Graduate School of Pharmaceutical Sciences, Chiba University, 1-8-1 Inohana, Chuo-ku, Chiba 260-8675, Japan.
Department of Biochemistry & Molecular Biology, Kyoto Pharmaceutical University, Kyoto 607-8414, Japan.
Int J Mol Sci. 2022 Oct 14;23(20):12300. doi: 10.3390/ijms232012300.
The nucleolus is a non-membranous structure in the nucleus and forms around ribosomal DNA repeats. It plays a major role in ribosomal biogenesis through the transcription of ribosomal DNA and regulates mRNA translation in response to cellular stress including DNA damage. Rad17 is one of the proteins that initiate and maintain the activation of the ATR pathway, one of the major DNA damage checkpoints. We have recently reported that the central basic domain of Rad17 contains a nuclear localization signal and that the nuclear translocation of Rad17 promotes its proteasomal degradation. Here, we show that the central basic domain contains the nucleolar localization signal as well as the nuclear localization signal. The nucleolar localization signal overlaps with the nuclear localization signal and is capable of transporting an exogenous protein into the nucleolus. Phosphomimetic mutations of the central basic domain inhibit nucleolar accumulation, suggesting that the post-translational modification sites regulate the nucleolar localization. Nucleolar accumulation of Rad17 is promoted by proteasome inhibition and UV irradiation. Our data show the nucleolar localization of Rad17 and suggest a possible role of Rad17 in the nucleolus upon UV irradiation.
核仁是细胞核中的一种无膜结构，围绕核糖体 DNA 重复序列形成。它通过转录核糖体 DNA 在核糖体生物发生中发挥主要作用，并响应包括 DNA 损伤在内的细胞应激调节 mRNA 翻译。Rad17 是启动和维持 ATR 途径激活的蛋白质之一，ATR 途径是主要的 DNA 损伤检查点之一。我们最近报道称，Rad17 的中心碱性结构域包含核定位信号，并且 Rad17 的核转位促进其蛋白酶体降解。在这里，我们表明中心碱性结构域包含核仁定位信号和核定位信号。核仁定位信号与核定位信号重叠，并能够将外源蛋白运送到核仁中。中心碱性结构域的磷酸模拟突变抑制核仁积累，表明翻译后修饰位点调节核仁定位。蛋白酶体抑制和 UV 照射促进 Rad17 的核仁积累。我们的数据显示 Rad17 的核仁定位，并表明 Rad17 在 UV 照射时可能在核仁中发挥作用。
