Structure and function of Zucchini endoribonuclease in piRNA biogenesis
Structure and function of Zucchini endoribonuclease in piRNA biogenesis
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DOI：
10.1038/nature11509
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发表时间：
2012-11-08
期刊：
影响因子：
64.8
通讯作者：
Nureki, Osamu
中科院分区：
文献类型：
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作者：
Nishimasu, Hiroshi;Ishizu, Hirotsugu;Nureki, Osamu
Piwi相互作用RNA(PiRNAs)沉默转座子以维持动物生殖系基因组的完整性(1-4)。根据其生物发生机制，piRNAs被分为初级和次级piRNAs(5-10)。初级piRNA是由从基因组中的piRNA簇转录而来的长非编码RNA前体通过初级加工途径(5，8-10)加工而成的。虽然已经预测了参与这一途径的核糖核酸酶的存在，但其分子身份仍不清楚。在这里，我们发现西葫芦(Zucchini，Zuc)是线粒体磷脂酶D(PLD)超家族成员(11)，是初级piRNA生物发生所必需的内切核酸酶。我们在1.75埃分辨率下解析了果蝇的晶体结构。结构表明，DmZuc在二聚体界面上有一个带正电的窄催化槽，可以容纳单链RNA，但不能容纳双链RNA。DmZuc和小鼠同系物MmZuc(也称为Pld6和MitoPLD)(12-14)在体外显示了单链RNA的内切核酸酶活性。RNA裂解产物带有一个5‘-单磷酸基团，这是成熟的piRNAs的标志。突变分析表明，DmZuc保守的活性位点残基对体外核糖核酸酶活性、体内piRNA成熟和转座子沉默至关重要。我们提出了一个动物胚系中piRNA生物发生的模型，其中Zuc内切核酸酶在初级piRNA成熟过程中起着关键作用。
PIWI-interacting RNAs (piRNAs) silence transposons to maintain genome integrity in animal germ lines(1-4). piRNAs are classified as primary and secondary piRNAs, depending on their biogenesis machinery(5-10). Primary piRNAs are processed from long non-coding RNA precursors transcribed from piRNA clusters in the genome through the primary processing pathway(5,8-10). Although the existence of a ribonuclease participating in this pathway has been predicted, its molecular identity remained unknown. Here we show that Zucchini (Zuc), a mitochondrial phospholipase D (PLD) superfamily member(11), is an endoribonuclease essential for primary piRNA biogenesis. We solved the crystal structure of Drosophila melanogaster Zuc (DmZuc) at 1.75 angstrom resolution. The structure revealed that DmZuc has a positively charged, narrow catalytic groove at the dimer interface, which could accommodate a single-stranded, but not a double-stranded, RNA. DmZuc and the mouse homologue MmZuc (also known as Pld6 and MitoPLD)(12-14) showed endoribonuclease activity for single-stranded RNAs in vitro. The RNA cleavage products bear a 5'-monophosphate group, a hallmark of mature piRNAs. Mutational analyses revealed that the conserved active-site residues of DmZuc are critical for the ribonuclease activity in vitro, and for piRNA maturation and transposon silencing in vivo. We propose a model for piRNA biogenesis in animal germ lines, in which the Zuc endoribonuclease has a key role in primary piRNA maturation.