作　　者： (亓合媛）; (李艳明）; (熊倩）; (谢兵兵）; (张昭军）; (方向东）;

机构地区： 中国科学院北京基因组研究所基因组科学与信息重点实验室,北京100101 中国科学院大学生命科学学院,北京100049

出　　处： 《发育医学电子杂志》 2017年第1期1-6,共6页

摘　　要： 目的研究CTCF对红系分化的影响及相应调控机制。方法敲低K562细胞中的CTCF,通过Hemin诱导和荧光实时定量聚合酶链反应(quantitative real-time polymerase chain reaction,qRT-PCR)技术,分析Hemin诱导0到4天的对照和CTCF敲低细胞的珠蛋白基因表达水平,研究CTCF敲低对红系分化的影响。利用基因表达芯片全基因组范围分析CTCF敲低造成的影响。根据公共数据分析推测CTCF调控红系分化的潜在机制,通过染色质免疫共沉淀(chromatin immunoprecipitation,ChIP)验证。结果随着诱导的进行,对照和敲低细胞中的珠蛋白基因HBE和HBG表达均逐渐升高,但CTCF敲低细胞中HBE和HBG基因水平均低于对照细胞,提示CTCF敲低能够抑制K562细胞分化过程中珠蛋白基因的表达。通过检测全基因组范围的基因表达水平,共筛选到1 128个差异表达基因,其中616个基因在CTCF敲低样本中表达上调,512个基因表达下调。利用IPA软件进行功能富集分析,显示主要相关的生理系统发育与功能包括血液系统的发育与功能。公共数据显示存在CTCF-GATA1-ALAS2作用关系,ALAS2基因上存在GATA1结合;对照和CTCF敲低细胞的ChIP-qPCR结果表明,敲低CTCF能够降低GATA1在ALAS2基因区的结合。结论 CTCF可能通过影响GATA1在ALAS2基因区的结合来调控K562细胞红系分化。 Objective To study the function of CTCF during erythroid differentiation and the relevant mechanism. Methods CTCF was knockdown in K562 cells. Heroin inducing was performed for four days. Quantitative real-time polymerase chain reaction （qRT-PCR） were performed to detect expression of globin genes in control and CTCF knockdown K562 cells, and to evaluate the role of CTCF in erythroid differentiation. Global gene expression pattern after CTCF knockdown was analyzed. Both array data and public data were analyzed to speculate potential regulating mechanism. Chromatin immunoprecipitation （CHIP） was performed to further validate our hypothesis. Results Along with Hemin inducing, expressions of globin genes HBE and HBG both increased in control and CTCF knockdown K562 cells, while expressions of globin genes HBE and HBG in CTCF knockdown K562 cells were lower than that in control cells, indicating CTCF knockdown inhibited expressions of globin genes during erythroid differentiation of K562 cells. 1 128 differentially expressed genes （DEGs） were identified through genome wide gene expression array. Among them, 616 genes were up-regulated in CTCF knockdown cells while 512 genes were down-regulated. IPA analyses showed that the main function items of DEGs included development and function of hematological system. Public data suggested the potential correlation of CTCF-GATA1-ALAS2, and GATA1 was found binding at the ALAS2 gene locus. The ChIP-qPCR results indicated that knockdown CTCF in K562 was able to decrease the binding of GATA1 at ALAS2 gene locus. Conclusion CTCF plays a role in K562 erythroid differentiation may through regulating interaction between GATA1 and ALAS2 gene locus.

关 键 词： 红系分化