以介导CYCA-Luc降解。此外，利用无创生物发光成像技术，我们证实了CYCA-Luc在体内和体外对10-羟基喜树碱（HCPT）的反应中积累，HCPT是一种s期靶向抗癌药物。我们的研究结果表明，CYCA-Luc融合报告系统可以实时监测细胞周期s期，并在体外和体内评估抗癌药物HCPT的药理活性，可能为筛选此类药物提供重要工具。
Bioluminescence reporter proteins have been widely used in the development of tools for monitoring biological events in living cells. Currently, some assays like flow cytometry analysis are available for studying DNA synthetic phase (S-phase) targeted anti-cancer drug activity in vitro; however, techniques for imaging of in vivo models remain limited. Cyclin A2 is known to promote S-phase entry in mammals. Its expression levels are low during G1-phase, but they increase at the onset of S-phase. Cyclin A2 is degraded during prometaphase by ubiquitin-dependent, proteasome-mediated proteolysis. In this study, we have developed a cyclin A2-luciferase (CYCA-Luc) fusion protein targeted for ubiquitin-proteasome dependent degradation, and have evaluated its utility in screening S-phase targeted anti-cancer drugs. Similar to endogenous cyclin A2, CYCA-Luc accumulates during S-phase and is degraded during G2/M-phase. Using Cdc20 siRNA we have demonstrated that Cdc20 can mediate CYCA-Luc degradation. Moreover, using noninvasive bioluminescent imaging, we demonstrated accumulation of CYCA-Luc in response to 10-hydroxycamptothecin (HCPT), an S-phase targeted anti-cancer drug, in human tumor cells in vivo and in vitro. Our results indicate that a CYCA-Luc fusion reporter system can be used to monitor S-phase of cell cycle, and evaluate pharmacological activity of anti-cancer drug HCPT in real time in vitro and in vivo, and is likely to provide an important tool for screening such drugs.
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影响因子：
8
作者：
Mateo, F.;Vidal-Laliena, M.;Canela, N.;Busino, L.;Martinez-Balbas, M. A.;Pagano, M.;Agell, N.;Bachs, O.
通讯作者：
Bachs, O.
影响因子：
5.8
作者：
Abdel-Aziz, W;Hickey, RJ;Malkas, LH
通讯作者：
Malkas, LH
影响因子：
4.8
作者：
Zhang, Guo-Jun;Chen, Tsing-Bau;Williams, David L., Jr.
通讯作者：
Williams, David L., Jr.
影响因子：
3.7
作者：
Honda A;Valogne Y;Bou Nader M;Bréchot C;Faivre J
通讯作者：
Faivre J
DOI：
10.1083/jcb.153.1.137
发表时间：
2001-04-02
期刊：
The Journal of cell biology
影响因子：
--
作者：
Geley S;Kramer E;Gieffers C;Gannon J;Peters JM;Hunt T
通讯作者：
Hunt T