Translational Repression of the Disintegrin and Metalloprotease ADAM10 by a Stable G-quadruplex Secondary Structure in Its 5′-Untranslated Region
Translational Repression of the Disintegrin and Metalloprotease ADAM10 by a Stable G-quadruplex Secondary Structure in Its 5′-Untranslated Region
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DOI：
10.1074/jbc.m111.296921
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发表时间：
2011-12-30
影响因子：
4.8
通讯作者：
Haass, Christian
中科院分区：
文献类型：
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作者：
Lammich, Sven;Kamp, Frits;Haass, Christian
α-分泌酶对淀粉样前体蛋白APP的抗淀粉样蛋白生成加工防止了淀粉样β肽的形成，淀粉样β肽在阿尔茨海默病患者的老年斑中积累。α-分泌酶属于解整合素和金属蛋白酶（亚当斯）家族，并且ADAM 10是这种抗淀粉样蛋白生成活性的主要候选物。我们最近证明了ADAM 10的翻译被其5 '-UTR抑制，特别是ADAM 10 5'-UTR的前半部分负责翻译抑制。在这里，我们询问在ADAM 10 5 '-UTR中是否存在能够形成复杂二级结构从而潜在地抑制ADAM 10翻译的特定序列基序。使用圆二色光谱，我们证明了在ADAM 10 5 '-UTR的核苷酸66和94之间的富含G的区域在生理条件下形成高度稳定的分子内平行G-四链体二级结构。该序列中鸟嘌呤的突变消除了G-四链体结构的形成。尽管G-四链体结构在体外翻译测定和活细胞中有效地抑制荧光素酶报告基因的翻译，但G-四链体形成的抑制未能做到这一点。此外，ADAM 10的表达同样受到G-四链体的抑制。G-四链体基序的突变导致ADAM 10水平显著增加，从而导致APP α分泌显著增加。因此，我们确定了5 '-UTR内的关键RNA二级结构，其有助于ADAM 10的翻译抑制。
Anti-amyloidogenic processing of the amyloid precursor protein APP by alpha-secretase prevents formation of the amyloid-beta peptide, which accumulates in senile plaques of Alzheimer disease patients. alpha-Secretase belongs to the family of a disintegrin and metalloproteases (ADAMs), and ADAM10 is the primary candidate for this anti-amyloidogenic activity. We recently demonstrated that ADAM10 translation is repressed by its 5'-UTR and that in particular the first half of ADAM10 5'-UTR is responsible for translational repression. Here, we asked whether specific sequence motifs exist in the ADAM10 5'-UTR that are able to form complex secondary structures and thus potentially inhibit ADAM10 translation. Using circular dichroism spectroscopy, we demonstrate that a G-rich region between nucleotides 66 and 94 of the ADAM10 5'-UTR forms a highly stable, intramolecular, parallel G-quadruplex secondary structure under physiological conditions. Mutation of guanines in this sequence abrogates the formation of the G-quadruplex structure. Although the G-quadruplex structure efficiently inhibits translation of a luciferase reporter in in vitro translation assays and in living cells, inhibition of G-quadruplex formation fails to do so. Moreover, expression of ADAM10 was similarly repressed by the G-quadruplex. Mutation of the G-quadruplex motif results in a significant increase of ADAM10 levels and consequently APPs alpha secretion. Thus, we identified a critical RNA secondary structure within the 5'-UTR, which contributes to the translational repression of ADAM10.