Department of Developmental Biology, Sloan-Kettering Institute, New York, New York, USA.
Nat Struct Mol Biol. 2013 Jul;20(7):789-95. doi: 10.1038/nsmb.2606. Epub 2013 May 26.
Homeostatic mechanisms regulate the abundance of several components in small-RNA pathways. We used Drosophila and mammalian systems to demonstrate a conserved homeostatic system in which the status of miRNA biogenesis controls Argonaute protein stability. Clonal analyses of multiple mutants of core Drosophila miRNA factors revealed that stability of the miRNA effector AGO1 is dependent on miRNA biogenesis. Reciprocally, ectopic transcription of miRNAs within in vivo clones induced accumulation of AGO1, as did genetic interference with the ubiquitin-proteasome system. In mouse cells, we found that the stability of Ago2 declined in Dicer-knockout cells and was rescued by proteasome blockade or introduction of either Dicer plasmid or Dicer-independent miRNA constructs. Notably, Dicer-dependent miRNA constructs generated pre-miRNAs that bound Ago2 but did not rescue Ago2 stability. We conclude that Argonaute levels are finely tuned by cellular availability of mature miRNAs and the ubiquitin-proteasome system.
内稳态机制调节小 RNA 通路中几种成分的丰度。我们使用果蝇和哺乳动物系统证明了一个保守的内稳态系统，其中 miRNA 生物发生的状态控制 Argonaute 蛋白稳定性。多个核心果蝇 miRNA 因子的克隆分析表明，miRNA 效应物 AGO1 的稳定性依赖于 miRNA 的生物发生。反过来，体内克隆中 miRNA 的异位转录诱导 AGO1 的积累，泛素-蛋白酶体系统的遗传干扰也是如此。在小鼠细胞中，我们发现 Dicer 敲除细胞中 Ago2 的稳定性下降，蛋白酶体阻断或引入 Dicer 质粒或 Dicer 非依赖性 miRNA 构建体可挽救其稳定性。值得注意的是，Dicer 依赖性 miRNA 构建体生成与 Ago2 结合但不能挽救 Ago2 稳定性的 pre-miRNA。我们得出结论，Argonaute 水平受到成熟 miRNA 和泛素-蛋白酶体系统的细胞可用性的精细调节。
