Piwi 在果蝇卵巢体细胞中的核定位及其调控机制。

Piwi Nuclear Localization and Its Regulatory Mechanism in Drosophila Ovarian Somatic Cells.

机构信息

Department of Biological Sciences, Graduate School of Science, The University of Tokyo, Tokyo 113-0032, Japan.

Yokohama City University School of Medicine, Yokohama, Kanagawa 236-0004, Japan.

出版信息

Cell Rep. 2018 Jun 19;23(12):3647-3657. doi: 10.1016/j.celrep.2018.05.051.

DOI:10.1016/j.celrep.2018.05.051

PMID:29925005

Abstract

In Drosophila ovarian somatic cells (OSCs), Piwi represses transposons transcriptionally to maintain genome integrity. Piwi nuclear localization requires the N terminus and PIWI-interacting RNA (piRNA) loading of Piwi. However, the underlying mechanism remains unknown. Here, we show that Importinα (Impα) plays a pivotal role in Piwi nuclear localization and that Piwi has a bipartite nuclear localization signal (NLS). Impα2 and Impα3 are highly expressed in OSCs, whereas Impα1 is the least expressed. Loss of Impα2 or Impα3 forces Piwi to be cytoplasmic, which is rectified by overexpression of any Impα members. Extension of Piwi-NLS with an additional Piwi-NLS leads Piwi to be imported to the nucleus in a piRNA-independent manner, whereas replacement of Piwi-NLS with SV40-NLS fails. Limited proteolysis analysis suggests that piRNA loading onto Piwi triggers conformational change, exposing the N terminus to the environment. These results suggest that Piwi autoregulates its nuclear localization by exposing the NLS to Impα upon piRNA loading.

摘要

在果蝇卵巢体细胞（OSCs）中，Piwi 通过转录抑制转座子来维持基因组完整性。Piwi 的核定位需要其 N 端和 PIWI 相互作用 RNA（piRNA）的加载。然而，其潜在的机制仍不清楚。在这里，我们表明 Importinα（Impα）在 Piwi 的核定位中起着关键作用，并且 Piwi 具有双部分核定位信号（NLS）。Impα2 和 Impα3 在 OSCs 中高度表达，而 Impα1 的表达最少。Impα2 或 Impα3 的缺失会迫使 Piwi 细胞质定位，而过表达任何 Impα 成员都可以纠正这一现象。用额外的 Piwi-NLS 扩展 Piwi-NLS 会导致 Piwi 以 piRNA 非依赖的方式被导入细胞核，而用 SV40-NLS 替换 Piwi-NLS 则不行。有限蛋白酶分析表明，piRNA 加载到 Piwi 上会触发构象变化，使 N 端暴露在环境中。这些结果表明，Piwi 通过在 piRNA 加载时将 NLS 暴露给 Impα 来自我调节其核定位。