PO.MCB06.01 · 分子与细胞生物学
+1 核小体稳定性与 DNA 超螺旋之间的关系
The relationships between the stability of the +1 nucleosome and DNA superhelicity
作者与单位 Authors & Affiliations
摘要 Abstract
中文摘要
在一项涉及琼脂糖包埋细胞核的原位分析中，比较了各种组蛋白尾部翻译后修饰（PTM）对核小体稳定性的影响，结果发现启动子近端 H3K4me3、H3K27ac 阳性核小体的一个亚群相较于整体 H3-GFP 或携带以下任一标记的核小体表现出对嵌入剂的相对敏感性：H3K27me1、H3K27me2、H3K27me3、H3K9me1、H3K9me2、H3K9me3、H3K36me3、H3K4me0、H3K4me1、H3K4me2、H3K9ac 和 H3K14ac。对细胞核进行切口酶或 DNase I 处理，或对活细胞进行博来霉素处理，均不影响携带 H3K4me3 或 H3K27ac 的核小体的稳定性，而第二组核小体在嵌入剂处理下均被去稳定化。这些观察结果支持以下可能性：指定动态核小体的启动子近端标记与因体内产生的 DNA 断裂而松弛的 DNA 序列并置。与此解释一致，在人单个核细胞以及小鼠胚胎干细胞（mES）中，内源性 3'OH 切口被定位于启动子的核小体游离区内。我们还提供证据表明，含有断裂的染色质区域在拓扑上与含有超螺旋染色质的结构域相分离。关于引发断裂的机制，我们使用敲除细胞、抑制剂和降解子系统，基于 TOP2 活性或基于组蛋白去甲基化诱导的氧化性 DNA 损伤及其修复，实验测试了两种备选模型。这些观察结果支持这样一个模型：DNA 链不连续性在转录调控和高阶染色质组织中的作用得到整合。基金：OTKA/NKFIH 138524，ERA-Net NEURON 2024-1.2.2-ERA_NET-2024-00009 *PN 和 LI 贡献相同。
查看英文原文 English abstract
When the effect of various posttranslational histone tail modifications (PTMs) on nucleosome stability was compared in an in situ assay involving agarose-embedded nuclei, a subpopulation of the promoter proximal H3K4me3, H3K27ac positive nucleosomes exhibited relative sensitivity to intercalators as compared to bulk H3-GFP or nucleosomes carrying any of the following marks: H3K27me1, H3K27me2, H3K27me3, H3K9me1, H3K9me2, H3K9me3, H3K36me3, H3K4me0, H3K4me1, H3K4me2, H3K9ac, and H3K14ac. Nickase or DNase I treatment of the nuclei, or bleomycin treatment of live cells, did not affect the stability of nucleosomes carrying H3K4me3 or H3K27ac, while those of the second group were all destabilized upon treatment with intercalators. These observations support the possibility that the promoter proximal marks specifying dynamic nucleosomes are juxtaposed with relaxed DNA sequences due to DNA breaks generated in vivo . In line with this interpretation, endogeneous, 3'OH nicks were mapped within the nucleosome free region of promoters in human mononuclear cells as well as in mES. We also present evidence that the chromatin regions harboring the breaks are topologicaly separated from the domains containing superhelical chromatin. Regarding the mechanism eliciting the breaks, two alternative models were tested experimentally using kncok out cells, inhibitors and degrader systems, based on TOP2 activity or on histone demethylation induced oxydative DNA lesions and their repair. These observations lend support for a model where the role of DNA strand discontinuities in transcriptional regulation and in higher-order chromatin organization are integrated. Grants: OTKA/NKFIH 138524, ERA-Net NEURON 2024-1.2.2-ERA_NET-2024-00009 *PN and LI contributed equally
利益披露 Disclosure
P. Nanasi Jr., None..
L. Imre, None..
I. Szatmari, None..
V. Dombradi, None..
G. Szabo, None.