Nakuci Enkeleda, Xu Mai, Pujana Miguel Angel, Valls Joan, Elshamy Wael M
Dana-Farber Cancer Institute and Harvard Medical School, Boston, MA 02115, USA.
Int J Biochem Cell Biol. 2006;38(7):1207-20. doi: 10.1016/j.biocel.2005.12.017. Epub 2006 Feb 3.
Previous studies suggested that geminin plays a vital role in both origin assembly and DNA re-replication during S-phase; however, no data to support a role for geminin in G2/M cells have been described. Here it is shown that in G2/M-phase, geminin participates in the promotion of proper cytokinesis. This claim can be supported through a series of observations. First, geminin in G2/M is loaded onto chromatin after it is tyrosine phosphorylated. It is unlike S-phase geminin that resides in the nuclear soluble fraction, where it is exclusively S/T phosphorylated. Secondly, on chromatin, geminin gets S/T phosphorylated in late G1; this modification causes the release of geminin from the chromatin. Cyclins bind and phosphorylate geminin in a sequential, cell cycle-dependent manner. These modifications correlated well with geminin departure from the chromatin. This suggests that cyclin functions to either release geminin from chromatin or at least keep it at bay until late S-phase. Thirdly, depletion of geminin from a diploid mammary epithelial cell line (HME) causes cells to arrest in late G2/M-phase. Massive serine-10 phosphorylated histone H3 staining and survivin localization to mid-body were observed; this suggests that they could be arrested in either mitosis or at cytokinesis. Finally, while in the absence of geminin, cyclin B1, chk1 and cdc7 are all over expressed. This paper will demonstrate that only cdc7 is important in maintaining the cytokinesis arrest in the absence of geminin. Only double depletion of geminin and cdc7 induce apoptosis. Our results taken together show, for the first time, that phosphorylation-induction activates oscillation of geminin between both nuclear soluble and chromatin compartments. Chromatin-bound geminin species functions to initiate or maintain proper cytokineses. In the absence of geminin, cells arrest in cytokinesis; this defines a novel checkpoint, monitored by cdc7, rather than cyclin B1 or chk1.
先前的研究表明，geminin在S期的起始装配和DNA再复制过程中都起着至关重要的作用；然而，尚无数据支持geminin在G2/M期细胞中发挥作用。本文表明，在G2/M期，geminin参与促进正常的胞质分裂。这一观点可通过一系列观察结果得到支持。首先，G2/M期的geminin在酪氨酸磷酸化后加载到染色质上。这与S期存在于核可溶性部分的geminin不同，后者仅发生丝氨酸/苏氨酸（S/T）磷酸化。其次，在染色质上，geminin在G1晚期发生S/T磷酸化；这种修饰导致geminin从染色质上释放。细胞周期蛋白以连续的、细胞周期依赖性的方式结合并磷酸化geminin。这些修饰与geminin从染色质上脱离密切相关。这表明细胞周期蛋白的作用是要么将geminin从染色质上释放出来，要么至少在S期晚期之前抑制它。第三，从二倍体乳腺上皮细胞系（HME）中去除geminin会导致细胞在G2/M期晚期停滞。观察到大量丝氨酸-10磷酸化组蛋白H3染色以及生存素定位于中体；这表明它们可能在有丝分裂或胞质分裂时停滞。最后，在没有geminin的情况下，细胞周期蛋白B1、chk1和cdc7均过度表达。本文将证明，在没有geminin的情况下，只有cdc7对维持胞质分裂停滞很重要。只有同时去除geminin和cdc7才会诱导细胞凋亡。我们的研究结果共同表明，磷酸化诱导激活了geminin在核可溶性和染色质区室之间的振荡。与染色质结合的geminin物种起到启动或维持正常胞质分裂的作用。在没有geminin的情况下，细胞在胞质分裂时停滞；这定义了一个由cdc7而非细胞周期蛋白B1或chk1监测的新检查点。
