摘要
目的:检测小鼠1-细胞期受精卵各个细胞周期中蛋白激酶Wee1B的表达水平及其定位水平,初步阐明Wee1B对哺乳动物受精卵早期发育调控的机制。方法:超排卵获得小鼠1-细胞期受精卵,收集卵细胞后用RT-PCR和Western blotting方法检测Wee1B的mRNA和蛋白表达水平;放射自显影测定Wee1B活性;间接免疫荧光观察Wee1B在各个细胞时期的定位。结果:小鼠1-细胞期受精卵G1、S、G2和M期Wee1BmRNA相对表达量比较差异均无统计学意义(P>0.05);S、G2和M期Wee1B蛋白表达量与G1期比较差异有统计学意义(P<0.01);与G1期比较,S、G2期Wee1B活性逐渐增高(P>0.05,P<0.01),M期Wee1B活性迅速下降(P<0.01)。G1期和S期细胞Wee1B蛋白偶联的红色荧光信号主要集中分布于细胞核,但在G2期细胞红色荧光信号弥散分布于细胞浆,在M末期和2-细胞期可见红色荧光信号又重新分布于细胞核,细胞浆内荧光减弱。结论:蛋白激酶Wee1B在小鼠1-细胞期受精卵各个细胞周期动态表达,Wee1B蛋白存在核浆转位,Wee1B的定位变化可能调控小鼠受精卵有丝分裂。
Objective To observe the expression and localization of protein kinase Wee1Bin mouse one-cell-stage embryos,and to pay foundation for further study on the mechanism of the early development of mouse embryos.Methods One-cell-stage mouse embryos were collected after superovulation,and the mRNA and protein expression levels of Wee1B were examined by RT-PCR and Western blotting,respectively.The Wee1Bactivity in mouse embryos was detected by autoradiography and the localization of Wee1Bin each cell cycle was observed by indirect immunofluorescence.Results There were no significant differences in the expression levels of Wee1B mRNA between G1,S,G2 and M phases（P0.05）;but there were significant differences in the expression levels of Wee1Bprotein between S,G2,M phases and G1 phase（P0.01）.Compared with G1 phase,the activities of Wee1Bin G2 phase and S phase were increased（P0.05,P0.01）,but the activity of Wee1Bin M phase was decreased（P0.01）.Wee1Bdistributed in the nucleus at the G1 and S phases and translocated to the cytoplasm at the G2 /M transition in one-cell-stage embryos.Conclusion The protein and activity of Wee1Bare dynamically expressed in one-cell-stage embryos and the nucleocytoplasmic shuttling of Wee1Bmay be the mechanism of early development of mouse embryos.
出处
《吉林大学学报（医学版）》
CAS
CSCD
北大核心
2013年第5期863-867,I0001,共6页
Journal of Jilin University：Medicine Edition
基金
国家自然科学基金资助课题(81270654
81270698)
辽宁省教育厅科研基金资助课题(L2012303)
