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N-terminal huntingtin and Huntington disease neuropathology

N 末端亨廷顿蛋白和亨廷顿病神经病理学

基本信息

批准号：
10117290
负责人：
Andrew P Escayg
金额：
$ 44.84万
依托单位：
EMORY UNIVERSITY
依托单位国家：
美国
项目类别：
财政年份：
2017
资助国家：
美国
起止时间：
2017-03-01 至 2023-02-28
项目状态：
已结题

来源：
https://reporter.nih.gov/project-details/10117290
关键词：
Address Adult Affect Age Amino Acids Animals Antisense Oligonucleotides Brain Brain Diseases Brain region CRISPR/Cas technology Cell Nucleus Cells Conserved Sequence Corpus striatum structure Development Disease Embryo Embryonic Development Event Excision Gene Targeting Genes Genetic Transcription Genets Guide RNA Huntington Disease Huntington gene Huntington protein Impairment Injections Intervention Knock-in Mouse Knockout Mice Lead Length Mediating Methods Motor Mus Mutation N-terminal Nerve Degeneration Neurodegenerative Disorders Neurologic Neurons Nuclear Pathogenesis Pathologic Pathology Patients Phenotype Proteins Proteolysis Small Interfering RNA Testing Therapeutic Therapeutic Effect Toxic effect age related experimental study genome editing inhibitor/antagonist insight mutant neuronal survival neuropathology neurotoxic novel therapeutic intervention offspring polyglutamine preservation protective effect side effect therapeutically effective tool treatment strategy

项目摘要

Huntington's disease (HD) is a devastating neurodegenerative disease caused by expansion of a polyglutamine (polyQ) domain in distinct proteins with different functions. In HD, the polyQ domain is located in the N-terminal region of huntingtin (Htt). This N-terminal region is well conserved in a wide range of species, but polyQ expansion can lead to misfolding and subsequent toxicity of N-terminal fragments of Htt. Since a lack of Htt causes embryonic lethality in mice, Htt is also thought to be essential for animal development and survival. Reducing the expression of mutant Htt is widely accepted as an important strategy for treating HD, so considerable efforts have gone into developing siRNA and anti-sense oligonucleotides to suppress the expression of mutant Htt. These approaches have also raised concerns that markedly suppressing Htt expression could lead to side effects by diminishing the normal function of Htt; however, whether Htt can preserve critical functions without the N-terminal domain that contains the polyQ domain remains unknown. Addressing this issue is important if we are to develop a new strategy to treat HD: if the N-terminal polyQ domain is not required for essential Htt functions and can be removed, complete elimination of the N-terminal region of Htt is now possible since the recent development of the genomic editing tool, CRISPR/Cas9. In this competitive renewal application, we will use CRISPR/Cas9 to investigate the toxicity of N-terminal mutant Htt fragments and therapeutic effects by removing the polyQ-containing N-terminal region. In Aim 1, we will use CRISPR/Cas9 to introduce mutations in the mouse Htt gene in embryos from HD 140Q KI mice to generate truncated mutant Htt genes that express different N-terminal Htt fragments and can be transmitted to offspring via the germline. Using the newly established HD KI mice that express different N-terminal mHtt fragments containing the same polyQ repeat (140Q) at the endogenous level, we will examine the relationship between the length of N-terminal mutant Htt fragments and their nuclear accumulation and toxicity in striatal neurons. In Aim 2, we will use CRISPR/Cas9 to remove the N-terminal polyQ domain as a therapeutic strategy. We will explore whether removing the N-terminal polyQ domain in Htt can eliminate neuropathology without affecting neuronal survival and function in adult mice. These studies will use HD knock-in mice in which mutant Htt is expressed at the same endogenous level as in HD patients. We hope these studies will not only provide new insight into the pathogenesis of N-terminal mutant Htt fragments, but also allow us to develop a novel therapeutic strategy to treat Huntington's disease and other polyQ diseases.

亨廷顿氏病（HD）是一种破坏性的神经退行性疾病，多聚谷氨酰胺（polyQ）结构域在不同的蛋白质具有不同的功能。在HD中，polyQ结构域位于亨廷顿蛋白的N-末端区域（Htt）。该N-末端区域在广泛的物种中是很保守的，但polyQ扩增可导致Htt的N-末端片段的错误折叠和随后的毒性。由于缺乏Htt会导致小鼠胚胎死亡，Htt也被认为是动物发育所必需的，生存降低突变型Htt的表达被广泛接受为治疗HD的重要策略，因此，相当大的努力已经投入到开发siRNA和反义寡核苷酸以抑制突变体Htt的表达。这些方法也引起了人们的担忧，表达可能通过减少Htt的正常功能而导致副作用;然而，Htt是否可以保留关键功能而不包含polyQ结构域的N-末端结构域仍然未知。如果我们要开发治疗HD的新策略，解决这个问题是重要的：如果N-末端polyQ 结构域不是必需的Htt功能所必需的，并且可以被去除，完全消除N-末端由于最近开发了基因组编辑工具CRISPR/Cas9，因此Htt区域现在是可能的。在这竞争性更新申请，我们将使用CRISPR/Cas9来研究N-末端突变体Htt的毒性片段和治疗效果。在目标1中，我们将使用 CRISPR/Cas9在来自HD 140 Q KI小鼠的胚胎中的小鼠Htt基因中引入突变，以产生表达不同N末端Htt片段并可遗传给后代的截短突变Htt基因通过种系。使用新建立的表达不同N端mHtt片段的HD KI小鼠，在内源性水平上含有相同的polyQ重复序列（140 Q），我们将研究 N-末端突变Htt片段的长度及其在纹状体神经元中的核积累和毒性。在目的2，我们将使用CRISPR/Cas9去除N端polyQ结构域作为治疗策略。我们将探索去除Htt中的N末端polyQ结构域是否可以消除神经病理学而不影响神经元存活和功能。这些研究将使用HD基因敲入小鼠，其中突变型Htt 表达水平与HD患者相同。我们希望这些研究不仅能提供新的深入了解N-末端突变Htt片段的发病机制，也使我们能够开发一种新的治疗亨廷顿病和其他polyQ疾病的治疗策略。

项目成果

期刊论文数量（20）

专著数量（0）

科研奖励数量（0）

会议论文数量（0）

专利数量（0）

Promoting Cas9 degradation reduces mosaic mutations in non-human primate embryos.

促进 Cas9 降解减少非人类灵长类动物胚胎中的嵌合突变

DOI：
10.1038/srep42081
发表时间：
2017-02-03
期刊：
Scientific reports
影响因子：
4.6
作者：
Tu Z;Yang W;Yan S;Yin A;Gao J;Liu X;Zheng Y;Zheng J;Li Z;Yang S;Li S;Guo X;Li XJ
通讯作者：
Li XJ

HAP1 Is Required for Endocytosis and Signalling of BDNF and Its Receptors in Neurons.

DOI：
10.1007/s12035-016-0379-0
发表时间：
2018-03
期刊：
Molecular neurobiology
影响因子：
5.1
作者：
Lim Y;Wu LL;Chen S;Sun Y;Vijayaraj SL;Yang M;Bobrovskaya L;Keating D;Li XJ;Zhou XF
通讯作者：
Zhou XF

CRISPR: Established Editor of Human Embryos?