Parisi Tiziana, Beck Andreas R, Rougier Nathalie, McNeil Tom, Lucian Linda, Werb Zena, Amati Bruno
DNAX Research Institute, 901 California Avenue, Palo Alto, CA 94304, USA.
EMBO J. 2003 Sep 15;22(18):4794-803. doi: 10.1093/emboj/cdg482.
In mammalian cells, cyclin E-CDK2 complexes are activated in the late G1 phase of the cell cycle and are believed to have an essential role in promoting S-phase entry. We have targeted the murine genes CCNE1 and CCNE2, encoding cyclins E1 and E2. Whereas single knockout mice were viable, double knockout embryos died around midgestation. Strikingly, however, these embryos showed no overt defects in cell proliferation. Instead, we observed developmental phenotypes consistent with placental dysfunction. Mutant placentas had an overall normal structure, but the nuclei of trophoblast giant cells, which normally undergo endoreplication and reach elevated ploidies, showed a marked reduction in DNA content. We derived trophoblast stem cells from double knockout E3.5 blastocysts. These cells retained the ability to differentiate into giant cells in vitro, but were unable to undergo multiple rounds of DNA synthesis, demonstrating that the lack of endoreplication was a cell-autonomous defect. Thus, during embryonic development, the needs for E-type cyclins can be overcome in mitotic cycles but not in endoreplicating cells.
在哺乳动物细胞中，细胞周期蛋白E - CDK2复合物在细胞周期的G1晚期被激活，并且被认为在促进进入S期过程中起关键作用。我们靶向了编码细胞周期蛋白E1和E2的小鼠基因CCNE1和CCNE2。单基因敲除小鼠是存活的，而双基因敲除胚胎在妊娠中期左右死亡。然而，令人惊讶的是，这些胚胎在细胞增殖方面没有明显缺陷。相反，我们观察到与胎盘功能障碍一致的发育表型。突变胎盘的整体结构正常，但通常进行核内复制并达到高倍体的滋养层巨细胞的细胞核显示DNA含量显著降低。我们从双基因敲除的E3.5囊胚中获得了滋养层干细胞。这些细胞在体外保留了分化为巨细胞的能力，但无法进行多轮DNA合成，这表明核内复制的缺失是一种细胞自主缺陷。因此，在胚胎发育过程中，有丝分裂周期中对E型细胞周期蛋白的需求可以被克服，但在进行核内复制的细胞中则不能。
