Department of Molecular Biology, Keio University School of Medicine, Tokyo 160-8582, Japan.
Mol Cell. 2013 Feb 21;49(4):680-91. doi: 10.1016/j.molcel.2012.12.024. Epub 2013 Jan 31.
Endogenous small interfering RNAs (endo-siRNAs) in Drosophila are processed by Dicer-2 (Dcr-2) and loaded onto Ago2 by the Dcr-2/R2D2 heterodimer. In r2d2 mutants, the level of endo-siRNAs is unchanged, but endo-siRNAs are misloaded onto Ago1. However, the mechanism underlying the control of endo-siRNA sorting by R2D2 remains unknown. Here, we show that R2D2 controls endo-siRNA sorting by localizing Dcr-2, and presumably endo-siRNA duplexes, to cytoplasmic foci, D2 bodies. Ago2, but not Ago1, localized to D2 bodies. dsRNA-binding-deficient mutant, but not wild-type, R2D2 failed to localize D2 bodies and caused endo-siRNA misdirection to Ago1 in R2D2-depleted cells. However, R2D2 was dispensable for sorting miRNAs and exogenous siRNAs onto Ago1 and Ago2, respectively, in vivo. Endo- and exo-siRNA guide selection also occurred R2D2 independently. The functions of R2D2 are required to avoid endo-siRNA misdirection to Ago1, because Ago1 is capable of loading incompletely complementary miRNA duplexes and endo-siRNA duplexes.
果蝇内源性小干扰 RNA (endo-siRNAs) 由 Dicer-2 (Dcr-2) 加工，并由 Dcr-2/R2D2 异二聚体加载到 Ago2 上。在 r2d2 突变体中，endo-siRNAs 的水平不变，但 endo-siRNAs 错误地加载到 Ago1 上。然而，R2D2 控制 endo-siRNA 分类的机制尚不清楚。在这里，我们表明 R2D2 通过将 Dcr-2 及其推测的 endo-siRNA 双链体定位到细胞质焦点 D2 体上来控制 endo-siRNA 的分类。Ago2，但不是 Ago1，定位到 D2 体。dsRNA 结合缺陷突变体，而不是野生型，R2D2 未能定位 D2 体，并导致 R2D2 耗尽细胞中 endo-siRNA 错误导向 Ago1。然而，R2D2 在体内分别将 miRNAs 和外源性 siRNAs 分类到 Ago1 和 Ago2 上是可有可无的。内源性和外源性 siRNA 引导选择也独立于 R2D2 发生。R2D2 的功能对于避免 endo-siRNA 错误导向 Ago1 是必需的，因为 Ago1 能够加载不完全互补的 miRNA 双链体和 endo-siRNA 双链体。
