劣性遺伝性脊髄小脳変性症の分子病態の解明

隐性脊髓小脑变性的分子发病机制的阐明

• 批准号: 20023012
• 负责人: 西澤 正豊
• 金额: $ 3.58万
• 依托单位: Niigata University
• 依托单位国家: 日本
• 项目类别: Grant-in-Aid for Scientific Research on Priority Areas
• 财政年份: 2008
• 资助国家: 日本
• 起止时间: 2008 至 2009
• 项目状态: 已结题
• 来源: https://kaken.nii.ac.jp/en/grant/KAKENHI-PROJECT-20023012/
• 关键词: 脊髄小脳変性症; 劣性遺伝; アプラタキシン欠損症; 1本鎖DNA損傷修復; 分子病態; 運動失調症; 核小体

眼球運動失行と低アルブミン血症を伴う早期発症型脊髄小脳失調症(EAOH/AOA1)の原因遺伝子アプラタキシン(APTX)を同定し,その遺伝子産物APTXについて解析してきた.本研究期間では,疾患関連ミスセンス変異型APTX,本邦で最多であるフレームシフト変異型APTXの病態機序の解明を目的とした.野生型,および疾患関連ミスセンス変異型,疾患非関連ミスセンス変異型APTXの組換えタンパク質を作成し.3'末端にリン酸基またはPG基の付加した20merのオリゴヌクレオチドを基質として用いて,3'phosphatase活性や3'PG removal活性を検討した.APTXと蛍光蛋白との融合蛋白を用い,細胞内局在を共焦点レーザー顕微鏡で観察した.さらに,APTXの蛋白-蛋白結合を明らかとするため,N末V5 tag付き全長型APTXと,N末GFP tag付き全長型・断片型・ミスセンス変異型APTXを用い,共免疫沈降法にて解析した.野生型およびAPTX欠損マウス由来の小脳顆粒細胞を用いてコメットアッセイを行い,DNA切断損傷の蓄積量を検討した.APTXは損傷3'断片に関しては3'phosphatase活性や3'PG removal活性を示し,かっ損傷5'断片に関しては強い切断部5'-末断端のadenosine monophosphate (AMP)残基の除去活性を示した.疾患関連ミスセンス変異型APTXであるP206LおよびV263Gは,ともに3'phosphatase活性,3'PG removal活性,5'-AMP removal活性を示さなかった.一方,疾患非関連APTXミスセンス変異体H260Aは,5'-AMP removal活性は示さないが,3'phosphatase活性や3'PG removal活性を示した.また,APTXは二量体形成能をもち,これがAPTXの核小体のGranular Center (GC)への局在に重要であることを示した.一方,疾患関連ミスセンス変異型APTXはこ量体形成能を失い,GCへの局在能を失った.一方,疾患非関連APTXミスセンス変異体H260Aは核小体の二量体形成能を保持しGCへ局在した.APTXの生理活性として,3',5'損傷断端のremoval活性のうち何れが重要であるかを検討するために,APTX欠損神経細胞にH260A,野生型APTXを導入し,その修復機能を検討した.APTX欠損神経細胞では,ブレオマイシン負荷によりDNA単鎖切断損傷が有意に蓄積するが,その損傷は野生型APTXでもH260A変異型APTXでも同様に修復された.一方,本邦で最も多いフレームシフト変異型689 insTに関しては,ミスセンス依存性mRNA分解機構(NMD)により,mRNAの量が減少しておこっていることを見いだした.

The cause of hypomotor dyskinesia with early symptomatic microsomia (EAOH/AOA1) is the same as that of APTX, which is the same as that of APTX. During the period of this study, most of the patients were diagnosed as APTX, and the most common diseases in our country were the pathogenesis of APTX and the purpose of explaining the pathogenesis. Wild type, disease, disease The interior of the cell is in common focus. APTX protein-protein binding assay, N-terminal V5 tag to full-length APTX, N-terminal GFP tag to full-length fragment APTX, co-immunoprecipitation assay. The cause of the wild-type APTX deficiency is that the small grain cell is used to remove the activity of the adenosine monophosphate (AMP) residue from the cut-off section 5-end adenosine monophosphate (AMP). The APTX fragment 3 'fragment shows the 3'PG removal activity, and the APTX fragment 5' shows the activity of APTX. The 3'phosphatase activity, 3'PG removal activity and 5'-AMP removal activity of the disease were significantly higher than those of the control group (P < 0.05). On the one hand, the disease was not infected by APTX. The activity of 5'-AMP removal and 3'PG removal showed that the disease was H260A and 3'PG removal, respectively. APTX nucleosome Granular Center (GC) nucleosome is important to show that it is important. On the one hand, the disease is characterized by the loss of body formation energy in APTX, and the loss of energy in GC. On the one hand, the formation of two-dimensional nucleosome H260A nucleosomes can maintain the physiological activity of GC, the activity of removal, the activity of removal, the activity of APTX, APTX, APTX and APTX. The wild type APTX, the H260A type APTX, the same type of APTX, the same as the others. On the one hand, the most important information in our country is that the number of insT is 689, which is dependent on the mRNA decomposition mechanism (NMD), and the amount of mRNAs is low.

项目成果

Cardiac sympathetic denervation in Parkinson's disease Iinked to SNCA duplication

帕金森病的心脏交感神经去神经支配与 SNCA 重复有关

• 发表时间: 2008
• 期刊: Acta Neuropathol 116
• 作者: Orimo S;Uchihara T;Nakamura A;Mori F;Ikeuchi T;Onodera O;Nishizawa M;Ishikawa A;Kakita A;Wakabayashi K;Takahashi H
• 通讯作者: Takahashi H

Sporadic amyotrophic lateral sclerosis: two pathological patterns shown by analysis of distribution of TDP-43-immunoreactive neuronal and glial cytoplasmic inclusions

• DOI: 10.1007/s00401-008-0385-z
• 发表时间: 2008-08-01
• 期刊: ACTA NEUROPATHOLOGICA
• 影响因子: 12.7
• 作者: Nishihira, Yasushi;Tan, Chun-Feng;Takahashi, Hitoshi
• 通讯作者: Takahashi, Hitoshi

Development of a high- throughput microarray-based resequencing system for neurological disorders and its application to molecular genetics of amyotropic lateral sclerosis

神经系统疾病高通量微阵列重测序系统的开发及其在肌萎缩侧索硬化症分子遗传学中的应用

• 发表时间: 2008
• 期刊: Arch Neurol 65
• 作者: Takahashi Y;Seki N,Ishiura H;Mitsui J;Matsukawa T;Kishino A;Onodera 0;Aoki M;Shimozawa N;Murayama S;Itoyama Y;Suzuki Y;Sobue G;Nishizawa M;Goto J. Tsuii S
• 通讯作者: Goto J. Tsuii S

Frequency of nocturnal sudden death in patients with multiple system atrophy

• DOI: 10.1007/s00415-008-0941-4
• 发表时间: 2008-10-01
• 期刊: JOURNAL OF NEUROLOGY
• 影响因子: 6
• 作者: Shimohata, Takayoshi;Ozawa, Tetsutaro;Nishizawa, Masatoyo
• 通讯作者: Nishizawa, Masatoyo

Sporadic amyotrophic lateral sclerosis of long duration is associated with relatively mild TDP-43 pathology

• DOI: 10.1007/s00401-008-0443-6
• 发表时间: 2009-01-01
• 期刊: ACTA NEUROPATHOLOGICA
• 影响因子: 12.7
• 作者: Nishihira, Yasushi;Tan, Chun-Feng;Takahashi, Hitoshi
• 通讯作者: Takahashi, Hitoshi