Du Jian, Cheng Li, Chen Li-jian, Chen Ying, An Ran
Department of Biochemistry and Molecular Biology, School of Basic Medicine, Hefei, China.
Xi Bao Yu Fen Zi Mian Yi Xue Za Zhi. 2012 Aug;28(8):804-7.
To screen 14-3-3Zeta-interacting proteins by the yeast two-hybrid system and confirm the interaction between 14-3-3Zeta and Polo-like kinase 1 (Plk1).
A bait vector pGBKT7-14-3-3Zeta was constructed to screen the 14-3-3Zeta-interacting proteins from the HeLa cDNA library by yeast two-hybrid system. The interaction between 14-3-3Zeta and Plk1 was further confirmed by immunoprecipitation assay. We also examined the distribution of endogenous 14-3-3Zeta and Plk1 in mitosis using immunofluorescent staining.
Plk1 was identified as a partner of 14-3-3Zeta protein by the yeast two-hybrid system. Immunoprecipitation assay demonstrated the interaction between 14-3-3Zeta and Plk1. Immunofluorescence indicated 14-3-3Zeta was colocalized with Plk1 at the midbody during cytokinesis.
Plk1 is a serine/threonine protein kinase that plays multiple critical roles in centrosome maturation, mitotic chromosome segregation, cytokinesis, and the DNA damage response. The interaction between 14-3-3Zeta and Plk1 suggests 14-3-3 family emerges as a novel player in the mitotic regulation.
通过酵母双杂交系统筛选与14-3-3ζ相互作用的蛋白，并证实14-3-3ζ与Polo样激酶1（Plk1）之间的相互作用。
构建诱饵载体pGBKT7-14-3-3ζ，通过酵母双杂交系统从HeLa cDNA文库中筛选与14-3-3ζ相互作用的蛋白。通过免疫沉淀试验进一步证实14-3-3ζ与Plk1之间的相互作用。我们还使用免疫荧光染色检测有丝分裂过程中内源性14-3-3ζ和Plk1的分布。
通过酵母双杂交系统鉴定出Plk1是14-3-3ζ蛋白的一个相互作用蛋白。免疫沉淀试验证明了14-3-3ζ与Plk1之间的相互作用。免疫荧光显示在细胞分裂期间14-3-3ζ与Plk1在中体处共定位。
Plk1是一种丝氨酸/苏氨酸蛋白激酶，在中心体成熟、有丝分裂染色体分离、细胞分裂和DNA损伤反应中发挥多种关键作用。14-3-3ζ与Plk1之间的相互作用表明14-3-3家族成为有丝分裂调控中的一个新参与者。
