Finding out how cells prepare for fate change during differentiation commitment was our task. To address if the constitutive pericentromere associated domains (PADs) may be involved, we used a model system with known transcriptome data, MCF-7 breast cancer cells treated with the ErbB3 ligand heregulin (HRG), which induces differentiation and is used in the therapy of cancer. PAD-repressive heterochromatin (H3K9me3), centromere (CENPA)-specific, and active euchromatin (H3K4me3) antibodies, qPCR, acridine-orange DNA structural test (AOT), and microscopic image analysis were applied. We found a two-step DNA unfolding, after 15-20 min and 60 min HRG treatment, respectively. This behaviour was consistent with biphasic activation of the early response genes (c-fos - fosL1/myc) and the timing of two transcriptome avalanches reported in the literature. In control, the average number of PADs negatively correlated with their size by scale-free distribution, centromere clustering in turn correlated with PAD size, both indicating that PADs may create and modulate a supra-chromosomal network by fusing and splitting a constant proportion of the constitutive heterochromatin. By 15 min HRG treatment, the bursting unravelling of PADs from the nucleolus boundary occurred, coinciding with the first step of H3K4me3 chromatin unfolding confirmed by AOT. The second step after 60 min HRG treatment was associated with transcription of long-non-coding-RNA from PADs and peaking of fosL1/c-myc response. We hypothesize that bursting of PAD clusters under a critical silencing threshold pushes first transcription avalanche, while destruction of the PAD network enables genome rewiring needed for differentiation re-patterning, mediated by early response bivalent genes.

中文翻译：

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分化癌细胞揭示了中心域的早期大规模基因组调控

找出细胞在分化过程中如何为命运变化做准备是我们的任务。为了解决是否可能涉及组成型着丝粒周围相关结构域 (PAD)，我们使用了具有已知转录组​​数据的模型系统，用 ErbB3 配体调蛋白 (HRG) 处理的 MCF-7 乳腺癌细胞，其诱导分化并用于治疗癌症。应用了 PAD 抑制性异染色质 (H3K9me3)、着丝粒 (CENPA) 特异性和活性常染色质 (H3K4me3) 抗体、qPCR、吖啶橙 DNA 结构测试 (AOT) 和显微图像分析。我们发现两步 DNA 展开，分别在 15-20 分钟和 60 分钟 HRG 处理后。这种行为与早期反应基因 (c-fos - fosL1/myc) 的双相激活和文献中报道的两个转录组雪崩的时间一致。在对照中，PAD 的平均数量通过无标度分布与其大小呈负相关，着丝粒聚类反过来又与 PAD 大小相关，两者都表明 PAD 可以通过融合和分裂恒定比例的组成型异染色质。通过 15 分钟的 HRG 处理，PAD 从核仁边界突然解开，这与 AOT 证实的 H3K4me3 染色质展开的第一步一致。HRG 处理 60 分钟后的第二步与 PAD 中长链非编码 RNA 的转录和 fosL1/c-myc 反应的峰值有关。我们假设在临界沉默阈值下 PAD 簇的爆发会推动第一次转录雪崩，