EKLF 基因缺陷小鼠中的主要红细胞膜蛋白基因。

Major erythrocyte membrane protein genes in EKLF-deficient mice.

作者信息

Nilson Douglas G, Sabatino Denise E, Bodine David M, Gallagher Patrick G

机构信息

Hematopoiesis Section, Genetics and Molecular Biology Branch, National Human Genome Research Institute, National Institutes of Health, Bethesda, MD, USA.

出版信息

Exp Hematol. 2006 Jun;34(6):705-12. doi: 10.1016/j.exphem.2006.02.018.

DOI:10.1016/j.exphem.2006.02.018

PMID:16728274

Abstract

OBJECTIVES

Mice deficient in the transcription factor erythroid Krüppel-like factor, KLF1 (EKLF) die approximately 14.5 days postcoitum of anemia, attributed to decreased expression of the beta-globin gene. The objectives of this study were to rescue EKLF-deficient embryos with mice expressing gamma-globin from beta-spectrin or ankyrin promoters and to characterize expression of the major erythrocyte membrane genes in EKLF-deficient cells.

METHODS

Transgenic beta-spectrin/gamma-globin or ankyrin/gamma-globin mice were bred onto EKLF-deficient and wild-type backgrounds. Animals were genotyped, gamma-globin mRNA levels measured, and hemoglobin electrophoresis performed. Steady-state mRNA levels and transcriptional rates of the major erythrocyte membrane protein genes were assayed.

RESULTS

beta-spectrin/gamma-globin or ankyrin/gamma-globin mice on EKLF-deficient and wild-type backgrounds had identical levels of gamma-globin mRNA, indicating EKLF-independence of these promoters. gamma-Globin expression improved globin chain imbalance, but hemolysis was not improved and no live-born EKLF-deficient/(A)gamma-globin mice were obtained. Circulating erythroid cells from EKLF-deficient/(A)gamma-globin embryos exhibited hemolysis reminiscent of that seen in patients with severe erythrocyte membrane defects. Levels of beta-spectrin, ankyrin, and band 3 mRNA, but not alpha-spectrin, were decreased in EKLF-deficient fetal liver RNA. In a run-on assay, levels of transcription of the ankyrin and band 3 genes were decreased in EKLF-deficient fetal liver nuclei.

CONCLUSIONS

These results indicate that the EKLF-responsive regions of the ankyrin and beta-spectrin genes are outside their promoters and that EKLF is necessary for full transcriptional activity of the ankyrin and band 3 genes; the results also provide additional evidence that defects in addition to beta-globin deficiency, including an abnormal erythrocyte membrane, contribute to the anemia and embryonic lethality in EKLF-deficient mice.

摘要

目的

缺乏转录因子红系Krüppel样因子KLF1（EKLF）的小鼠在妊娠约14.5天时死于贫血，这归因于β-珠蛋白基因表达降低。本研究的目的是用从β-血影蛋白或锚蛋白启动子表达γ-珠蛋白的小鼠拯救EKLF缺陷胚胎，并表征EKLF缺陷细胞中主要红细胞膜基因的表达。

方法

将转基因β-血影蛋白/γ-珠蛋白或锚蛋白/γ-珠蛋白小鼠与EKLF缺陷和野生型背景的小鼠杂交。对动物进行基因分型，测量γ-珠蛋白mRNA水平，并进行血红蛋白电泳。测定主要红细胞膜蛋白基因的稳态mRNA水平和转录速率。

结果

在EKLF缺陷和野生型背景下的β-血影蛋白/γ-珠蛋白或锚蛋白/γ-珠蛋白小鼠具有相同水平的γ-珠蛋白mRNA，表明这些启动子不依赖于EKLF。γ-珠蛋白表达改善了珠蛋白链失衡，但溶血未得到改善，且未获得活产的EKLF缺陷/（A）γ-珠蛋白小鼠。来自EKLF缺陷/（A）γ-珠蛋白胚胎的循环红细胞表现出溶血，类似于严重红细胞膜缺陷患者所见。在EKLF缺陷的胎肝RNA中，β-血影蛋白、锚蛋白和带3 mRNA的水平降低，但α-血影蛋白的水平未降低。在核转录分析中，EKLF缺陷的胎肝细胞核中锚蛋白和带3基因的转录水平降低。