N6-Allyladenosine: A New Small Molecule for RNA Labeling Identified by Mutation Assay,Journal of the American Chemical Society

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N6-Allyladenosine: A New Small Molecule for RNA Labeling Identified by Mutation Assay
Journal of the American Chemical Society ( IF 14.4 ) Pub Date : 2017-11-15 00:00:00 , DOI: 10.1021/jacs.7b06837
Xiao Shu ₁ , Qing Dai ₂ , Tong Wu ₂ , Ian R. Bothwell ₃ , Yanan Yue ₁ , Zezhou Zhang ₁ , Jie Cao ₁ , Qili Fei ₂ , Minkui Luo ₃ , Chuan He ₂ , Jianzhao Liu _{1,

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Affiliation

RNA labeling is crucial for the study of RNA structure and metabolism. Herein we report N⁶-allyladenosine (a⁶A) as a new small molecule for RNA labeling through both metabolic and enzyme-assisted manners. a⁶A behaves like A and can be metabolically incorporated into newly synthesized RNAs inside mammalian cells. We also show that human RNA N⁶-methyladenosine (m⁶A) methyltransferases METTL3/METTL14 can work with a synthetic cofactor, namely allyl-SAM (S-adenosyl methionine with methyl replaced by allyl) in order to site-specifically install an allyl group to the N⁶-position of A within specific sequence to generate a⁶A-labeled RNAs. The iodination of N⁶-allyl group of a⁶A under mild buffer conditions spontaneously induces the formation of N¹,N⁶-cyclized adenosine and creates mutations at its opposite site during complementary DNA synthesis of reverse transcription. The existing m⁶A in RNA is inert to methyltransferase-assisted allyl labeling, which offers a chance to differentiate m⁶A from A at individual RNA sites. Our work demonstrates a new method for RNA labeling, which could find applications in developing sequencing methods for nascent RNAs and RNA modifications.

中文翻译：

N ^6-烯丙基腺苷：一种新的小分子，用于通过突变测定鉴定RNA的标签。

RNA标记对于RNA结构和代谢的研究至关重要。在这里，我们报道N ^6-烯丙基腺苷（a ⁶ A）作为通过代谢和酶辅助方式进行RNA标记的新小分子。a ⁶ A的行为类似于A，可以通过代谢方式整合到哺乳动物细胞内部新合成的RNA中。我们还显示，人RNA N ^6-甲基腺苷（m ⁶ A）甲基转移酶METTL3 / METTL14可以与合成辅因子，即烯丙基-SAM （甲基取代为烯丙基的S-腺苷甲硫氨酸）结合使用，以便于位点特异性地安装烯丙基。结合到特定序列内A的N ⁶位以生成一个⁶ A标记的RNA。在温和的缓冲液条件下，一个⁶ A的N ⁶烯丙基的碘化可自发诱导N ¹，N ⁶环化的腺苷的形成，并在互补的逆转录DNA合成过程中在其相反位点产生突变。RNA中现有的m ⁶ A对甲基转移酶辅助的烯丙基标记是惰性的，这为在单个RNA位点上将m ⁶ A与A区分提供了机会。我们的工作展示了一种用于RNA标记的新方法，该方法可以在开发新生RNA和RNA修饰的测序方法中找到应用。

更新日期：2017-11-16

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