Department of Molecular Biotechnology and Microbiology, University of Debrecen, Life Sciences Building 1.102, 1 Egyetem Square, 4010 Debrecen, Hungary.
Int J Mol Sci. 2022 Aug 22;23(16):9503. doi: 10.3390/ijms23169503.
Eukaryotic chromosomes have not been visualized during the interphase. The fact that chromosomes cannot be seen during the interphase of the cell cycle does not mean that there are no means to make them visible. This work provides visual evidence that reversible permeabilization of the cell membrane followed by the regeneration of cell membranes allows getting a glimpse behind the nuclear curtain. Reversibly permeable eukaryotic cells have been used to synthesize nascent DNA, analyze the 5'-end of RNA primers, view individual replicons and visualize interphase chromosomes. Dextran T-150 in a slightly hypotonic buffer prevented cells from disruption. Upon reversal of permeabilization, the nucleus could be opened at any time during the interphase. A broad spectrum of a flexible chromatin folding pattern was revealed through a series of transient geometric forms of chromosomes. Linear attachment of chromosomes was visualized in several mammalian and lower eukaryotic cells. The linear connection of chromosomes is maintained throughout the cell cycle showing that rather than individual chromosomes, a linear array of chromosomes is the functional giant macromolecule. This study proves that not only the prokaryotic genome but also linearly attached eukaryotic chromosomes form a giant macromolecular unit.
真核染色体在间期没有被观察到。在细胞周期的间期看不到染色体这一事实并不意味着没有方法使它们可见。这项工作提供了直观的证据，证明细胞膜的可逆通透化，随后细胞膜的再生，使得能够瞥见核膜的后面。可逆通透的真核细胞已被用于合成新生 DNA、分析 RNA 引物的 5'-末端、观察单个复制子和可视化间期染色体。在稍低渗的缓冲液中的葡聚糖 T-150 可防止细胞破裂。在通透化逆转后，核可以在间期的任何时间打开。通过一系列染色体的瞬态几何形态，揭示了广泛的灵活染色质折叠模式。在几种哺乳动物和低等真核细胞中观察到染色体的线性附着。染色体的线性连接贯穿整个细胞周期，表明线性连接的染色体是功能上的巨大大分子，而不是单个染色体。这项研究证明，不仅原核基因组，而且线性附着的真核染色体也形成一个巨大的大分子单元。
