Molecular Mechanisms for delta-Aminolevulinate Synthase Gene

人们普遍认为，哺乳动物体内产生的血红素中有近75%用于骨髓中的血红蛋白生物合成。在红系分化过程中，骨髓中血红素生物合成的关键酶——红系特异性三角洲氨基乙酰化合成酶受到红系型转录因子如GATA-1和NF-E2的调控。当我们分析x连锁铁母细胞性贫血患者的骨髓ALAS-E活性时，其水平低至正常对照的10%。然而，Western blot分析显示，酶蛋白的显著减少与酶活性缺陷成正比。因此，患者很可能发生了减少酶蛋白的突变。进一步研究表明，患者的ALAS-E基因存在Asp190Val突变，其蛋白在QT-6细胞中表达的特异性活性与正常对照酶一样高，转录-翻译活性相对相似。因此，我们研究了突变对酶的线粒体运输的影响。虽然突变的氨基酸位于大约。在ala - e蛋白加工位点下游100个氨基酸处，突变导致成熟酶的大小比正常蛋白大1.0和1.5 kDa （Furuyama, 1987）。我们还发现NF-E2、珠蛋白和ala - e受Ras-Raf-MAP激酶信号转导的控制。我们的分析明确了NF-E2的激活域和抑制域分别位于p45和p18 （Nagai, 1988）。就我们研究的非特异性δ -氨基乙酰酸合成酶（ALAS-N）而言，该基因通过NF-KB结合序列受到负调控（Fujita, 1998）。

It is generally accepted that almost 75% of heme produced in mammals are supplied for hemoglobin biosynthesis in the bone marrow. During erythroid differentiation the key enzyme for heme biosynthesis in the bone marrow, erythroid specific delta-aminolevulinate synthase, is regulated by erythroid type transcription factors, such as GATA-1 and NF-E2.When we analyzed bone marrow ALAS-E activity from a patient with X-linked sideroblastic anemia, the level was as low as 10% of that in normal control. Western blot analysis, however, demonstrated a remarkable decrease in enzyme protein proportional to the enzyme activity defect. Thus, it is likely the patient has a mutation (s) to reduce the enzyme protein. Further investigations indicated that ALAS-E gene of the patient has Asp190Val mutation, whose protein expressed in QT-6 cells revealed a specific activity as high as the normal control enzyme with a relatively similar transcription-translation activity. We, therefore, examined effects of the mutation for mitchondrial transport of the enzyme. Although the mutated amino acid located approx. 100 amino acids downstream of the processing site of ALAS-E protein, the mutation resulted in 1.0 and 1.5 kDa larger size mature enzyme when compared with the normal protein (Furuyama, 1987).We have also revealed that NF-E2, globin, and ALAS-E are controlled by Ras-Raf-MAP kinase signal transduction. Our analyzes clarified activate domain and suppress domain of NF-E2 were located within p45 and p18, respectively (Nagai, 1988).As far as we examined nonspecific delta-aminolevulinate synthase (ALAS-N), the gene was negatively regulated via NF-KB binding sequence (Fujita, 1998).