Alterations in Expression and Chromatin Configuration of the Alpha Hemoglobin-Stabilizing Protein Gene in Erythroid Krüppel-Like Factor-Deficient Mice
Alterations in Expression and Chromatin Configuration of the Alpha Hemoglobin-Stabilizing Protein Gene in Erythroid Krüppel-Like Factor-Deficient Mice
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DOI：
10.1128/mcb.02216-05
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发表时间：
2006-06
影响因子：
5.3
通讯作者：
A. M. Pilon;Douglas G. Nilson;Dewang Zhou;J. Sangerman;T. Townes;D. Bodine;P. Gallagher
中科院分区：
文献类型：
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作者：
A. M. Pilon;Douglas G. Nilson;Dewang Zhou;J. Sangerman;T. Townes;D. Bodine;P. Gallagher
摘要EKLf是一种红系锌指蛋白，它与̈-珠蛋白启动子中的一个CACCC序列相互作用，在β-珠蛋白启动子中建立局部染色质结构，允许β-珠蛋白基因转录。我们试图鉴定其他EKLF靶基因，并在EKLF存在和不存在的情况下确定这些基因的染色质状态。我们通过消减杂交鉴定了α-血红蛋白稳定蛋白(AHSP)，并证明在EKLF缺陷的细胞中AHSP的mRNA降低了95%到99.9%，并且AHSP缺失。与野生型染色质的高乙酰化相比，来自EKLF缺陷细胞的AHSP启动子上的染色质缺乏DNase I超敏感部位，并且在整个基因座上表现出组蛋白低乙酰化。野生型染色质在启动子区域CACCC盒上显示了与EKLF结合的峰，与EKLF共识相差一个核苷酸。在迁移率改变分析中，AHSP启动子CACCC以与β-珠蛋白启动子CACCC位点类似的方式与EKLF结合，表明EKLF共识结合位点有更广泛的识别序列。在缺乏EKLF的K562细胞中，AHSP启动子被EKLF反式激活。这些结果支持EKLf作为AHSP和β-珠蛋白基因的转录因子和染色质调节剂的假说，并表明EKLf可能在其他红系基因中发挥类似的作用。
ABSTRACT Erythroid Krüppel-like factor (EKLF) is an erythroid zinc finger protein identified by its interaction with a CACCC sequence in the β-globin promoter, where it establishes local chromatin structure permitting β-globin gene transcription. We sought to identify other EKLF target genes and determine the chromatin status of these genes in the presence and absence of EKLF. We identified alpha hemoglobin-stabilizing protein (AHSP) by subtractive hybridization and demonstrated a 95 to 99.9% reduction in AHSP mRNA and the absence of AHSP in EKLF-deficient cells. Chromatin at the AHSP promoter from EKLF-deficient cells lacked a DNase I hypersensitive site and exhibited histone hypoacetylation across the locus compared to hyperacetylation of wild-type chromatin. Wild-type chromatin demonstrated a peak of EKLF binding over a promoter region CACCC box that differs from the EKLF consensus by a nucleotide. In mobility shift assays, the AHSP promoter CACCC site bound EKLF in a manner comparable to the β-globin promoter CACCC site, indicating a broader recognition sequence for the EKLF consensus binding site. The AHSP promoter was transactivated by EKLF in K562 cells, which lack EKLF. These results support the hypothesis that EKLF acts as a transcription factor and a chromatin modulator for the AHSP and β-globin genes and indicate that EKLF may play similar roles for other erythroid genes.