カテコ-ルアミン関連酵素DNAプロ-ブによる遺伝性精神神経疾患の遺伝子解析

1990 年度 実績報告書

• 研究課題/領域番号: 01440047
• 研究機関: 名古屋大学
• 研究代表者: 永津 俊治 名古屋大学, 医学部, 教授 (40064802)
• 研究分担者: 萩原 昌子 名古屋大学, 医学部, 教務職員 ; 一瀬 宏 名古屋大学, 医学部, 助手 (90192492) ; 金田 典雄 名古屋大学, 医学部, 講師 (00144139) ; 直井 信 名古屋工業大学, 教授 (50022786)
• キーワード: ヒト・チロシン水酸化酵素 / ヒト・ド-パミンβー水酸化酵素 / ヒト・フェニルエタノ-ルアミンNーメチル基転移酵素 / ヒト・ド-パ脱炭酸酵素 / cDNA / DNAプロ-ブ / カテコ-ルアミン生合成酵素 / トランスジェニックマウス

研究概要

カテコ-ルアミン系神経伝達物質(ド-パミン、ノルアドレナリン、アドレナリン)のヒトの生合成酵素のDNAプロ-ブを用いて、遺伝性精神・神経疾患の発症の分子機構の解明を目指している。これまでに、クロ-ニングしたヒト・チロシン水酸化酵素(hTH)、ヒト・ド-パミンβー水酸化酵素(h DBH)、ヒト・ド-パ脱炭酸酵素(hDDC)、ヒト・フェニルエタノ-ルアミンNーメチル基転移酵素(h PNMT)、のcDNAプロ-ブを用いて、遺伝子異常の解明とモデル動物及び分子診断法の確立を試みる。(1)hTH遺伝子をC57BL/6マウス受精卵に注入したトランスジェニックス(tg)マウスを作製した。導入遺伝子mRNAは副腎と脳に特異的に発現していた。導入THのmRNA発現量は副腎では対照の5倍、脳では50倍であった。in situハイブリダイゼ-ションによる解析では導入遺伝子は中脳黒質と腹側被蓋野に強く発現していた。脳の各部位でのTHタンパク量および酵素活性は対照の3〜5倍の増加で、免疫組織化学法の結果も同じ傾向を示した。またカテコ-ルアミン量は対照に比べ有為な差はなかった。このtgマウスではhTHの遺伝子発現及びカテコ-ルアミン量について未知の調節機構が働いていることが示唆された。(2)2種類のhDBHcDNA(DBHー1、DBHー3)は、6塩基異なり304番目のアミノ酸がDBHー1のではアラニン、DBHー3ではセリンとなり、サル腎臓COS細胞にトランスフェクトして、活性のあるhDBHタンパク質を得ることに成功した。(3)hDDCcDNAをCOS細胞にトランスフェクションした。発現された酵素の分子量は50kDaであった。この酵素はLーDOPA、Lー5HTPおよびLーthreoーDOPSを脱炭酸した。LーDOPAに対する至適pHは6.5であったがLー5HTPに対してはより広範囲なpH領域で活性を示した。LーDOPA、Lー5HTPのいずれを基質とした場合もPLPの添加により活性は大きく上昇した。

研究成果

• [文献書誌] Nagatsu,T.: "Changes of tyrosine hydroxylase in the parkinsonian brain and in the brain of MPTPーtreated mice as revealed by homospecific activity." Neurochem.Res.15. 425-429 (1990)
• [文献書誌] Nagatsu,T.: "The functioning of tyrosine hydroxylase in parkinson's disease." Clin.Neuropharmacol.13. 45-46 (1990)
• [文献書誌] Niwa,T.: "Endogeneous synthesis of Nーmethylー1,2,3,4ーtetrahydroisoーquinoline,a precursor of Nーmethylisoquinolinium ion,in the brains of primates with parkinsonism after systemic administration of 1,2,3,4ーtetrahydroisoquinoline." J.Chromatogr.533. 145-151 (1990)
• [文献書誌] Kojima,T.: "3ーaminoー1ーmethylー5Hーpyrido [4,3ーb] indole (TrpーPー2) and other heterocyclic amines as inhibitors of mitochondrial monoamine oxidases separated from human brain synaptosomes." Neurochem.Int.16. 51-57 (1990)
• [文献書誌] Hirata,Y.: "Studies on the uptake of Nーmethylisoquinolinium ion into rat striatal slices using highーperformance liquid chromatography with fluorimetric detection." J.Chromatogr.503. 189-195 (1990)
• [文献書誌] Naoi,M.: "Inhibition of tryptophan hydroxylase by foodーderived carcinogenic heterocyclic amines,3ーaminoー1ーmethylー5Hーpyrido [4,3,6] indole and 3ーaminoー1,4ーdimethylー5Hーpyrido [4,3,6] indole." Biochem.Pharmacol.41. 199-203 (1991)
• [文献書誌] Uchida,K.: "Tetrahydrobiopterinーdependent production of LーDOPA in NRK fibroblasts transfected with tyrosine hydroxylase cDNA:future use for intracerebral grafting." Neurosci.Lett.109. 282-286 (1990)
