Sengupta Sibani, Jang Byeong-Churl, Wu Ming-Tao, Paik Ji-Hye, Furneaux Henry, Hla Timothy
Center for Vascular Biology, Department of Physiology, University of Connecticut Health Center, 263 Farmington Avenue, 06030-3501, USA.
J Biol Chem. 2003 Jul 4;278(27):25227-33. doi: 10.1074/jbc.M301813200. Epub 2003 Apr 18.
The cyclooxygenase-2 (COX-2) gene encodes the inducible prostaglandin synthase enzyme implicated in inflammation, cell growth, and tumorigenesis. Regulation of the COX-2 gene expression at the post-transcriptional level is poorly understood. For example, protein factors that regulate the post-transcriptional mRNA metabolism of COX-2 have not been fully characterized. In this study, we demonstrate that the RNA-binding protein HuR binds to COX-2 mRNA and regulates its expression. We show that there are three binding sites for HuR in the 3'-untranslated region of human COX-2. These sites are located at the following positions in the COX-2 3'-untranslated region: 39-84 nucleotides (nt), 1155-1187 nt, and 1244-1256 nt (hereinafter referred to as Sites I, II and III, respectively). Although all three sites are present in the 4.6-kb COX-2 mRNA, only site I is present in the shorter 2.8-kb isoform. HuR in MDA-MB-231 cell extracts associated with COX-2 mRNA at the identified sites. Further, HuR location in the cytoplasm was induced by serum withdrawal, a stimulus known to induce COX-2 mRNA. Down-regulation of HuR by two independent methods, namely RNA interference as well as antisense RNA expression, significantly attenuated serum withdrawal-induced increase in COX-2 mRNA (both the 4.6- and 2.8-kb isoforms) and protein levels. These data suggest that HuR binding to COX-2 is critical for its post-transcriptional mRNA stabilization.
环氧化酶-2（COX-2）基因编码诱导型前列腺素合酶，该酶与炎症、细胞生长和肿瘤发生有关。目前对COX-2基因在转录后水平的表达调控了解甚少。例如，调节COX-2转录后mRNA代谢的蛋白质因子尚未完全明确。在本研究中，我们证明RNA结合蛋白HuR与COX-2 mRNA结合并调节其表达。我们发现人COX-2的3'-非翻译区有三个HuR结合位点。这些位点在COX-2 3'-非翻译区的位置如下：39-84个核苷酸（nt）、1155-1187 nt和1244-1256 nt（以下分别称为位点I、II和III）。尽管所有三个位点都存在于4.6 kb的COX-2 mRNA中，但较短的2.8 kb异构体中仅存在位点I。MDA-MB-231细胞提取物中的HuR在确定的位点与COX-2 mRNA结合。此外，血清饥饿可诱导HuR在细胞质中的定位，血清饥饿是一种已知可诱导COX-2 mRNA的刺激因素。通过两种独立方法，即RNA干扰和反义RNA表达下调HuR，可显著减弱血清饥饿诱导的COX-2 mRNA（4.6 kb和2.8 kb异构体）和蛋白水平的增加。这些数据表明，HuR与COX-2的结合对其转录后mRNA的稳定性至关重要。
