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首页> 外文期刊>Nucleus >Differential staining of peripheral nuclear chromatin with Acridine orange implies an A-form epichromatin conformation of the DNA
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Differential staining of peripheral nuclear chromatin with Acridine orange implies an A-form epichromatin conformation of the DNA

机译：A啶橙对外周核染色质的差异染色意味着DNA的A型表观染色质构象

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The chromatin observed by conventional electron microscopy under the nuclear envelope constitutes a single layer of dense 30–35?nm granules, while ～30?nm fibrils laterally attached to them, form large patches of lamin-associated domains (LADs). This particular surface “epichromatin” can be discerned by specific (H2A+H2B+DNA) conformational antibody at the inner nuclear envelope and around mitotic chromosomes. In order to differentiate the DNA conformation of the peripheral chromatin we applied an Acridine orange (AO) DNA structural test involving RNAse treatment and the addition of AO after acid pre-treatment. MCF-7 cells treated in this way revealed yellow/red patches of LADs attached to a thin green nuclear rim and with mitotic chromosomes outlined in green, topologically corresponding to epichromatin epitope staining by immunofluorescence. Differentially from LADs, the epichromatin was unable to provide metachromatic staining by AO, unless thermally denatured at 94~(o)C. DNA enrichment in GC stretches has been recently reported for immunoprecipitated ～ 1Kb epichromatin domains. Together these data suggest that certain epichromatin segments assume the relatively hydrophobic DNA A-conformation at the nuclear envelope and surface of mitotic chromosomes, preventing AO side dimerisation.? We hypothesize that epichromatin domains form nucleosome superbeads. Hydrophobic interactions stack these superbeads and align them at the nuclear envelope, while repulsing the hydrophilic LADs. The hydrophobicity of epichromatin explains its location at the surface of mitotic chromosomes and its function in mediating chromosome attachment to the restituting nuclear envelope during telophase.
机译：通过常规电子显微镜在核膜下观察到的染色质构成了一层单层致密的30–35？nm颗粒，而横向附着于其上的〜30？nm的原纤维则形成了大片的层粘连蛋白相关区域（LAD）。这种特定的表面“上染素”可以通过内核被膜和有丝分裂染色体周围的特异性（H2A + H2B + DNA）构象抗体来识别。为了区分外周染色质的DNA构象，我们进行了cr啶橙（AO）DNA结构测试，涉及RNAse处理和酸预处理后添加AO。以这种方式处理的MCF-7细胞显示出LAD的黄色/红色斑块，该LAD附着在绿色核薄边缘上，并以绿色勾勒出有丝分裂染色体，在拓扑学上对应于通过免疫荧光染色的表观染色质表位。与LAD不同的是，除非在94℃下进行热变性，否则原染色质不能通过AO提供变色染色。最近报道了免疫沉淀〜1Kb表观染色质域中GC片段中的DNA富集。这些数据加在一起表明某些​​表观染色质片段在核膜和有丝分裂染色体表面具有相对疏水的DNA A构象，从而阻止了AO侧二聚化。我们假设表观染色质域形成核小体超级珠。疏水相互作用将这些超微珠堆叠在一起，并使它们在核被膜处对齐，同时排斥亲水性LAD。 Epichromatin的疏水性解释了其在有丝分裂染色体表面的位置及其在介导末期介导染色体与恢复核膜结合中的功能。

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