• [文献書誌] Alexander,N.: "Rightーleft asymmetry of tyrosine hydroxylase in rat median eminence:influence of arterial baroreflex nerves." Brain Res.523. 195-198 (1990)
• [文献書誌] Hashimoto,R.: "The plasma tetrahydrobiopterin levels in patients with affective disorders." Biolog.Psychiat.28. 526-528 (1990)
• [文献書誌] Ichinose,H.: "Isolation and characterization of a cDNA clone encoding human aromatic Lーamino acid decarboxylase." Biochem.Biophys.Res.Commun.164. 1024-1030 (1989)
• [文献書誌] Sumi,C.: "Characterization of recombinant human aromatic Lーamino acid decarboxylase expressed in COS cells." J.Neurochem.55. 1075-1082 (1990)
• [文献書誌] Ichikawa,S.: "Multiple mRNAs of monkey tyrosine hydroxylase." Biochem.Biophys.Res.Commun.173. 1331-1336 (1990)
• [文献書誌] Hagihara,M.: "Dietary calcium deprivation increased the levels of plasma catecholamines and catecholamineーsynthesizing enzymes of adrenal glands in rats." Biochem.Pharmacol.39. 1229-1231 (1990)
• [文献書誌] Hagihara,M.: "Concentration of neopterin and biopterin in serum from patients with rheumatoid arthritis or systemic lupus erythematous and in synovial fluid from patients with rheumatoid or osteoarthritis." Clin.Chem.36. 705 (1990)
• [文献書誌] Ishii,A.: "Effect of (6R)ーand (6S)ーtetrahydrobiopterin on Lー3,4ーDihydroxyphenylalanine (DOPA) formation in NRK fibroblasts transfected with human tyrosine hydroxylase type 2 cDNA." Neurochem.Int.17. 625-632 (1990)
• [文献書誌] Nagatsu,I.: "Antibodies raised against different oligonucleotid segments of human dopamineーβーhydroxylase." Neurosci.Lett.120. 141-145 (1990)
• [文献書誌] Kaneda,N.: "Tissueーspecific and highーlevel expression of human tyrosine hydroxylase gene in transgenic mice." Neuron. 6. 1-12 (1991)
• [文献書誌] Ishii,A.: "Expression of two forms of human dopamineーβーhydroxylase in COS cells." Neurosci.Lett.(1991)
• [文献書誌] Uchida,K.: "Tetrahydrobiopterinーdependent production of LーDOPA in NRK fibroblasts transfected with tyrosine hydroxylase cDNA:future use for intracerebral grafting." Neurosci.Lett.109. 282-286 (1990)
• [文献書誌] Alexander,N.: "Rightーleft asymmetry of tyrosine hydroxylase in rat median eminence:influence of arterial baroreflex nerves." Brain Res.523. 195-198 (1990)
• [文献書誌] Hashimoto,R.: "The plasma tetrahydrobiopterin levels in patients with affective disorders." Biolog.Psychiat.28. 526-528 (1990)
• [文献書誌] Ichinose,H.: "Isolation and characterization of a cDNA clone encoding human aromatic Lーamino acid decarboxylase." Biochem.Biophys.Res.Commun.164. 1024-1030 (1989)
• [文献書誌] Sumi,C.: "Characterization of recombinant human aromatic Lーamino acid decarboxylase expressed in COS cells." J.Neurochem.55. 1075-1082 (1990)
