Easwaran Hariharan P, Leonhardt Heinrich, Cardoso M Cristina
Max Delbrück Center for Molecular Medicine, Berlin, Germany.
Cell Cycle. 2005 Mar;4(3):453-5. doi: 10.4161/cc.4.3.1525. Epub 2005 Mar 7.
Many cellular processes are regulated by cell cycle dependent changes in protein dynamics and localization. Studying these changes in vivo requires methods to distinguish the different cell cycle stages. Here we demonstrate the use of DNA Ligase I fused to DsRed1 as an in situ marker to identify S phase and the subsequent transition to G2 in live cells. Using this marker, we observed changes in the nuclear distribution of Dnmt1 during cell cycle progression. Based on the different nuclear distribution of DNA Ligase I and Dnmt1 in G2 and G1, we demonstrate that the combination of both proteins allows the direct discrimination of all cell cycle phases using either immunostainings or fusions with fluorescent proteins. These markers are new tools to directly study cell cycle dependent processes in both, fixed and living cells.
许多细胞过程受蛋白质动力学和定位的细胞周期依赖性变化调控。在体内研究这些变化需要能够区分不同细胞周期阶段的方法。在此，我们展示了将与DsRed1融合的DNA连接酶I用作原位标记物，以识别活细胞中的S期以及随后向G2期的转变。使用该标记物，我们观察到细胞周期进程中Dnmt1核分布的变化。基于G2期和G1期DNA连接酶I和Dnmt1不同的核分布，我们证明这两种蛋白质的组合能够通过免疫染色或与荧光蛋白融合直接区分所有细胞周期阶段。这些标记物是直接研究固定细胞和活细胞中细胞周期依赖性过程的新工具。
