Mikulenkova Erika, Neradil Jakub, Zitterbart Karel, Sterba Jaroslav, Veselska Renata
Laboratory of Tumor Biology, Department of Experimental Biology, School of Science, Masaryk University, Kotlarska 2, 611 37, Brno, Czech Republic.
Regional Centre for Applied Molecular Oncology, Masaryk Memorial Cancer Institute, Zluty kopec 7, 656 53, Brno, Czech Republic.
Tumour Biol. 2015 Sep;36(10):7483-91. doi: 10.1007/s13277-015-3474-3. Epub 2015 Apr 25.
The p73 protein is a member of the p53 family, and this protein is known to be essential for the maintenance of genomic stability, DNA repair, and apoptosis regulation. Transcription from two promoters leads to two main N-terminal isoforms: the TAp73 isoform is reported to have tumor suppressor function, whereas the ΔNp73 isoform likely has oncogenic potential. The present study is focused on the investigation of a possible role of both these p73 N-terminal isoforms in the process of centrosome amplification. HGG-02 and GM7 glioblastoma cell lines and the Daoy medulloblastoma cell line were used in this study. The cells were analyzed using indirect immunofluorescence to determine TAp73 and ΔNp73 expression patterns and possible co-localization with the BubR1 protein, as well as the number of centrosomes. A transiently transfected GM7 cell line was used to verify the results concerning the N-terminal isoforms in relation to centrosome amplification. We found that increased immunoreactivity for the ΔNp73 isoform is associated with the occurrence of an abnormal number of centrosomes in particular cells. Using the transiently transfected GM7 cell line, we confirmed that centrosome amplification is present in cells with overexpression of the ΔNp73 isoform. In contrast, the immunoreactivity for the TAp73 isoform was weak or medium in most of the cells with an aberrant number of centrosomes. To determine the putative counterpart of the p73 N-terminal isoforms among spindle assembly checkpoint (SAC) proteins, we also evaluated possible interactions between the N-terminal isoforms and BubR1 protein, but no co-localization of these proteins was observed.
p73蛋白是p53家族的成员之一，已知该蛋白对于维持基因组稳定性、DNA修复及细胞凋亡调控至关重要。由两个启动子转录产生两种主要的N端异构体：据报道，TAp73异构体具有肿瘤抑制功能，而ΔNp73异构体可能具有致癌潜力。本研究聚焦于探究这两种p73 N端异构体在中心体扩增过程中可能发挥的作用。本研究使用了HGG - 02和GM7胶质母细胞瘤细胞系以及Daoy髓母细胞瘤细胞系。通过间接免疫荧光分析细胞，以确定TAp73和ΔNp73的表达模式以及它们与BubR1蛋白的可能共定位情况，同时确定中心体的数量。使用瞬时转染的GM7细胞系来验证与中心体扩增相关的N端异构体的结果。我们发现，ΔNp73异构体免疫反应性增加与特定细胞中中心体数量异常的发生有关。使用瞬时转染的GM7细胞系，我们证实中心体扩增存在于过表达ΔNp73异构体的细胞中。相比之下，在大多数中心体数量异常的细胞中，TAp73异构体的免疫反应性较弱或中等。为了确定纺锤体组装检查点（SAC）蛋白中p73 N端异构体的假定对应物，我们还评估了N端异构体与BubR1蛋白之间的可能相互作用，但未观察到这些蛋白的共定位。
