AUTOPHAGY, DNA DAMAGE AND ONSET OF HUNTINGTON'S DISEASE.

自噬、DNA 损伤和亨廷顿病的发作。

• 批准号: 7085095
• 负责人: JUNYING YUAN
• 金额: $ 33.83万
• 依托单位: HARVARD MEDICAL SCHOOL
• 依托单位国家: 美国
• 财政年份: 2006
• 资助国家: 美国
• 起止时间: 2006-04-01 至 2011-03-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/7085095
• 关键词: DNA damage; Huntington' s disease; aging; amidohydrolases; autophagy; cell line; cyclin dependent kinase; disease /disorder onset; free radical oxygen; gene expression; gene induction /repression; gene mutation; genetic promoter element; genetically modified animals; laboratory mouse; mitochondria; nerve /myelin protein; neural degeneration; neurogenetics; neuropathology; oxidative stress; transcription factor

The goal of this proposal is to test the hypothesis that the reduction of key autophagy gene expression and autophagy function as a result of DNA damage during the aging process plays a key role in mediating the onset of Huntington's disease (HD). Mouse, Drosophila and C. elegans models of HD suggest that the cytotoxicity of expanded polyglutamine is highly dependent upon protein context and protein expression levels of mutant Htt. Autophagy plays an important role in regulating the intracellular accumulation of mutant Htt with expanded polyQ. The expression of beclin 1, a key gene involved in autophagy, decreased in an age-dependent fashion in human brains. Since beclin 1 gene is haploid insufficient in regulating autophagosome function, age-dependent decrease of beclin 1 expression may lead to a reduction of autophagic activity during aging. The hypothesis is that reduction of autophagy function in aging results in both increased oxidative stress induced DNA damage and reduced long-lived protein turnover which promotes the accumulation of mutant Htt. Increased accumulation of mutant Htt and oxidative stress may play an important role in promoting the onset of HD. This hypothesis will be tested in the following specific aims. Specific Aim 1 is to test,the hypothesis that the reduction of beclin 1 expression in aging human brains contributes to the onset of HD by generating beclin 1+/-; HdhQ111 mice and examining if 50% reduction of beclin 1 expression led to an increased accumulation of mutant Htt as well as to determine the consequence of autophagy deficiency on neuronal survival and functions. Specific Aim 2 is to examine the mechanism which led to the age-dependent reduction of beclin 1 expression by testing if the promoter of beclin 1 is preferentially damaged in aging human brains and particularly susceptible to oxidative damage in cellular models. The contribution of reduced expression of transcriptional factors regulating beclin 1 expression will also be considered. Specific Aim 3 is to test the hypothesis that the reduction of autophagy function exacerbates the DNA damage during aging by increasing the accumulation of damaged mitochondria which further promotes the levels of intracellular ROS by examining aging beclin 1+/- mice and autophagy deficient cells for evidence of increased damaged mitochondria. Specific Aim 4 is to investigate the functional role of autophagy to oxidative DNA damage using CK-p25 mice as a model and to examine the roles of of DNA damage and autophagy deficiency to the accumulation of mutant Htt in HdhQ111; CKp25 mice. The ability of SIRT1 activating molecules (STACs) to restore the autophagy function in CK-p25 mice and to delay the onset of motor dysfunction in HD models will be determined. Understanding the mechanism by which DNA damage negatively regulates autophagy during aging would allow us to develop strategies to maintain normal autophagy function during aging process which may delay or prevent the onset of HD and other aging related neurodegenerative diseases.

该提案的目的是检验以下假设：关键自噬基因表达的减少和 衰老过程中 DNA 损伤导致的自噬功能在介导衰老过程中发挥着关键作用 亨廷顿病 (HD) 发作。小鼠、果蝇和秀丽隐杆线虫的 HD 模型表明 扩展的聚谷氨酰胺的细胞毒性高度依赖于蛋白质背景和蛋白质表达 突变体Htt的水平。自噬在调节细胞内突变体的积累中发挥着重要作用 Htt 与扩展的 PolyQ。参与自噬的关键基因 beclin 1 的表达在 人类大脑中与年龄相关的时尚。由于beclin 1基因是单倍体，不足以调节 自噬体功能，年龄依赖性 beclin 1 表达的减少可能导致 衰老过程中的自噬活动。该假设认为，衰老过程中自噬功能的降低会导致 氧化应激增加会导致 DNA 损伤，并减少长寿命蛋白质更新，从而 促进突变体Htt的积累。突变体 Htt 和氧化应激积累的增加可能 对HD的发生起着重要的促进作用。该假设将在以下具体情况中得到检验 目标。具体目标 1 是检验衰老人脑中 beclin 1 表达减少的假设 通过产生 beclin 1+/- 促进 HD 的发生； HdhQ111 小鼠并检查是否减少 50% beclin 1 表达导致突变体 Htt 积累增加，并确定 自噬缺陷对神经元存活和功能的影响。具体目标 2 是检查 通过测试是否启动子，导致 beclin 1 表达随年龄而减少的机制 beclin 1 在衰老的人类大脑中优先受损，并且特别容易受到氧化损伤 细胞模型。调节 beclin 1 的转录因子表达减少的贡献 表达也会被考虑。具体目标 3 是检验自噬减少的假设 功能通过增加受损细胞的积累，加剧衰老过程中的 DNA 损伤 线粒体通过检查衰老的 beclin 1+/- 小鼠进一步提高细胞内 ROS 水平 自噬缺陷细胞寻找线粒体受损增加的证据。具体目标 4 是调查 以 CK-p25 小鼠为模型，研究自噬对 DNA 氧化损伤的功能作用并进行检验 DNA损伤和自噬缺陷对HdhQ111突变体Htt积累的作用；肌酸激酶p25 老鼠。 SIRT1 激活分子 (STAC) 恢复 CK-p25 自噬功能的能力 小鼠并延迟 HD 模型中运动功能障碍的发生将被确定。了解 衰老过程中 DNA 损伤对自噬负调节的机制将使我们能够开发 在衰老过程中维持正常自噬功能的策略可能会延迟或预防发生 HD 和其他与衰老相关的神经退行性疾病。