Structure and function of the regulatory region of the gene encoding non-specific form delta-aminolevulinate synthase

编码非特异性形式δ-氨基乙酰丙酸合酶的基因调控区的结构和功能

• 批准号: 04454165
• 负责人: HAYASHI Norio
• 金额: $ 3.71万
• 依托单位: Tohoku University
• 依托单位国家: 日本
• 项目类别: Grant-in-Aid for General Scientific Research (B)
• 财政年份: 1992
• 资助国家: 日本
• 起止时间: 1992 至 1993
• 项目状态: 已结题
• 来源: https://kaken.nii.ac.jp/en/grant/KAKENHI-PROJECT-04454165/
• 关键词: delta-aminolevulinate synthase; heme biosynthesis; gene; rat; regulatory region; δーアミノレブリン酸合成酵素; フィードバック調節

Heme is essential for all eukaryotic cells because of its role as a prosthetic group for a number of proteins, and its intracellular concentration is highly regulated. The first and the rate-limiting step of heme biosynthesis in animals is catalyzed by delta-aminolevulinate(ALA) synthase. There are two tissue-specific isozymes of ALA synthase : the erythroid- specific ALA synthase (ALAS-E) and the non-specific ALA synthase (ALAS-N). To understand the molecular mechanisms contorolling the heme biosynthetic pathway, it is a prerequisite to analyze the structure and function of the ALA synthase genes.We attempted isolation and characterization of the gene encoding rat ALAS-N.The ALAS-N gene was found to span more than 14kb in the rat genome, encompassing 11 exons, Analysis of the promoter region of the gene revealed several potential cos-acting regulatory elements. Comparison of the organization of rat ALAS-N gene with that of the ALAS-E gene in mouse suggests that the ancestral gene for ALA synthase in animals was probably composed of 11 exons, and both ALAS-N and ALAS-E genes were derived from this ancestral gene.The proximal regulatory region of the human ALAS-E gene was also analyzed. The results indicated that binding sites for erythroid transcription factors are clustered within 120 bp upstream form the transcription initiation site.RNA blot hybridization analysis was used to examine the developmental stage-specific transcription of ALAS-N and ALAS-E genes in fetal, newborn and adult rat liver. The results demonstrated that, while ALAS-E is the key enzyme which supplies large quantities of heme for hemoglobin synthesis, ALAS-N functions to supply heme to the P-450 system in the liver and also functions as a housekeeper gene to supply heme for respiratory cytochromes and other hemoproteins in various tissues.

血红素对所有真核细胞都是必需的，因为它作为许多蛋白质的辅基，并且其细胞内浓度受到高度调节。δ-氨基乙酰丙酸（ALA）合成酶是动物血红素生物合成的第一步和限速步骤。ALA合酶有两种组织特异性同工酶：红系特异性ALA合酶（ALAS-E）和非特异性ALA合酶（ALAS-N）。为了了解血红素生物合成途径的分子调控机制，首先要分析ALA合成酶基因的结构和功能，我们尝试分离和鉴定大鼠ALAS-N基因，发现ALAS-N基因在大鼠基因组中的长度超过14 kb，包含11个外显子，对该基因启动子区的分析揭示了几个潜在的相互作用的调控元件。比较大鼠ALAS-N基因和小鼠ALAS-E基因的结构，提示动物ALA合成酶的祖先基因可能由11个外显子组成，ALAS-N和ALAS-E基因均来源于该祖先基因，并对人ALAS-E基因的近端调控区进行了分析。结果表明，ALAS-N和ALAS-E基因在转录起始位点上游120 bp处有一簇转录因子的结合位点，利用RNA印迹杂交技术检测了ALAS-N和ALAS-E基因在胚胎、新生和成年大鼠肝脏中的发育阶段特异性转录。结果表明，ALAS-E是为血红蛋白合成提供大量血红素的关键酶，ALAS-N的功能是为肝脏中的P-450系统提供血红素，并且还作为管家基因为各种组织中的呼吸细胞色素和其他血红素蛋白提供血红素。

项目成果

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Munakata,H.et al.: "Purification and structure of rat erythroid-specific δ-aminolevulinate synthase" J.Biochem.114. 103-111 (1993)

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Munakata, H.et al.: "Purification and structure of rat erythroid-specific delta-aminolevulinate synthase" J.Biochem.114. 103-111 (1993)

Munakata, H.et al.：“大鼠红系特异性 δ-氨基乙酰丙酸合酶的纯化和结构”J.Biochem.114。