Fluorescence-based quantification of nucleocytoplasmic transport
Fluorescence-based quantification of nucleocytoplasmic transport
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DOI：
10.1016/j.ymeth.2018.11.002
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发表时间：
2019-03-15
期刊：
影响因子：
4.8
通讯作者：
Paschal, Bryce M.
中科院分区：
文献类型：
--
作者：
Kelley, Joshua B.;Paschal, Bryce M.
DNA在膜结合的细胞核内的隔离是真核细胞的定义特征。因此，复制和转录仅限于细胞核，然而，这些事件的调节依赖于细胞质过程，包括蛋白质合成和信号转导途径。由于各种细胞活动都依赖于核转运，来自不同领域的研究人员发现研究感兴趣的蛋白质的核定位是有用的。在这里，我们提出了一些重要的技术考虑，研究细胞核和细胞质定位，并提供指导，定量蛋白质水平使用荧光显微镜和ImageJ软件。我们包括使用感兴趣的区域和图像分割的蛋白质定位的定量讨论。核质转运对于控制细胞核或细胞质中的蛋白质水平和活性至关重要，定量分析可以深入了解生物输出是如何实现的。
The sequestration of DNA within the membrane-bound nucleus is a defining characteristic of eukaryotic cells. Replication and transcription are therefore restricted to the nucleus, however, the regulation of these events relies on cytoplasmic processes including protein synthesis and signal transduction pathways. Because a variety of cellular activities depend on nuclear transport, researchers from diverse fields have found it useful to examine the nuclear localization of proteins of interest. Here we present some important technical considerations for studying nuclear and cytoplasmic localization, and provide guidance for quantifying protein levels using fluorescence microscopy and ImageJ software. We include discussion of the use of regions of interest and image segmentation for quantification of protein localization. Nucleocytoplasmic transport is fundamentally important for controlling protein levels and activity in the nucleus or cytoplasm, and quantitative analysis can provide insight into how biological output is achieved.