Gene Center and Department of Biochemistry, Ludwig-Maximilians-Universität München, Feodor-Lynen-Str. 25, 81377 Munich, Germany.
Nucleic Acids Res. 2011 May;39(9):3836-51. doi: 10.1093/nar/gkq1324. Epub 2011 Jan 17.
In Drosophila, siRNAs are classified as endo- or exo-siRNAs based on their origin. Both are processed from double-stranded RNA precursors by Dcr-2 and then loaded into the Argonaute protein Ago2. While exo-siRNAs serve to defend the cell against viruses, endo-siRNAs restrict the spread of selfish DNA in somatic cells, analogous to piRNAs in the germ line. Endo- and exo-siRNAs display a differential requirement for double-stranded RNA binding domain proteins (dsRBPs): R2D2 is needed to load exo-siRNAs into Ago2 while the PD isoform of Loquacious (Loqs-PD) stimulates Dcr-2 during the nucleolytic processing of hairpin-derived endo-siRNAs. In cell culture assays, R2D2 antagonizes Loqs-PD in endo-siRNA silencing and Loqs-PD is an inhibitor of RNA interference. Loqs-PD can interact via the C-terminus unique to this isoform with the DExH/D-helicase domain of Drosophila Dcr-2, where binding of R2D2 has also been localized. Separation of the two pathways is not complete; rather, the dicing and Ago2-loading steps appear uncoupled, analogous to the corresponding steps in miRNA biogenesis. Analysis of deep sequencing data further demonstrates that in r2d2 mutant flies, siRNAs can be loaded into Ago2 but not all siRNA classes are equally proficient for this. Thus, the canonical Ago2-RISC loading complex can be bypassed under certain circumstances.
在果蝇中，根据其来源，siRNA 被分为内源性或外源性 siRNA。两者均由 Dcr-2 从双链 RNA 前体加工而来，然后装载到 Argonaute 蛋白 Ago2 中。外源性 siRNA 可用于保护细胞免受病毒侵害，而内源性 siRNA 则限制了自私 DNA 在体细胞中的传播，类似于生殖系中的 piRNA。内源性和外源性 siRNA 对双链 RNA 结合域蛋白（dsRBPs）的需求存在差异：R2D2 用于将外源性 siRNA 装载到 Ago2 中，而 Loquacious 的 PD 同工型（Loqs-PD）在发夹衍生的内源性 siRNA 的核酶加工过程中刺激 Dcr-2。在细胞培养测定中，R2D2 在内源性 siRNA 沉默中拮抗 Loqs-PD，而 Loqs-PD 是 RNA 干扰的抑制剂。Loqs-PD 可以通过其独特的 C 末端与果蝇 Dcr-2 的 DExH/D-解旋酶结构域相互作用，R2D2 的结合也已定位在此结构域。两条途径的分离并不完全；相反，切割和 Ago2 装载步骤似乎是解耦的，类似于 miRNA 生物发生的相应步骤。对深度测序数据的分析进一步表明，在 r2d2 突变体果蝇中，siRNA 可以装载到 Ago2 中，但并非所有 siRNA 类都同样擅长于此。因此，在某些情况下，可以绕过规范的 Ago2-RISC 装载复合物。
