Sato Yuko, Stasevich Timothy J, Kimura Hiroshi
Cell Biology Center, Institute of Innovative Research, Tokyo Institute of Technology, Yokohama, Japan.
World Research Hub Initiative, Institute of Innovative Research, Tokyo Institute of Technology, Yokohama, Japan.
Methods Mol Biol. 2018;1861:91-102. doi: 10.1007/978-1-4939-8766-5_8.
The inactive X chromosome (Xi) harbors characteristic epigenetic features, including the enrichment of histone H3 lysine 27 trimethylation (H3K27me3) and H4 lysine 20 monomethylation (H4K20me1) as well as a lack of histone acetylation. Recently, these modifications have been visualized not only in fixed specimen, but also in living cells via probes derived from modification-specific antibodies. The probes include fluorescently labeled antigen binding fragments (Fabs), which can be loaded into cells, as well as genetically encoded single-chain variable fragments tagged with the green fluorescent protein. We refer to the latter as modification specific intracellular antibodies, or "mintbodies" for short. By using Fabs or mintbodies to target Xi-specific modifications, the dynamics of Xi in living cells can be visualized.
失活的X染色体(Xi)具有特征性的表观遗传特征,包括组蛋白H3赖氨酸27三甲基化(H3K27me3)和H4赖氨酸20一甲基化(H4K20me1)的富集以及组蛋白乙酰化的缺乏。最近,这些修饰不仅在固定标本中得以可视化,还通过源自修饰特异性抗体的探针在活细胞中实现了可视化。这些探针包括可加载到细胞中的荧光标记抗原结合片段(Fabs),以及标记有绿色荧光蛋白的基因编码单链可变片段。我们将后者称为修饰特异性细胞内抗体,或简称为“薄荷体”。通过使用Fabs或薄荷体靶向Xi特异性修饰,可以在活细胞中可视化Xi的动态变化。