Gene manipulation of heme synthetic pathway enzymes

• Project/Area Number: 10557015
• Research Category: Grant-in-Aid for Scientific Research (B)
• Allocation Type: Single-year Grants
• Section: 展開研究
• Research Field: Pathological medical chemistry
• Research Institution: Tohoku University
• Principal Investigator: HAYASHI Norio Tohoku University, Graduate School of Medicine, Professor (00004606)
• Co-Investigator(Kenkyū-buntansha): YAMAMOTO Masayuki Tohoku University, Graduate School of Medicine, Professor (50166823) ; TAKETANI Shigeru Kyoto Institute of Technology, Faculty of Fiber Science, Associate Professor (20121949) ; KOBAYASHI Akira Tohoku University, Graduate School of Medicine, Assistant Professor (50292214)
• Project Period (FY): 1998 – 1999
• Project Status: Completed (Fiscal Year 1999)
• Budget Amount: ¥10,700,000 (Direct Cost: ¥10,700,000); Fiscal Year 1999: ¥4,500,000 (Direct Cost: ¥4,500,000); Fiscal Year 1998: ¥6,200,000 (Direct Cost: ¥6,200,000)
• Keywords: Heme / Erythroid cells / Knockout mice / Transgenic mice / 5-アミノレブリン酸合成酵素 / ヘム生合成 / 遺伝子改変マウス / 鉄芽球性貧血 / 鉄代謝 / 環状鉄芽球 / 赤血球分化 / モデル動物

Research Abstract

Heme biosynthesis pathway contains a battery of tissue-specific enzymes, including ALAS-E (Alas2). Our purpose is to clarify the pathogenesis of congenital diseases caused by the defective heme synthesis, such as X-linked sideroblastic anemia (XLSA), through generating an ALAS-E knockout line of mice. We generated ALAS-E knockout mouse line through gene targeting experiments. The ALAS-E-deficient mice were embryonic lethal. The cause of the lethality appeared to be the defective primitive hematopoiesis. Chimeric analysis of ALAS-E-null ES cells revealed the presence of sideroblasts that are similar to those observed in human XLSA patients. It was highly expected that the ring sideroblasts were originated from the ALAS-E-null ES cells. We then generated transgenic (TG) mouse lines expressing ALAS-E mutant molecule isolated from the human samples of XLSA patients under the regulation of GATA1-hematopoietic regulatory domain (G1HRD). When G1HRD-ALAS-Emut TG mice were crossed into ALAS-E-null background, severe phenotypes of ALAS-E-null mice were rescued only partially, suggesting that full-activity of ALAS-E is required to fully rescue the knockout phenotype. The results indicate that the ALAS-E mutant molecule is functionally impaired. These results thus demonstrate that close relationship between heme synthesis and erythropoiesis.

