Cdc25B在小鼠受精卵的表达和定位

为阐明细胞分裂周期(Cdc)25B调控小鼠受精卵发育的机制,利用Western印迹检测小鼠受精卵各时期Cdc25B的表达及Cdc2-Tyr15的磷酸化状态。利用间接免疫荧光技术观察Cdc25B在小鼠受精卵的定位。构建pEGFP-Cdc25B融合表达载体并显微注射到受精卵中,观察Cdc25B在受精卵M期的定位变化。结果表明Cdc25B在G1和S期被磷酸化,在G2和M期去磷酸化。Cdc2-Tyr15在G1和S期处于磷酸化状态,G2期只检测到Cdc2-Tyr15轻微的磷酸化信号,M期未检测到任何Cdc2-Tyr15的磷酸化信号。Cdc25B在G1期定位于细胞质和细胞核中,S和G2期定位于细胞质的皮质部分,M期由细胞质转向核区。证明Cdc25B核输出后激活有丝分裂促进因子,从而启动小鼠受精卵的有丝分裂。

Expression and Localization of Cdc25B in Mouse Fertilized Eggs

The purpose of this study is to elucidate the mechanism of Cdc25B regulating the development of mouse fertilized eggs. Using Western blot to detect the expression of Cdc25B and the phosphorylation status of Cdc2-Tyr15 at various phases in mouse fertilized eggs. The localization of Cdc25B was investigated using fluores- cence microscopy at one-cell stage. pEGFP-Cdc25B plasmid was microinjected to mouse fertilized eggs in order to detect location variation of Cdc25B at mitosis. Western blots revealed that Cdc25B was phosphorylated at G1 and S phase, dephosphorylated at G2 and M phase. There was strong phosphorylation signal of Cdc2-Tyr15 detected at G1 and S whereas only slight phosphorylation signal of Cdc2-Tyr15 found at G2 and no phosphorylation signal of Cdc2- Tyr15 was identified at M phase in mouse fertilized eggs. The subcellular localization of Cdc25B showed that Cdc25B was mainly in cytoplasm and nucleus at G1, but in cytoplasm cortex at S and G2 phase. Cdc25B transferred from cytoplasm to nucleus region in part at M phase in mouse fertilized eggs. Our findings identify that Cdc25B exported from nucleus initiates the mitosis by activating M-phase promoting factor in mouse fertilized eggs.