羟基脲诱导前后HEL细胞内GATA转录因子与人β—类珠蛋白基因调控元件的结合模式分析

ANALYSIS OF THE BINDING OF GATA FACTORS TO THE REGULATORY ELEMENTS OF HUMAN β- GLOBIN GENE IN HYDROXYUREA-INDUCED AND UNINDUCED HEL CELLS

HEL细胞是一株人红白血病细胞株,其中成年型β-珠蛋白基因不能表达。我们以往的试验证明,羟基脲诱导以后,能使HEL细胞内成年型β-珠蛋白基因表达。本文以此为模型,探索了β-珠蛋白基因在HEL细胞内诱导表达的分子机制。结果表明,羟基脲诱导之后,与人β-珠蛋白基因5’远侧端DNaseⅠ超敏感点2核心DNA序列以及近侧端启动子DNA序列相结合的GATA-1蛋白因子量明显增多,而GATA-2蛋白因子量明显减少。结果显示,GATA蛋白家族各成员在HEL细胞分化及珠蛋白基因表达的过程中扮演着不同的角色。推测GATA-1可能有促进β-珠蛋白基因的表达,使HEL细胞趋向终末分化的作用,GATA-2则可能与胚胎型珠蛋白基因表达有关,并有抑制红细胞向终末分化的功能。

HEL cells, a human erythroleukemia cell line, expressing mainly the γ-globin genes, small amount of ε-globin gene, but not β-globin gene. Our previous studies demonstrated that β-globin gene could be expressed in HEL cells induced by hydroxyurea. However, the molecular mechen-ism is still unknown. Here the binding patterns of GATA factors (GATA-1 and GATA-2)to the regulatory elements of human β-globin gene were examined with the nuclear extracts from hydrox-yurea-induced and uninduced HEL cells. Our results showed in EMSA assay that GATA factors could bind to the core sequence of HS2(-10681 to -10971 bp),the 3' flanking sequence of HS2 core( -10323 to-10680 bp)and the promoter ofhuman 2-globin gene( + 20 to-112 bp). However, the binding patterns between hydroxyurea-in-duced and uninduced HEL cells were different. Furthermore, by using Western-blot analysis, our data showed that the amount of GATA-2 was decreased in hydroxyurea-induced HEL cells. In contrast to GATA-2, the amount of GATA-1 was increased in hydroxyurea-induced HEL cells. These results showed that the different members of GATA family might play different roles during the differentiation of erythrocytes. GATA-1 may stimulate the differentiation of HEL cells, while GATA-2 can probably inhibit the differentiation of HEL cells.