Keys Janelle R, Tallack Michael R, Hodge Denise J, Cridland Simon O, David Rakesh, Perkins Andrew C
Institute for Molecular Bioscience, University of Queensland, St Lucia, Qld, Australia.
Br J Haematol. 2007 Jan;136(1):150-7. doi: 10.1111/j.1365-2141.2006.06381.x. Epub 2006 Oct 27.
Alpha haemoglobin stabilising protein (AHSP) binds free alpha-globin chains and plays an important role in the protection of red cells, particularly during beta-thalassaemia. Murine ASHP was discovered as a GATA-1 target gene and human AHSP is directly regulated by GATA-1. More recently, AHSP was rediscovered as a highly erythroid Kruppel-like factor (EKLF) -dependent transcript. We have determined the organisation of the murine AHSP gene and compared it to orthologs. There are two CACC box elements in the proximal promoter. The proximal element is absolutely conserved, but does not bind EKLF as it is not a canonical binding site. In rodents, the distal element contains a 3 bp insertion that disrupts the typical EKLF binding consensus region. Nevertheless, EKLF binds this atypical site by gel mobility shift assay, specifically occupies the AHSP promoter in vivo in a chromatin immunoprecipitation assay, and transactivates AHSP through this CACC site in promoter-reporter assays. Our results suggest EKLF can occupy CACC elements in vivo that are not predictable from the consensus binding site inferred from structural studies. We also propose that absence of AHSP in EKLF-null red cells exacerbates the toxicity of free alpha-globin chains, which exist because of the defect in beta-globin gene activation.
α血红蛋白稳定蛋白（AHSP）可结合游离的α珠蛋白链，在红细胞保护中发挥重要作用，尤其是在β地中海贫血期间。小鼠ASHP被发现是一种GATA-1靶基因，人类AHSP直接受GATA-1调控。最近，AHSP被重新发现是一种高度依赖红系Kruppel样因子（EKLF）的转录本。我们已经确定了小鼠AHSP基因的结构，并将其与直系同源基因进行了比较。近端启动子中有两个CACC框元件。近端元件是绝对保守的，但由于它不是典型的结合位点，所以不与EKLF结合。在啮齿动物中，远端元件包含一个3 bp的插入序列，该序列破坏了典型的EKLF结合共有区域。然而，通过凝胶迁移率变动分析，EKLF可结合这个非典型位点，在染色质免疫沉淀分析中，它在体内特异性占据AHSP启动子，并在启动子报告基因分析中通过这个CACC位点反式激活AHSP。我们的结果表明，EKLF在体内可占据CACC元件，而这些元件无法从结构研究推断出的共有结合位点预测出来。我们还提出，在EKLF缺失的红细胞中，由于β珠蛋白基因激活缺陷而存在的游离α珠蛋白链的毒性会因AHSP的缺失而加剧。
