Apak的亚细胞定位机制研究

Subcellular localization of Apak,a negative regulator of P53

目的探讨P53的调控分子Apak亚细胞定位的结构基础。方法在热休克及DNA损伤诱导剂MMS处理下,通过免疫荧光技术观察Apak的亚细胞定位变化,并通过激光共聚焦荧光显微镜观察Apak截短体的亚细胞定位。结果 Apak多以均匀的形式分布在细胞核质,偶见在核仁中聚集;在外界刺激后Apak核内的均匀分布减少,主要以点状的大颗粒形式在核仁中呈聚集样分布;Apak的9种截短体均可定位于核内,且在核质和核仁中具有明显的比例差异。结论在DNA损伤信号下Apak有一个从核质向核仁移位的动态过程;KRAB区可以拮抗Apak在核仁的定位,而锌指尤其是锌指数的数目(锌指数>14)则决定了Apak能否在核仁定位。

Objective To study the mechanism determining the subcellular localization of Apak which negatively regulates P53.Methods Immunofluorescence assay（IF） was used to observe the subcellular localization of Apak in response to stimuli such as heat shock and DNA damage agent MMS.The localization of Apak truncated forms was analyzed under the confocal laser fluorescence microscope.Results Apak was evenly distributed in cell nuclei and gathered in nucleoli on occasion,but was accumulated mainly in the form of blob-shaped large particles in the nucleolus in response to stimuli.All the truncated forms of Apak were localized in the nucleus but presented with different ratios between the nucleolus and nucleoplasm.Conclusion Our data here demonstrate that DNA damage stimuli promote the translocation of Apak from the nucleoplasm to the nucleolus.The number of Zn-finger domains in Apak determines its nucleolus localization while the KRAB region performs a contrary function.