• [文献書誌] Ichikawa,S.: "Multiple mRNAs of monkey tyrosine hydroxylase." Biochem.Biophys.Res.Commun.173. 1331-1336 (1990)
• [文献書誌] Nagatsu,T.: "Changes of tyrosine hydroxylase in the parkinsonian brain and in the brain of MPTPーtreated mice as revealed by homospecific activity." Neurochem.Res.15. 425-429 (1990)
• [文献書誌] Nagatsu,T.: "The functioning of tyrosine hydroxylase in parkinson's disease." Clin.Neuropharmacol.13. 45-46 (1990)
• [文献書誌] Niwa,T.: "Endogeneous synthesis of Nーmethylー1,2,3,4ーtetrahydroisoーquinoline,a precursor of Nーmethylisoquinolinium ion,in the brains of primates with parkinsonism after systemic administration of 1,2,3,4ーtetrahydroisoquinoline." J.Chromatogr.533. 145-151 (1990)
• [文献書誌] Kojima,T.: "3ーaminoー1ーmethylー5Hーpyrido [4,3ーb] indole (TrpーPー2) and other heterocyclic amines as inhibitors of mitochondrial monoamine oxidases separated from human brain synaptosomes." Neurochem.Int.16. 51-57 (1990)
• [文献書誌] Hirata,Y.: "Studies on the uptake of Nーmethylisoquinolinium ion into rat striatal slices using highーperformance liquid chromatography with fluorimetric detection." J.Chromatogr.503. 189-195 (1990)
• [文献書誌] Naoi,M.: "Inhibition of tryptophan hydroxylase by foodーderived carcinogenic heterocyclic amines,3ーaminoー1ーmethylー5Hーpyrido [4,3,6] indole and 3ーaminoー1,4ーdimethylー5Hーpyrido [4,3,6] indole." Biochem.Pharmacol.41. 199-203 (1991)
• [文献書誌] Hagihara,M.: "Dietary calcium deprivation increased the levels of plasma catecholamines and catecholamineーsynthesizing enzymes of adrenal glands in rats." Biochem.Pharmacol.39. 1229-1231 (1990)
• [文献書誌] Hagihara,M.: "Concentration of neopterin and biopterin in serum from patients with rheumatoid arthritis or systemic lupus erythematous and in synovial fluid from patients with rheumatoid or osteoarthritis." Clin.Chem.36. 705 (1990)
• [文献書誌] Ishii,A.: "Effect of (6R)ーand (6S)ーtetrahydrobiopterin on Lー3,4ーDihydroxyphenylalanine (DOPA) formation in NRK fibroblasts transfected with human tyrosine hydroxylase type 2 cDNA." Neurochem.Int.17. 625-632 (1990)
• [文献書誌] Nagatsu,I.: "Antibodies raised against different oligonucleotid segments of human dopamineーβーhydroxylase." Neurosci.Lett.120. 141-145 (1990)
• [文献書誌] Kaneda,N.: "Tissueーspecific and highーlevel expression of human tyrosine hydroxylase gene in transgenic mice." Neuron. 6. 1-12 (1991)
• [文献書誌] Ishii,A.: "Expression of two forms of human dopamineーβーhydroxylase in COS cells." Neurosci.Lett.(1991)
• [文献書誌] Kiuchi,Y.(Ollat,H.,Parvez,S.and Parvez,H.(Eds.)): "Role of CCK peptides in behavior and neurochemical regulation:Possible interrelation with alcoholism.In:Progress in Alcohol Research" Alcohol & Behavior,VSP Press,The Netherlands, 185-228 (1990)
