Pende A, Giacchè M, Castigliola L, Contini L, Passerone G, Patrone M, Port J D, Lotti G
Division of Internal Medicine 3-DI.M.I., University of Genoa School of Medicine, Genoa, Italy.
Biochem Biophys Res Commun. 1999 Dec 20;266(2):609-14. doi: 10.1006/bbrc.1999.1862.
An important mechanism of regulation of the expression of the AT(1) receptors is the modulation of the mRNA stability. AUF1, a human RNA-binding protein, may play an important role. Since AUF1 seems to bind to AU-rich regions of the 3'-untranslated region of the mRNAs, we verified the nucleotide sequence of human AT(1) receptor 3'-untranslated region and we found possible binding sites. In addition we evaluated the expression of the AUF1 protein in human vascular smooth muscle cells: the administration of both isoproterenol and angiotensin II induced a significant increase of total anti-AUF1 immunoreactive isoforms. At the same time angiotensin II induced a significant decrease in the AT(1) receptor mRNA abundance. Moreover, we found that recombinant human AUF1 protein binds to human AT(1) receptor riboprobes. The protein was able to bind to the distal portion of the 3'-untranslated region, and also to the coding region. Since the clinically relevant AT(1) receptor polymorphism is located in the 3'-untranslated region, we created two DNAs, corresponding to the A and C polymorphism, without any differences. Our data demonstrate the presence of AUF1 in human vascular smooth muscle cells and its modulation by activation of the beta-adrenergic and the AT(1) pathways, a and specific binding of AUF1 to the human AT(1) receptor mRNA, suggesting a role of this protein in the modulation of the AT(1) receptor expression.
AT(1)受体表达调控的一个重要机制是mRNA稳定性的调节。人RNA结合蛋白AUF1可能起重要作用。由于AUF1似乎与mRNA 3'-非翻译区富含AU的区域结合，我们验证了人AT(1)受体3'-非翻译区的核苷酸序列，并发现了可能的结合位点。此外，我们评估了人血管平滑肌细胞中AUF1蛋白的表达：异丙肾上腺素和血管紧张素II的给药均导致总抗AUF1免疫反应性亚型显著增加。同时，血管紧张素II导致AT(1)受体mRNA丰度显著降低。此外，我们发现重组人AUF1蛋白与人AT(1)受体核糖探针结合。该蛋白能够结合到3'-非翻译区的远端部分，也能结合到编码区。由于临床上相关的AT(1)受体多态性位于3'-非翻译区，我们创建了两个对应于A和C多态性的DNA，二者无差异。我们的数据证明了AUF1在人血管平滑肌细胞中的存在及其通过β-肾上腺素能和AT(1)途径激活的调节，以及AUF1与人AT(1)受体mRNA的特异性结合，表明该蛋白在AT(1)受体表达调节中起作用。
