Chemogenetic control of protein localization and mammalian cell signaling by SLIPT
Chemogenetic control of protein localization and mammalian cell signaling by SLIPT
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SLIPT 对蛋白质定位和哺乳动物细胞信号传导的化学遗传学控制
DOI：
10.1007/978-1-0716-1441-9_14
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发表时间：
2021
期刊：
影响因子：
--
通讯作者：
Shinya Tsukiji
中科院分区：
文献类型：
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作者：
Sachio Suzuki;Yuka Hatano;Tatsuyuki Yoshii;Shinya Tsukiji
蛋白质定位的化学控制是操纵哺乳动物细胞过程的有力方法。自定位配体诱导的蛋白质易位（SLIPT）是一种新兴的平台，其使得能够使用合成的自定位配体（SL）控制活哺乳动物细胞中的蛋白质定位。我们最近建立了一个化学发生SLIPT系统，其中任何感兴趣的蛋白质融合到大肠杆菌二氢叶酸还原酶的工程变体DHFRiK 6，可以快速和特异性地从细胞质转运到质膜（PM）的内小叶，使用甲氧苄啶（TMP）为基础的PM靶向SL，mDcTMP。通过随后添加游离TMP，可以将DHFRiK 6融合蛋白的mDcTMP介导的PM募集有效地返回到细胞质中，从而能够对蛋白定位进行时间和可逆的控制。在这里，我们描述了这种基于mDcTMP/DHFRiK 6的SLIPT系统用于诱导（1）可逆蛋白质易位和（2）Raf/ERK途径的合成活化的用途。该系统为哺乳动物合成生物学提供了一种简单而通用的工具，用于在PM上暂时操纵各种信号分子和途径。
Chemical control of protein localization is a powerful approach for manipulating mammalian cellular processes. Self-localizing ligand-induced protein translocation (SLIPT) is an emerging platform that enables control of protein localization in living mammalian cells using synthetic self-localizing ligands (SLs). We recently established a chemogenetic SLIPT system, in which any protein of interest fused to an engineered variant ofEscherichia colidihydrofolate reductase, DHFRiK6, can be rapidly and specifically translocated from the cytoplasm to the inner leaflet of the plasma membrane (PM) using a trimethoprim (TMP)-based PM-targeting SL, mDcTMP. The mDcTMP-mediated PM recruitment of DHFRiK6-fusion proteins can be efficiently returned to the cytoplasm by subsequent addition of free TMP, enabling temporal and reversible control over the protein localization. Here we describe the use of this mDcTMP/DHFRiK6-based SLIPT system for inducing (1) reversible protein translocation and (2) synthetic activation of the Raf/ERK pathway. This system provides a simple and versatile tool in mammalian synthetic biology for temporally manipulating various signaling molecules and pathways at the PM.