• [文献書誌] Nagatsu,T.(Tong,H.Joh (Ed.)): "Multiple mRNA of human tyrosine hydroxylase.In:Catecholamine Genes." Neurology and Neurobiology,New York, 9-22 (1990)
• [文献書誌] Sugimoto,T.(H.Ch.Curtius,S.Ghisla and N.Blau (Eds.)): "A new immunoassay for neopterin and biopterin.In:Chemistry and Biology of Pferidines Pteridines and Folic Acid Derivatives." Walter de Gruyter,Berlin,New York, 155-158 (1990)
• [文献書誌] Nagatsu,T.(K.Maurer,P.Riederer and H.Beckman (Eds.)): "In:Alzheimer's Disease:Epidemiology,Neuropathology,Neurochemistry,clinics." Springh Verlag,Wien,New York, 253-260 (1990)
• [文献書誌] Nagatsu,T.(M.B.Streifler,A.D.Korczyn,E.Melamed and M.B.H.Youdim (Eds.)): "Changes of tyrosine hydroxylase in Parkinsonian Brains and in the Brains of MPTPーtreated mice.In:Advances in Neurology:Parkinson's Disease:Anatomy,Pathology,and Therapy" Raven Press,New York, 207-214 (1990)
• [文献書誌] Nagatsu,T.(Nagatsu,T.and Hayaishi,O.(Eds.)): "Normal brain aging versus pathological brain agingーSimilarity and dissimilarity between MPTP parkinsonism and Parkinson's disease in relation to brain aging.In:Aging of the brain:cellular and molecular aspects of brain aging and Alzheimer's disease." Japan Scientific Societies Press,Tokyo and Karger,Basel, 119-127 (1990)
• [文献書誌] Sugimoto,T.(H.Ch.Curtius,S.Ghisla and N.Blau (Eds.)): "A new immunoassay for neopterin and biopterin.In:Chemistry and Biology of Pferidines Pteridines and Folic Acid Derivatives." Walter de Gruyter,Berlin,New York, 155-158 (1990)
• [文献書誌] Nagatsu,T.(K.Maurer,P.Riederer and H.Beckman (Eds.)): "In:Alzheimer's Disease:Epidemiology,Neuropathology,Neurochemistry,clinics." Springh Verlag,Wien,New York, 253-260 (1990)
• [文献書誌] Kiuchi,Y.(Ollat,H.,Parvez,S.and Parvez,H.(Eds.)): "Role of CCK peptides in behavior and neurochemical regulation:Possible interrelation with alcoholism.In:Progress in Alcohol Research" Alcohol & Behavior,VSP Press,The Netherlands, 185-228 (1990)
• [文献書誌] Nagatsu,T.(Tong,H.Joh (Ed.)): "Multiple mRNA of human tyrosine hydroxylase.In:Catecholamine Genes." Neurology and Neurobiology,New York, 9-22 (1990)
• [文献書誌] Nagatsu,T.(M.B.Streifler,A.D.Korczyn,E.Melamed and M.B.H.Youdim (Eds.)): "Changes of tyrosine hydroxylase in Parkinsonian Brains and in the Brains of MPTPーtreated mice.In:Advances in Neurology:Parkinson's Disease:Anatomy,Pathology,and Therapy" Raven Press,New York, 207-214 (1990)
• [文献書誌] Nagatsu,T.(Nagatsu,T.and Hayaishi,O.(Eds.)): "Normal brain aging versus pathological brain agingーSimilarity and dissimilarity between MPTP parkinsonism and Parkinson's disease in relation to brain aging.In:Aging of the brain:cellular and molecular aspects of brain aging and Alzheimer's disease." Japan Scientific Societies Press,Tokyo and Karger,Basel, 119-127 (1990)