Although many protein labeling probes have been developed to elucidate the trafficking and turnover processes of cell surface proteins, real-time tracking of intracellular proteins remains a challenging task. Herein, we describe a new design to construct a cell-permeable, photostable, and far-red fluorescent turn-on probe to enable no-wash, organelle-specific, and long-term visualization of intracellular SNAP-tagged proteins in living cells. When the probe was used in dual-color pulse chase labeling experiments to differentiate between preLamin and mature Lamin, our results reveal that the shape of mature Lamin can be altered by the newly synthesized preLamin and that this alteration is progressive, cumulative, and due to a concentration-dependent dominant-negative effect of preLamin. We believe that this probe can also be applied to other intracellular proteins whose cellular localization and synthesis changes dynamically in response to external stimuli.

中文翻译：

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荧光蛋白标记探针，用于研究前Lamin和成熟Lamin之间的形态相互作用

尽管已经开发出许多蛋白质标记探针来阐明细胞表面蛋白质的运输和更新过程，但是实时跟踪细胞内蛋白质仍然是一项艰巨的任务。在这里，我们描述了一种新的设计，以构建一种细胞可渗透的，光稳定的和远红色的荧光开启探针，以实现对活细胞中细胞内SNAP标记蛋白的免冲洗，细胞器特异性和长期可视化。当将探针用于双色脉冲追逐标记实验中以区分preLamin和成熟Lamin时，我们的结果表明，新合成的preLamin可以改变成熟Lamin的形状，并且这种改变是渐进的，累积的，并且是由于preLamin的浓度依赖性显性负效应。