Molecular biology for Alzheimer's disease

阿尔茨海默病的分子生物学

• 批准号: 03557037
• 负责人: YUASA Tatsuhiko
• 金额: $ 11.01万
• 依托单位: Tokyo Medical and Dental University
• 依托单位国家: 日本
• 项目类别: Grant-in-Aid for Developmental Scientific Research (B)
• 财政年份: 1991
• 资助国家: 日本
• 起止时间: 1991 至 1993
• 项目状态: 已结题
• 来源: https://kaken.nii.ac.jp/grant/KAKENHI-PROJECT-03557037/
• 关键词: Alzheimer's diseaase; Growth inhibitory factor (GIF); Metallothionein; Gene targeting; Metal responsive-element; cDNA; 遺伝学; cDNAクロ-ニング; ゲノム; 神経成長抑制因子; 神経成長因子; gene targetting

Growth inhibitory factor (GIF) is down-regulated in Alzheimer's disease (AD) brains. We have cloned a full-length cDNA for human GIF gene and found a striking homology to metallothioneins (MTs). In contrast to MT gene, the GIF gene expression is found exclusively in the nervous system. Northern blot analysis revealed that the GIF mRNA expression is drastically decreased in AD brains. To analyze the mechanism of this down-regulation, we isolated the human and mouse GIF genes. These genes consist of three exons, are approximately 1 kb long. A comparison of the human and mouse GIF genes showed several conserved sequences, including the putative AP-2, SP-1, TATA-binding protein and metal-responsive elemnts (MRE). A sequence similar to human gfa common sequence (hgcs), recently identified as the sequence for an astrocyte-specific transcriptional factor, is present in the promoter of these GIF genes.Because GIFs are considered new members of the metallothionein family which are associated with resistance to the toxic effect of metals, we examined the effects of modulation of GIF transcriptional levels using heavy metals and cytokines in primary culture of mouse astrocyte cells. Stimulation by Cu^<2+> and Zn^<2+> induced GIF, whereas stimulation by IL-1 and EGF inhibited the transcription of GIF.These results indicate that several growth factors or cytokines regulate GIF synthesis and play a important part in regulation of the direction and extent of nerve growth. To disrupt GIF sesquences in mouse embryonic stem cells, we made a replacement vector containing a 6-kb genomic fragment that included the entire the GIF gene. The part of first exon was deleted and a neo cassette was inserted in the deleted position. Now we are trying to produce the GIF deficient mouse to elucidate the function of the GIF gene in vivo.

生长抑制因子 (GIF) 在阿尔茨海默病 (AD) 大脑中下调。我们克隆了人类 GIF 基因的全长 cDNA，并发现与金属硫蛋白 (MT) 具有惊人的同源性。与 MT 基因相反，GIF 基因仅​​在神经系统中表达。 Northern 印迹分析显示，AD 大脑中 GIF mRNA 表达急剧下降。为了分析这种下调的机制，我们分离了人和小鼠的 GIF 基因。这些基因由三个外显子组成，长约 1 kb。人类和小鼠 GIF 基因的比较显示了几个保守序列，包括假定的 AP-2、SP-1、TATA 结合蛋白和金属反应元件 (MRE)。这些 GIF 基因的启动子中存在与人类 gfa 共同序列 (hgcs) 相似的序列，最近被鉴定为星形胶质细胞特异性转录因子的序列。由于 GIF 被认为是金属硫蛋白家族的新成员，与金属毒性作用的抵抗力相关，因此我们在小鼠原代培养物中研究了使用重金属和细胞因子调节 GIF 转录水平的效果 星形胶质细胞。 Cu^2+和Zn^2+的刺激诱导GIF，而IL-1和EGF的刺激则抑制GIF的转录。这些结果表明，多种生长因子或细胞因子调节GIF的合成，在神经生长方向和程度的调节中发挥重要作用。为了破坏小鼠胚胎干细胞中的 GIF 序列，我们制作了一个包含 6 kb 基因组片段的替换载体，其中包含整个 GIF 基因。第一个外显子的部分被删除，并且neo盒被插入到删除的位置。目前我们正在尝试制备GIF缺陷小鼠，以阐明GIF基因在体内的功能。

项目成果

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Tanno,Y.: "Quantitation of mitochondrial DNA carrying tRNA Lys mutation in MERRF patients." Biochem.Biophys.Res.Commun.179. 880-885 (1991)

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Kobayashi,H.et al.: "Molecular cloning of rat growth inhibitory factor cDNA and the expression in the central nervous system." Mol.Brain.Res.19(3). 188-94 (1993)

Kobayashi,H.et al.：“大鼠生长抑制因子 cDNA 的分子克隆及其在中枢神经系统中的表达。”

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