Zou Tongtong, Mazan-Mamczarz Krystyna, Rao Jaladanki N, Liu Lan, Marasa Bernard S, Zhang Ai-Hong, Xiao Lan, Pullmann Rudolf, Gorospe Myriam, Wang Jian-Ying
Cell Biology Group, Department of Surgery, University of Maryland School of Medicine, Baltimore 21201, USA.
J Biol Chem. 2006 Jul 14;281(28):19387-94. doi: 10.1074/jbc.M602344200. Epub 2006 May 10.
Polyamines are essential for maintaining normal intestinal epithelial integrity, an effect that relies, at least in part, on their ability to keep low levels of nucleophosmin (NPM) and p53 mRNAs. The RNA-binding protein HuR associates with the p53 mRNA, as reported previously, and with the NPM mRNA, computationally predicted to be a target of HuR. Here, we show that HuR binds the NPM and p53 3'-untranslated regions and stabilizes these mRNAs in polyamine-depleted intestinal epithelial cells. Depletion of cellular polyamines by inhibiting ornithine decarboxylase with alpha-difluoromethylornithine dramatically enhanced the cytoplasmic abundance of HuR, whereas ectopic ornithine decarboxylase overexpression decreased cytoplasmic HuR; neither intervention changed whole-cell HuR levels. HuR was found to specifically bind the 3'-untranslated regions of NPN and p53 mRNAs. HuR silencing rendered the NPM and p53 mRNAs unstable and prevented increases in NPM and p53 mRNA and protein in polyamine-deficient cells. These results indicate that polyamines modulate cytoplasmic HuR levels in intestinal epithelial cells, in turn controlling the stability of the NPM and p53 mRNAs and influencing NPM and p53 protein levels.
多胺对于维持正常的肠道上皮完整性至关重要，这一作用至少部分依赖于它们维持低水平核磷蛋白（NPM）和p53 mRNA的能力。如先前报道，RNA结合蛋白HuR与p53 mRNA结合，并且通过计算预测其与NPM mRNA结合，NPM mRNA被认为是HuR的一个靶点。在此，我们表明HuR结合NPM和p53的3'非翻译区，并在多胺缺乏的肠道上皮细胞中稳定这些mRNA。用α-二氟甲基鸟氨酸抑制鸟氨酸脱羧酶来消耗细胞内的多胺，会显著增加HuR在细胞质中的丰度，而异位过表达鸟氨酸脱羧酶则会降低细胞质中HuR的水平；这两种干预措施均未改变全细胞HuR的水平。研究发现HuR特异性结合NPN和p53 mRNA的3'非翻译区。沉默HuR会使NPM和p53 mRNA不稳定，并阻止多胺缺乏细胞中NPM和p53 mRNA及蛋白水平的增加。这些结果表明，多胺调节肠道上皮细胞中细胞质HuR的水平，进而控制NPM和p53 mRNA的稳定性，并影响NPM和p53蛋白的水平。
