Ａｌｕ配列と遺伝性疾患の病態に関する研究

• Project Area: Functions of non-coding DNA region for genome integrity
• Project/Area Number: 24114507
• Research Category: Grant-in-Aid for Scientific Research on Innovative Areas (Research in a proposed research area)
• Allocation Type: Single-year Grants
• Review Section: Biological Sciences
• Research Institution: Gifu University
• Principal Investigator: 深尾 敏幸 岐阜大学, 医学(系)研究科(研究院), 教授 (70260578)
• Project Period (FY): 2012-04-01 – 2014-03-31
• Project Status: Completed (Fiscal Year 2013)
• Budget Amount: ¥11,440,000 (Direct Cost: ¥8,800,000、Indirect Cost: ¥2,640,000); Fiscal Year 2013: ¥5,850,000 (Direct Cost: ¥4,500,000、Indirect Cost: ¥1,350,000); Fiscal Year 2012: ¥5,590,000 (Direct Cost: ¥4,300,000、Indirect Cost: ¥1,290,000)
• Keywords: Alu配列 / 遺伝子内欠失 / 遺伝子内重複 / 相同組換え / スプライシング / エクソンスキップ / 遺伝病

Research Abstract

Alu配列はヒトゲノム中の100万コピー以上もあるとされ、全ゲノムの約１０％を占める。Alu 配列は霊長類の進化とともに出現した。Alu配列はこれまで意味のない配列と考えられてきたが、最近その機能についての知見が少しずつ報告されてきている。申請者はこれまで小児科の先天代謝異常症および臨床遺伝を専門として、ケトン体代謝異常症の分子病態の解析を行ってきた。その中で、遺伝子内における複数のエクソンの欠失、挿入などの遺伝子再構成を伴う症例を経験し、それがイントロン内のAlu配列間での相同組換えによることを、組み換え領域の配列を解析する事で報告してきた．
Alu配列間での組換えによるDNA断片の重複、欠失誘導の実際例の解析では、遺伝性疾患であるT2欠損症、SCOT欠損症、HMG-CoAリアーゼ欠損症、CPT2欠損症、CACT欠損症について独自にMLPA法を確立して遺伝子内欠失、重複を検出できるシステムを確立しており、それを患者解析に適応した．その結果T2欠損症におけるMLPA法の確立と欠失例の同定についてはMol Genet Metab 2013に報告した．またHMG-CoAリアーゼ欠損症についても欠失例を同定している．またAlu配列のスプライシングに与える影響の解析では、T2遺伝子のイントロン１０内に挿入したAlu配列のスプライシングへの影響の検討では、Alu配列の逆方向への挿入が、エクソン１０をスキップする方向に働くことを見いだした．これはイントロン１０の3’ スプライス部位の認識が弱いことに関連して生じていることが判った．

