European Molecular Biology Laboratory, Grenoble Outstation, 38042 Grenoble, France Unit for Virus Host-Cell Interactions, University of Grenoble Alpes-EMBL-CNRS, 38042 Grenoble, France.
Department of Oncological Sciences, Icahn School of Medicine at Mt. Sinai, New York, New York 10029, USA.
RNA. 2014 Jun;20(6):773-81. doi: 10.1261/rna.044701.114. Epub 2014 Apr 22.
Piwi-interacting RNAs (piRNAs) guide Piwi Argonautes to suppress transposon activity in animal gonads. Known piRNA populations are extremely complex, with millions of individual sequences present in a single organism. Despite this complexity, specific Piwi proteins incorporate piRNAs with distinct nucleotide- and transposon strand-biases (antisense or sense) of unknown origin. Here, we examined the contribution of structural domains in Piwi proteins toward defining these biases. We report the first crystal structure of the MID domain from a Piwi Argonaute and use docking experiments to show its ability to specify recognition of 5' uridine (1U-bias) of piRNAs. Mutational analyses reveal the importance of 5' end-recognition within the MID domain for piRNA biogenesis in vivo. Finally, domain-swapping experiments uncover an unexpected role for the MID-PIWI module of a Piwi protein in dictating the transposon strand-orientation of its bound piRNAs. Our work identifies structural features that allow distinguishing individual Piwi members during piRNA biogenesis.
Piwi 相互作用 RNA（piRNAs）指导 Piwi Argonautes 抑制动物性腺中转座子的活性。已知的 piRNA 群体极其复杂，在单个生物体中存在着数百万个单独的序列。尽管如此复杂，但特定的 Piwi 蛋白却将具有独特核苷酸和转座子链偏倚（反义或正义）的 piRNA 整合在一起，其来源未知。在这里，我们研究了 Piwi 蛋白结构域对定义这些偏倚的贡献。我们报告了第一个 Piwi Argonaute 的 MID 结构域的晶体结构，并通过对接实验表明其能够特异性识别 piRNA 的 5' 尿嘧啶（1U 偏倚）。突变分析揭示了 MID 结构域内 5' 端识别对于体内 piRNA 生物发生的重要性。最后，结构域交换实验揭示了 Piwi 蛋白的 MID-PIWI 模块在决定其结合的 piRNA 转座子链取向方面的意外作用。我们的工作确定了在 piRNA 生物发生过程中区分单个 Piwi 成员的结构特征。
