Institute of Bioorganic Chemistry, Polish Academy of Sciences, 61-704 Poznan, Poland.
Int J Mol Sci. 2021 Dec 10;22(24):13306. doi: 10.3390/ijms222413306.
The p53 protein is one of the major transcriptional factors which guards cell homeostasis. Here, we showed that poly(C)-binding protein 2 (PCBP2) can bind directly to the 5' terminus of p53 mRNA by means of electrophoretic mobility shift assay. Binding sites of PCBP2 within this region of p53 mRNA were mapped using Pb2+-induced cleavage and SAXS methods. Strikingly, the downregulation of PCBP2 in HCT116 cells resulted in a lower level of p53 protein under normal and stress conditions. Quantitative analysis of p53 mRNA in PCBP2-downregulated cells revealed a lower level of p53 mRNA under normal conditions suggesting the involvement of PCBP2 in p53 mRNA stabilisation. However, no significant change in p53 mRNA level was observed upon PCBP2 depletion under genotoxic stress. Moreover, a higher level of p53 protein in the presence of rapamycin or doxorubicin and the combination of both antibiotics was noticed in PCBP2-overexpressed cells compared to control cells. These observations indicate the potential involvement of PCBP2 in cap-independent translation of p53 mRNA especially occurring under stress conditions. It has been postulated that the PCBP2 protein is engaged in the enhancement of p53 mRNA stability, probably via interacting with its 3' end. Our data show that under stress conditions PCBP2 also modulates p53 translation through binding to the 5' terminus of p53 mRNA. Thus PCBP2 emerges as a double-function factor in the p53 expression.
p53 蛋白是守护细胞内稳态的主要转录因子之一。在这里,我们表明聚(C)结合蛋白 2(PCBP2)可以通过电泳迁移率变动分析直接与 p53 mRNA 的 5'末端结合。使用 Pb2+诱导切割和 SAXS 方法绘制了 PCBP2 在 p53 mRNA 该区域内的结合位点。引人注目的是,在 HCT116 细胞中下调 PCBP2 会导致在正常和应激条件下 p53 蛋白水平降低。对 PCBP2 下调细胞中的 p53 mRNA 进行定量分析显示,在正常条件下 p53 mRNA 水平较低,提示 PCBP2 参与 p53 mRNA 稳定。然而,在遗传毒性应激下 PCBP2 耗尽时,p53 mRNA 水平没有明显变化。此外,与对照细胞相比,在 rapamycin 或阿霉素存在或两者联合存在的情况下,PCBP2 过表达细胞中的 p53 蛋白水平更高。这些观察结果表明 PCBP2 可能参与 p53 mRNA 的无帽依赖性翻译,尤其是在应激条件下。据推测,PCBP2 蛋白通过与其 3' 端相互作用,参与增强 p53 mRNA 的稳定性。我们的数据表明,在应激条件下,PCBP2 还通过与 p53 mRNA 的 5' 末端结合来调节 p53 翻译。因此,PCBP2 作为 p53 表达的双重功能因子出现。