Identification of a Novel Enhancer/Chromatin Opening Element Associated with High-Level γ-Globin Gene Expression.
Identification of a Novel Enhancer/Chromatin Opening Element Associated with High-Level γ-Globin Gene Expression.
复制标题
DOI：
10.1128/mcb.00197-18
复制
发表时间：
2018-10-01
影响因子：
5.3
通讯作者：
Bungert J
中科院分区：
文献类型：
--
作者：
Shen Y;Bassett MA;Gurumurthy A;Nar R;Knudson IJ;Guy CR;Perez A;Mellen RW;Ikeda M;Hossain MA;Huang S;Igarashi K;Bungert J
11号染色体上5个β-型珠蛋白基因的组织结构反映了红系细胞发育过程中的表达时间，其中胚胎ε-珠蛋白基因位于5‘端，其次是两个胎儿γ-珠蛋白基因，成人β-和δ-珠蛋白基因位于3’端。在这里，我们从功能上鉴定了位于Gγ-珠蛋白基因上游4kb的DNA酶I超敏感位点(HbG-4kb HS)。11号染色体上5个β-型珠蛋白基因的组织结构反映了红系细胞发育过程中的表达时间，其中胚胎ε-珠蛋白基因位于5‘端，其次是两个胎儿γ-珠蛋白基因，成人β-和δ-珠蛋白基因位于3’端。在这里，我们从功能上鉴定了位于Gγ-珠蛋白基因上游4kb的DNA酶I超敏感位点(HbG-4kb HS)。在人类红白血病细胞系K562中，转录因子USF1、USF2、Egr1、MafK和NF-E2占据了这个位置，并且表现出典型的增强子的组蛋白修饰。我们合成了针对HBG-4kb HS的锌指(ZF)DNA结合域(HBG-4kb ZF)。HBG-4kb ZF在体外和细胞内与靶部位相互作用，具有很高的亲和力和特异性。将hBG-4kb ZF直接导入K562和原代人红系细胞，可导致γ-珠蛋白基因表达降低，这与Hs及其下游转录因子和活性组蛋白标记结合减少有关。这些数据表明，HBG-4kb HS对胎儿珠蛋白的产生是重要的，并提示它可能通过定向打开染色质而起作用。
The organization of the five β-type globin genes on chromosome 11 reflects the timing of expression during erythroid cell development, with the embryonic ε-globin gene being located at the 5′ end, followed by the two fetal γ-globin genes, and with the adult β- and δ-globin genes being located at the 3′ end. Here, we functionally characterized a DNase I-hypersensitive site (HS) located 4 kb upstream of the Gγ-globin gene (HBG-4kb HS). The organization of the five β-type globin genes on chromosome 11 reflects the timing of expression during erythroid cell development, with the embryonic ε-globin gene being located at the 5′ end, followed by the two fetal γ-globin genes, and with the adult β- and δ-globin genes being located at the 3′ end. Here, we functionally characterized a DNase I-hypersensitive site (HS) located 4 kb upstream of the Gγ-globin gene (HBG-4kb HS). This site is occupied by transcription factors USF1, USF2, EGR1, MafK, and NF-E2 in the human erythroleukemia cell line K562 and exhibits histone modifications typical for enhancers. We generated a synthetic zinc finger (ZF) DNA-binding domain targeting the HBG-4kb HS (HBG-4kb ZF). The HBG-4kb ZF interacted with the target site in vitro and in the context of cells with a high affinity and specificity. Direct delivery of the HBG-4kb ZF to K562 and primary human erythroid cells caused a reduction in γ-globin gene expression which was associated with decreased binding of transcription factors and active histone marks at and downstream of the HS. The data demonstrate that the HBG-4kb HS is important for fetal globin production and suggest that it may act by opening chromatin in a directional manner.