Research Products

• [Journal Article] Molecular cloning and functional characterization of a new CNC family transcription factor Nrf3. (1999)
  • Author(s): Kobayashi, A., Itoh, E., Toki, T., Takahashi, S-i, Igarashi, K., Hayashi, N., Yamamoto, M.
  • Journal Title: J. Biol. Chem. 274 Pages: 6443-6452
  • Description: 「研究成果報告書概要(和文)」より
• [Journal Article] Molecular cloning and functional characterization of a new CNC family transcription factor Nrf3 (1999)
  • Author(s): Kobayashi, A., Itoh, E., Toki, T., Takahashi, S-i, Igarashi, K., Hayashi, N., Yamamoto, M.
  • Journal Title: J.Biol.Chem. 274 Pages: 6443-6452
  • Description: 「研究成果報告書概要(欧文)」より
• [Journal Article] Deficient heme and globin synthesis in ES cells lacking the erythroid-specific δ-aminolevulinate synthase gene. (1998)
  • Author(s): Harigae, H., Suwabe, N., Weinstock, P.H., Nagai, M., Fujita, H., Yamamoto, M., Sassa, S.
  • Journal Title: Blood 91 Pages: 798-805
  • Description: 「研究成果報告書概要(和文)」より
• [Journal Article] Alternative promoters regulate transcription of the mouse GATA-2 gene. (1998)
  • Author(s): Minegishi, N., Ohta, J., Suwabe, N., Nakauchi, H., Ishihara, H., Hayashi, N., Yamamoto, M.
  • Journal Title: J. Biol. Chem. 273 Pages: 3625-3634
  • Description: 「研究成果報告書概要(和文)」より
• [Journal Article] Regulation of NF-E2 activity in erythroleukemia cell differentiation. (1998)
  • Author(s): Nagai, T., Igarashi, K., Akasaka, J., Furuyama, K., Fujita, H., Hayashi, N., Yamamoto, M., Sassa, S.
  • Journal Title: J. Biol. Chem. 273 Pages: 5358-5365
  • Description: 「研究成果報告書概要(和文)」より
• [Journal Article] Differential regulation of mouse coproporphyrinogen oxidase geneexpression in erythroid and non-erythroid cells. (1998)
  • Author(s): Takahashi, S-i., Taketani, S., Akasaka, J-e., Kobayashi, A., Hayashi, N., Yamamoto, M., Nagai, T.
  • Journal Title: Blood 92 Pages: 3436-3444
  • Description: 「研究成果報告書概要(和文)」より
• [Journal Article] Deficient heme and globin synthesis in ES cells lacking the erythroid-specific δ-aminolevulinate synthase gene (1998)
  • Author(s): Harigae, H., Suwabe, N., Weinstock, P.H., Nagai, M., Fujita, H., Yamamoto, M., Sassa, S.
  • Journal Title: Blood 91 Pages: 798-805
  • Description: 「研究成果報告書概要(欧文)」より
• [Journal Article] Alternative promoters regulate transcription of the mouse GATA-2 gene (1998)
  • Author(s): Minegishi, N., Ohta, J., Suwabe, N., Nakauchi, H., Ishihara, H., Hayashi, N., Yamamoto, M.
  • Journal Title: J.Biol.Chem. 273 Pages: 3625-3634
  • Description: 「研究成果報告書概要(欧文)」より
• [Journal Article] Regulation of NF-E2 activity in erythroleukemia cell differentiation (1998)
  • Author(s): Nagai, T., Igarashi, K., Akasaka, J., Furuyama, K., Fujita, H., Hayashi, N., Yamamoto, M., Sassa, S.
  • Journal Title: J.Biol.Chem. 273 Pages: 5358-5365
  • Description: 「研究成果報告書概要(欧文)」より
• [Journal Article] Differential regulation of mouse coproporphyrinogen oxidase gene expression in erythroid and non-erythroid cells (1998)
  • Author(s): Takahashi, S-i., Taketani, S., Akasaka, J-e., Kobayashi, A., Hayashi, N., Yamamoto, M., Nagai, T.
  • Journal Title: Blood 92 Pages: 3436-3444
  • Description: 「研究成果報告書概要(欧文)」より
• [Publications] Minegishi Naoko: "The mouse GATA-2 gene is expressed in the para-aortic splanchnopleura and aortagonad-mesonephros region"Blood. 93・12. 4196-4207 (1999)
• [Publications] Harigae Hideo: "A novel mutation of the erythroid-specific d-aminolaevulinate synthase gene in a patient with X-linked sideroblastic anaemia"Br.J. Haematol. 106・1. 175-177 (1999)
• [Publications] Furuyama Kazumichi: "Absence of C282Y and H63D mutations of the hemochromatosis gene in Japanese patients with sideroblastic anemia"Am.J. Hematol. 61・4. 276 (1999)
• [Publications] Harigae Hideo: "A novel mutation of the erythroid-specific d-aminolevulinate synthase gene in a patient with non-inherited pyridoxine-responsive sideroblastic anemia"Am.J. Hematol. 62・2. 112-114 (1999)
• [Publications] Nakajima Osamu: "Heme deficiency in erythroid lineage causes differentiation arrest and cytoplasmic iron overload"EMBO J.. 18・22. 6282-6289 (1999)
• [Publications] Furuyama,Kazumichi: "ALAS2 gene encodes a pyridoxine-responsive enzyme with low activity." British J.Haematol.103. 839-841 (1998)
• [Publications] Takahashi,Shinichiro: "Differential regulation of coporoporphyrinogen oxidase gene between erythroid and nonerythroid cells." Blood. 92. 3436-3444 (1998)
• [Publications] Kobayashi,Akira: "Molecular cloning and functional characterization of a new CNCfamily transcription factor Nrf3." J.Biol.Chem.274. 6443-6452 (1999)