Growth Factor Independence 1b (Gfi1b) Is Important for the Maturation of Erythroid Cells and the Regulation of Embryonic Globin Expression

Growth factor independence 1b (GFI1B) is a DNA binding repressor of transcription with vital functions in hematopoiesis. Gfi1b-null embryos die at midgestation very likely due to defects in erythro- and megakaryopoiesis. To analyze the full functionality of Gfi1b, we used conditionally deficient mice that harbor floxed Gfi1b alleles and inducible (Mx-Cre, Cre-ERT) or erythroid specific (EpoR-Cre) Cre expressing transgenes. In contrast to the germline knockout, EpoR-Cre mediated erythroid specific ablation of Gfi1b allows full gestation, but causes perinatal lethality with very few mice surviving to adulthood. Both the embryonic deletion of Gfi1b by EpoR-Cre and the deletion in adult mice by Mx-Cre or Cre-ERT leads to reduced numbers of erythroid precursors, perturbed and delayed erythroid maturation, anemia and extramedullary erythropoiesis. Global expression analyses showed that the Hba-x, Hbb-bh1 and Hbb-y embryonic globin genes were upregulated in Gfi1b deficient TER119+ fetal liver cells over the gestation period from day 12.5–17.5 p.c. and an increased level of Hbb-bh1 and Hbb-y embryonic globin gene expression was even maintained in adult Gfi1b deficient mice. While the expression of Bcl11a, a regulator of embryonic globin expression was not affected by Gfi1b deficiency, the expression of Gata1 was reduced and the expression of Sox6, also involved in globin switch, was almost entirely lost when Gfi1b was absent. These findings establish Gfi1b as a regulator of embryonic globin expression and embryonic and adult erythroid maturation.生长因子独立1b（Growth factor independence 1b, GFI1B）是一类具备DNA结合能力的转录阻遏蛋白，在造血调控过程中发挥至关重要的作用。Gfi1b生殖系敲除胚胎会于妊娠中期死亡，其诱因极可能为红细胞生成与巨核细胞生成缺陷。为全面解析Gfi1b的完整生物学功能，本研究构建了携带floxed Gfi1b等位基因的条件性敲除小鼠，并搭配三类Cre重组酶表达转基因系统：诱导型（Mx-Cre、Cre-ERT）与红细胞特异性（EpoR-Cre）。 与生殖系敲除模型不同，经EpoR-Cre介导的Gfi1b红细胞特异性敲除可使胚胎完成完整妊娠，但会引发围产期致死，仅极少数小鼠可存活至成年阶段。无论是通过EpoR-Cre在胚胎期敲除Gfi1b，还是借助Mx-Cre或Cre-ERT在成年小鼠体内敲除Gfi1b，均会导致红细胞前体细胞数量减少、红细胞成熟过程紊乱且延迟，进而引发贫血与髓外造血。 全转录组表达分析结果显示，在妊娠第12.5~17.5天（p.c.）的TER119阳性胎肝细胞中，Hba-x、Hbb-bh1及Hbb-y这三种胚胎珠蛋白基因的表达均显著上调；且在成年Gfi1b缺陷小鼠体内，Hbb-bh1与Hbb-y胚胎珠蛋白基因的高表达水平仍可维持。 尽管胚胎珠蛋白表达调控因子Bcl11a的表达未受Gfi1b缺陷的影响，但Gata1的表达水平出现下调，而同样参与珠蛋白基因转换调控的Sox6的表达则在Gfi1b缺失时几乎完全丧失。本研究结果证实，Gfi1b是调控胚胎珠蛋白表达以及胚胎与成年期红细胞成熟的关键调控因子。