Huang Xiuqing, Liu Jin, Shen Tao, Meng Xiangyu, Dou Lin, Lin Yajun, Li Jian
The Key Laboratory of Geriatrics, Beijing Hospital & Beijing Institute of Geriatrics, Ministry of Health, Beijing, 100730, China.
College of Life Sciences, Beijing Normal University, Beijing, 100875, China.
Cell Prolif. 2016 Apr;49(2):219-35. doi: 10.1111/cpr.12249. Epub 2016 Apr 3.
Protein phosphatase 4 (PP4) has been reported to be indispensable for cell proliferation and survival. Deletion of PP4 has been shown to induce abnormal and even lethal events in growth and development both in lower eukaryotes and in mammals. However, until now, effects of PP4 up-regulation have remained unclear.
To test effects of PP4 on cell proliferation, cell cycle and morphology in HepG2 cells, it was down-regulated using PP4 siRNA or its activity was inhibited using PP4RL (a PP4 phosphatase-dead mutant) adenoviruses. Alternatively, PP4 was up-regulated using PP4 adenoviruses. Next, we used a functional proteomic approach to identify proteins that may interact with PP4. Furthermore, we performed rescue experiments to verify the possible mechanisms.
To our surprise, we found that both up-regulation and inhibition of PP4 inhibited cell proliferation. Unlike PP4 inhibition, PP4 up-regulation induced prominent arrest at the prometaphase/metaphase transition by causing defects in chromosome alignment and spindle assembly. Moreover, we identified scaffold attachment factor A (SAF-A) (an important protein required for kinetochore-microtubule attachment that participates in the prometaphase/metaphase transition), to be a novel protein that interacts with PP4, using a proteomic approach. Thus, mutual regulatory mechanisms exist between PP4 and SAF-A. Interactions between PP4 and SAF-A played a role in prometaphase/metaphase transition.
Our data demonstrate a novel regulatory mechanism involving PP4 in cell proliferation.
据报道，蛋白磷酸酶4（PP4）对于细胞增殖和存活不可或缺。在低等真核生物和哺乳动物中，PP4的缺失已被证明会在生长发育过程中引发异常甚至致死事件。然而，迄今为止，PP4上调的影响仍不清楚。
为了测试PP4对HepG2细胞中细胞增殖、细胞周期和形态的影响，使用PP4 siRNA下调PP4，或使用PP4RL（一种PP4磷酸酶失活突变体）腺病毒抑制其活性。或者，使用PP4腺病毒上调PP4。接下来，我们采用功能蛋白质组学方法来鉴定可能与PP4相互作用的蛋白质。此外，我们进行了挽救实验以验证可能的机制。
令我们惊讶的是，我们发现PP4的上调和抑制均抑制细胞增殖。与PP4抑制不同，PP4上调通过导致染色体排列和纺锤体组装缺陷，在有丝分裂前期/中期转换阶段诱导显著停滞。此外，我们采用蛋白质组学方法鉴定出支架附着因子A（SAF-A）（一种参与有丝分裂前期/中期转换的着丝粒-微管附着所需的重要蛋白质）是一种与PP4相互作用的新蛋白质。因此，PP4与SAF-A之间存在相互调节机制。PP4与SAF-A之间的相互作用在有丝分裂前期/中期转换中发挥作用。
我们的数据证明了一种涉及PP4的细胞增殖新调控机制。
