Nucleo-cytoplasmic shuttling of Id2, a negative regulator of basic helix-loop-helix transcription factors
Nucleo-cytoplasmic shuttling of Id2, a negative regulator of basic helix-loop-helix transcription factors
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DOI：
10.1074/jbc.m412614200
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发表时间：
2005-02-11
影响因子：
4.8
通讯作者：
Yokota, Y
中科院分区：
文献类型：
--
作者：
Kurooka, H;Yokota, Y
Id蛋白作为碱性螺旋-环-螺旋转录因子的负调控因子，在细胞命运决定中发挥重要作用。它们优先与碱性螺旋-环-螺旋家族的普遍表达的E蛋白结合，并阻止它们与DNA结合并激活转录。虽然它们的小尺寸表明Id蛋白通过被动扩散进入和离开细胞核，但一些研究表明其他途径可能调节它们的亚细胞定位。在本研究中，我们获得了Id 2具有在细胞核和细胞质之间穿梭的能力的证据。当通过与绿色荧光蛋白（GFP）融合来防止被动扩散时，Id 2主要定位于细胞质中。使用GFP融合构建体，我们证明了C-末端区域是细胞质定位所需的。用核输出抑制剂来普霉素B处理的细胞中GFP-Id 2的核积累表明核输出受体染色体区域维持蛋白1介导Id 2的细胞质定位。Id 2含有两个假定的富含亮氨酸的核输出信号，并且在C-末端区域的核输出信号是核输出所必需的。另一方面，螺旋-环-螺旋结构域对于核定位是重要的。最后，使用报告分析的实验揭示了核出口和转录抑制通过E盒序列之间的负相关性。基于所有这些发现，我们提出，核质穿梭是一种新的机制，调节Id 2的功能。
Id proteins function as negative regulators for basic helix-loop-helix transcriptional factors that play important roles in cell fate determination. They preferentially associate with ubiquitously expressed E proteins of the basic helix-loop-helix family and prevent them from binding to DNA and activating transcription. Although their small size suggests that Id proteins enter and exit the nucleus by passive diffusion, several studies have indicated that other pathways may regulate their subcellular localization. In this study, we obtained evidence that Id2 has the ability to shuttle between the nucleus and the cytoplasm. When passive diffusion was prevented by fusion with green fluorescent protein (GFP), Id2 was predominantly localized in the cytoplasm. Using GFP fusion constructs, we demonstrated that the C-ter-minal region is required for cytoplasmic localization. Nuclear accumulation of GFP-Id2 in cells treated with the nuclear export inhibitor leptomycin B suggests that the nuclear export receptor chromosome region maintenance protein 1 mediates the cytoplasmic localization of Id2. Id2 contains two putative leucine-rich nuclear export signals, and the nuclear export signal in the C-terminal region is essential for nuclear export. On the other hand, the helix-loop-helix domain is important for nuclear localization. Finally, experiments using reporter assays revealed an inverse correlation between nuclear export and transcriptional repression via the E-box sequence. Based on all these findings, we propose that nucleo-cytoplasmic shuttling is a novel mechanism for the regulation of Id2 function.