Poly(A) tail length-dependent stabilization of GAP-43 mRNA by the RNA-binding protein HuD
Poly(A) tail length-dependent stabilization of GAP-43 mRNA by the RNA-binding protein HuD
复制标题
DOI：
10.1074/jbc.m201982200
复制
发表时间：
2002-08-02
影响因子：
4.8
通讯作者：
Perrone-Bizzozero, NI
中科院分区：
文献类型：
--
作者：
Beckel-Mitchener, AC;Miera, A;Perrone-Bizzozero, NI
神经元类ELAV的RNA结合蛋白HuD与生长相关蛋白 - 43（GAP - 43）mRNA的3' - 非翻译区的一个调控元件结合。在此我们报道，在PC12细胞中HuD蛋白的过表达通过延迟mRNA降解的起始来稳定GAP - 43 mRNA，并且这一过程取决于多聚（A）尾的长度。利用基于多核糖体的体外mRNA降解实验，我们发现向系统中添加重组HuD蛋白增加了全长的、加帽的且多聚腺苷酸化的GAP - 43 mRNA的半衰期，并且这种效应部分是由于mRNA的去腺苷酸化速率降低所致。这种稳定作用对于在3' - 非翻译区包含HuD结合元件且具有至少150个A核苷酸的多聚（A）尾的GAP - 43 mRNA是特异性的。与HuD对GAP - 43 mRNA稳定性的影响相关，我们发现HuD结合长尾巴（A150）的GAP - 43 mRNA的亲和力比结合短尾巴（A30）的高10倍。我们得出结论，HuD通过一种依赖于多聚（A）尾长度且涉及对mRNA亲和力变化的机制来稳定GAP - 43 mRNA。
The neuronal ELAV-like RNA-binding protein HuD binds to a regulatory element in the 3'-untranslated region of the growth-associated protein-43 (GAP-43) mRNA. Here we report that overexpression of HuD protein in PC12 cells stabilizes the GAP-43 mRNA by delaying the onset of mRNA degradation and that this process depends on the size of the poly(A) tail. Using a polysomebased in vitro mRNA decay assay, we found that addition of recombinant HuD protein to the system increased the half-life of full-length, capped, and polyadenylated GAP-43 mRNA and that this effect was caused in part by a decrease in the rate of deadenylation of the mRNA. This stabilization was specific for GAP-43 mRNA containing the HuD binding element in the 3'-untranslated region and a poly(A) tail of at least 150 A nucleotides. In correlation with the effect of HuD on GAP-43 mRNA stability, we found that HuD binds GAP-43 mRNAs with long tails (A150) with 10-fold higher affinity than to those with short tails (A30). We conclude that HuD stabilizes the GAP-43 mRNA through a mechanism that is dependent on the length of the poly(A) tail and involves changes in its affinity for the mRNA.