Self-renewal of human embryonic stem cells is supported by a shortened G1 cell cycle phase
Self-renewal of human embryonic stem cells is supported by a shortened G1 cell cycle phase
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DOI：
10.1002/jcp.20776
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发表时间：
2006-12-01
影响因子：
5.6
通讯作者：
Stein, Gary S.
中科院分区：
文献类型：
--
作者：
Becker, Klaus A.;Ghule, Prachi N.;Stein, Gary S.
人类胚胎干细胞（ES）的自我更新能力与多能性有关。然而，人类胚胎干细胞分裂的基本参数非常缺乏。在这项研究中，我们发现与体细胞（如正常的二倍体IMR90成纤维细胞和NT-2畸胎癌细胞）相比，人类胚胎干细胞（HI和H9； nih指定的WA01和WA09）由于整体细胞周期非常短（15-16小时）而快速增殖。人类胚胎干细胞周期维持G1、S、G2和M四个典型的细胞周期阶段，但G1的持续时间明显缩短。溴脱氧尿苷（Bromodeoxyuridine, BrdU）结合和FACS分析表明，65%的异步生长的人胚胎干细胞处于S期。检测BrdU标记的有丝分裂染色体、Ki67结构域和含有Cajal小体的p220（NPAT）的免疫荧光显微镜研究显示，ES和体细胞的S期（类似8小时）、G2期（类似4小时）和M期（类似1小时）的持续时间相似。我们确定人类胚胎干细胞在与诺可达唑或抗肿瘤药物紫杉醇（紫杉醇）同步后仍然具有活力，并且G1期缩短仅为2.5-3小时，明显短于体细胞。利用定量RT-PCR的分子分析表明，人类胚胎干细胞和体细胞表达相似的细胞周期标记。然而，在细胞周期蛋白和细胞周期蛋白依赖性激酶（CDKs）中，我们观察到G1相关的CDK4和细胞周期蛋白D2基因的mRNA水平较高。我们得出结论，人类胚胎干细胞表现出独特的G1细胞周期动力学，并利用CDK4/周期蛋白D2相关机制获得DNA复制能力。
Competency for self-renewal of human embryonic stem (ES) cells is linked to pluripotency. However, there is a critical paucity of fundamental parameters of human ES cell division. In this study we show that human ES cells (HI and H9; NIH-designated WA01 and WA09) rapidly proliferate due to a very short overall cell cycle (15-16 h) compared to somatic cells (e.g., normal diploid IMR90 fibroblasts and NT-2 teratocarcinoma cells). The human ES cell cycle maintains the four canonical cell cycle stages G1, S, G2, and M, but the duration of G1 is dramatically shortened. Bromodeoxyuridine (BrdU) incorporation and FACS analysis demonstrated that 65% of asynchronously growing human ES cells are in S phase. Immunofluorescence microscopy studies detecting BrdU labeled mitotic chromosomes, Ki67 domains, and p220(NPAT) containing Cajal bodies revealed that the durations of the S (similar to 8 h), G2 (similar to 4 h), and M phases(similar to 1 h) are similar in ES and somatic cells. We determined that human ES cells remain viable after synchronization with either nocodazole or the anti-tumor drug Paclitaxel (taxol) and have an abbreviated G1 phase of only 2.5-3 h that is significantly shorter than in somatic cells. Molecular analyses using quantitative RT-PCR demonstrate that human ES cells and somatic cells express similar cell cycle markers. However, among cyclins and cyclin-dependent kinases (CDKs), we observed high mRNA levels for the G1 related CDK4 and cyclin D2 genes. We conclude that human ES cells exhibit unique G1 cell cycle kinetics and use CDK4/cyclin D2 related mechanisms to attain competency for DNA replication.