Current Status of Research Progress

Reason

25年度が最終年度であるため、記入しない。

Strategy for Future Research Activity

25年度が最終年度であるため、記入しない。

Research Products

• [Journal Article] Development of MLPA for Human ACAT1 Gene and Identification of a Heterozygous Alu-mediated Deletion of Exons 2 and 3 in a Patient with Mitochondrial Acetoacetyl-CoA Thiolase (T2) Deficiency. (2013)
  • Author(s): Fukao T, Aoyama Y, Murase K, Hori T, Wierenga R, Boneh A, Kondo N
  • Journal Title: Mol Genet Metab Volume: 110 Issue: 1-2 Pages: 184-187
  • DOI: 10.1016/j.ymgme.2013.07.004
  • Peer Reviewed
• [Journal Article] Molecular basis of two exon skipping (exons 12 and 13) by c.1248+5g>a in OXCT1 gene. Study on intermediates of OXCT1 transcripts in fibroblasts. (2013)
  • Author(s): Hori T, Fukao T, Murase K, Sakaguchi N, Harding CO, Kondo N.
  • Journal Title: Hum Mutat Volume: 34 Issue: 3 Pages: 473-480
  • DOI: 10.1002/humu.22258
  • Peer Reviewed
• [Journal Article] Intracellular in vitro probe acylcarnitine assay for identifying deficiencies ofcarnitine transporter and carnitine palmitoyltransferase-1 (2013)
  • Author(s): Purevsuren J
  • Journal Title: Anal Bioanal Chem Volume: 405 Issue: 4 Pages: 1345-1351
  • DOI: 10.1007/s00216-012-6532-3
  • Peer Reviewed
• [Journal Article] Clinical and molecular aspects of Japanese children with medium chain acyl-CoA dehydrogenase deficiency (2012)
  • Author(s): Purevsuren J, Fukao T (8/9), Yamaguchi S (9/9), et al
  • Journal Title: Mol Genet Metab Volume: 107 Issue: 1-2 Pages: 237-240
  • DOI: 10.1016/j.ymgme.2012.06.010
  • Peer Reviewed
• [Journal Article] Bezafibrate can be a new treatment option for mitochondrial fatty acid oxidation disorders : Evaluation by in vitro probe acylcarnitine assay (2012)
  • Author(s): Yamaguchi S (1/12), Fukao T (11/12), et al
  • Journal Title: Mol Genet Metab Volume: 107 Issue: 1-2 Pages: 87-91
  • DOI: 10.1016/j.ymgme.2012.07.004
  • Peer Reviewed
• [Journal Article] Alu配列の関連した遺伝子異常が遺伝性疾患を引き起こす (2012)
  • Author(s): 深尾敏幸
  • Journal Title: 実験医学 Volume: 14 Pages: 2240-2246
• [Journal Article] Three Japanese patients with beta-ketothiolase deficiency whoshare a mutation, c.431A>C(H144P) in ACAT1 : subtle abnormality in urinary organic acid analysis and blood acylvcarnitine analysis using tandem mass spectrometry (2011)
  • Author(s): Fukao T
  • Journal Title: JIMD Reports Volume: 3 Pages: 108-115
  • DOI: 10.1007/8904_2011_72
  • ISBN: 9783642249358, 9783642249365
  • Peer Reviewed
• [Presentation] Development of MLPA and identification of a heterozygous alu-associated deletion including exons 2-4 in a patient with HMGCL deficiency. (2013)
  • Author(s): Aoyama Y, Ishige M, Tanaka T, Fukao T
  • Organizer: The 3rd Asian Congress for Inherited Metabolic Diseases / The 55th Annual meeting of the Japanese Society for Inherited Metabolic Diseases.
  • Place of Presentation: 幕張
• [Presentation] Ketone body metabolism and its defects. Update. (2013)
  • Author(s): Fukao T
  • Organizer: 12th International Congress of Inborn Errors of Metabolism,
  • Place of Presentation: バルセロナ，（スペイン)
  • Invited
• [Presentation] Inborn errors of ketogenesis and ketone body utilization. (2013)
  • Author(s): Fukao T
  • Organizer: The 3rd Asian Congress for Inherited Metabolic Diseases / The 55th Annual meeting of the Japanese Society for Inherited Metabolic Diseases
  • Place of Presentation: 幕張
  • Invited
• [Presentation] ケトン体代謝異常症の病態解明に関する研究．日本先天代謝異常学会学会賞受賞講演 (2012)
  • Author(s): 深尾敏幸
  • Organizer: 第５４回日本先天代謝異常学会総会
  • Place of Presentation: 十六プラザ、岐阜
  • Year and Date: 2012-11-15
  • Invited
• [Presentation] An Exonic splicing enhancer mutation indetified in German beta-ketothiolase deficient patients. (2012)
  • Author(s): Fukao T, Sass JO, Konstantopoulou V, Marquardt T, Frauendienst-Egger G, Kondo N.
  • Organizer: Annual symposium of Society for the Study of Inborn Errors of Metabolism.
  • Place of Presentation: Birmingham, UK
  • Year and Date: 2012-09-04
• [Presentation] Molecular basis of two exon skipping (exons 12 and 13) by c. 1248+5GA in fibroblasts from a SCOT deficient patient. Study of splicing order in SCOT transcripts in fibroblasts from controls and the patient. (2012)
  • Author(s): Hori T, Fukao T, Harding CO, Kondo N.
  • Organizer: Annual symposium of Society for the Study of Inborn Errors of Metabolism.
  • Place of Presentation: Birmingham, UK
  • Year and Date: 2012-09-04