Department of Biomolecular Science, Faculty of Engineering, Gifu University, 1-1 Yanagido, Gifu 501-1193, Japan.
BMC Mol Biol. 2012 Jan 17;13:2. doi: 10.1186/1471-2199-13-2.
Glial cell line-derived neurotrophic factor (GDNF) is a potent survival factor for many cell types, and its expression is widespread both within and outside of the nervous system. The regulation of GDNF expression has been extensively investigated but is not fully understood.
Using a luciferase reporter assay, we identified the role of the 3'-untranslated region (3'-UTR) of the mouse GDNF gene in the regulation of gene expression. We focused on a well-conserved A- and T-rich region (approximately 200 bp in length), which is located approximately 1000 bp downstream of the stop codon in exon 4 of the gene and contains three typical AU-rich elements (AREs), AUUUA. Interestingly, these AREs are well conserved in several GDNF genes. By testing reporter constructs containing various regions and lengths of the 3'-UTR fused to the end of the luciferase gene, we demonstrated that the ARE-induced decrease in luciferase activity correlates with the attenuation of the mRNA stability. Furthermore, we found that several regions around the AREs in the 3'-UTR suppressed the luciferase activity. Moreover, the expression level of the GDNF protein was negligible in C6 glioma cells transfected with the ARE-containing GDNF expression vector.
Our study is the first characterization of the possible role of AREs and other suppressive regions in the 3'-UTR in regulating the amounts of GDNF mRNA in C6 cells.
胶质细胞源性神经营养因子（GDNF）是许多细胞类型的有效存活因子，其表达在神经系统内外广泛存在。GDNF 表达的调节已被广泛研究，但尚未完全理解。
我们使用荧光素酶报告基因检测，确定了小鼠 GDNF 基因 3'-非翻译区（3'-UTR）在基因表达调控中的作用。我们集中研究了一个高度保守的 A 和 T 富含区（约 200 个碱基对长），位于基因外显子 4 的终止密码子下游约 1000 个碱基对处，包含三个典型的 AU 富含元件（AREs），AUUUA。有趣的是，这些 ARE 在几个 GDNF 基因中都得到了很好的保守。通过测试包含 3'-UTR 不同区域和长度与荧光素酶基因末端融合的报告基因构建体，我们证明了 ARE 诱导的荧光素酶活性降低与 mRNA 稳定性减弱相关。此外，我们发现 3'-UTR 中 ARE 周围的几个区域抑制了荧光素酶活性。此外，用含有 ARE 的 GDNF 表达载体转染 C6 神经胶质瘤细胞后，GDNF 蛋白的表达水平可忽略不计。
我们的研究首次描述了 3'-UTR 中的 ARE 和其他抑制区在调节 C6 细胞中 GDNF mRNA 含量方面的可能作用。
