Muséum National d'Histoire Naturelle, CNRS UMR7196, INSERM U1154, 57 rue Cuvier, B.P. 26, 75231, Paris cedex 05, France.
Biochimie. 2018 Jun;149:122-134. doi: 10.1016/j.biochi.2018.03.008. Epub 2018 Apr 5.
Pericentromeric heterochromatin plays important roles in controlling gene expression and cellular differentiation. Fluorescent pyrrole-imidazole polyamides targeting murine pericentromeric DNA (major satellites) can be used for the visualization of pericentromeric heterochromatin foci in live mouse cells. New derivatives targeting human repeated DNA sequences (α-satellites) were synthesized and their interaction with target DNA was characterized. The possibility to use major satellite and α -satellite binding polyamides as tools for staining pericentromeric heterochromatin was further investigated in fixed and living mouse and human cells. The staining that was previously observed using the mouse model was further characterized and optimized, but remained limited regarding the fluorophores that can be used. The promising results regarding the staining in the mouse model could not be extended to the human model. Experiments performed in human cells showed chromosomal DNA staining without selectivity. Factors limiting the use of fluorescent polyamides, in particular probe aggregation in the cytoplasm, were investigated. Results are discussed with regards to structure and affinity of probes, density of target sites and chromatin accessibility in both models.
着丝粒异染色质在控制基因表达和细胞分化方面发挥着重要作用。针对鼠着丝粒 DNA（主要卫星）的荧光吡咯-咪唑聚酰胺可用于活鼠细胞着丝粒异染色质焦点的可视化。合成了针对人重复 DNA 序列（α-卫星）的新衍生物，并对其与靶 DNA 的相互作用进行了表征。进一步研究了主要卫星和α-卫星结合聚酰胺在固定和活的鼠和人细胞中作为着丝粒异染色质染色剂的可能性。以前在鼠模型中观察到的染色进一步进行了表征和优化，但在可以使用的荧光团方面仍然受到限制。在鼠模型中观察到的有希望的结果不能扩展到人类模型。在人细胞中进行的实验显示出没有选择性的染色体 DNA 染色。研究了限制荧光聚酰胺使用的因素，特别是在细胞质中探针聚集。结果根据探针的结构和亲和力、两种模型中靶位点的密度和染色质可及性进行了讨论。
