Linking genotype to phenotype in autism: mechanisms of cell-type specific presynaptic dysfunction in Chd8 haploinsufficiency

将自闭症基因型与表型联系起来：Chd8 单倍体不足中细胞类型特异性突触前功能障碍的机制

基本信息

批准号：
MR/X010481/1
负责人：
Laura Andreae
金额：
$ 65.66万
依托单位：
King's College London
依托单位国家：
英国
项目类别：
Research Grant
财政年份：
2023
资助国家：
英国
起止时间：
2023 至无数据
项目状态：
未结题

来源：
https://gtr.ukri.org/projects?ref=MR%2FX010481%2F1
关键词：
Linking genotype phenotype autism mechanisms

项目摘要

Autism is a neurodevelopmental disorder for which there is a severe lack of effective treatments for those who need them. A major problem in the development of new treatments is our poor understanding of its underlying biology. While there is clearly a strong genetic component, we now know that there are hundreds of different gene mutations that increase the risk of developing autism. Many of these risk genes are involved in synapse function or formation, suggesting that synapses (the site of communication between neurons in the brain) may be a key part of autism biology. However, an even larger group of autism risk genes simply regulate the expression of other genes. What happens downstream of these genes? Might they also affect synaptic function or formation? In this proposal, we aim to identify the molecules and pathways that link gene mutation in one of these 'regulator genes' to changes in synapse function, in a part of the brain that is well known to be affected in autism, the prefrontal cortex. We will focus on a specific gene called Chd8, which is one of the highest confidence risk genes for autism, and where we have already identified a robust synaptic phenotype in prefrontal cortex. In particular, there is increasing evidence that the effects of missing a copy of Chd8 (and other autism risk genes) depends critically on which type of cell is affected. We will use transgenic technology to selectively delete a copy of Chd8 from excitatory neurons or different types of inhibitory neurons and find out exactly how synapse function and/or formation is affected. We will then collect individual cells of each type - excitatory and inhibitory - and use single nuclei sequencing to discover which genes show either increased or decreased expression when that cell type is missing a copy of Chd8. While this kind of approach usually results in many different genes that show changes, we will focus on those that we already know are involved in synaptic function / formation. We will carry out extensive validation of potential targets, including testing whether simply increasing or decreasing expression of the candidate in cortical neurons in a dish leads to the expected alteration to synaptic phenotype. Finally, we will take advantage of recent advances in CRISPR/Cas9 technology (the so-called genetic 'scissors' that allow researchers to edit DNA) to bring expression levels of candidate genes back to normal in the living brain, and see whether this can rescue the synaptic phenotypes.This will allow us to work out the molecules and pathways that link the original genetic mutation to changes in cell and synapse function, thereby helping to fill a major gap in our understanding of this disorder. These molecules and pathways may provide novel targets for therapeutic intervention, potentially opening up new avenues in the effort to develop new treatments.

自闭症是一种神经发育障碍，对需要患者的人严重缺乏有效的治疗方法。开发新疗法的一个主要问题是我们对其潜在生物学的理解不足。虽然显然有强大的遗传成分，但我们现在知道，有数百种不同的基因突变会增加自闭症的风险。这些风险基因中的许多都参与突触功能或形成，这表明突触（大脑中神经元之间的通信位点）可能是自闭症生物学的关键部分。但是，更大的自闭症风险基因只是调节其他基因的表达。这些基因下游会发生什么？它们是否还会影响突触功能或形成？在此提案中，我们旨在确定这些“调节基因”之一将基因突变与突触功能变化联系起来的分子和途径，这是大脑的一部分，众所周知在自闭症中受影响的前额叶皮层。我们将重点关注一个称为CHD8的特定基因，该基因是自闭症的最高置信风险基因之一，并且我们已经确定了前额叶皮层中强大的突触表型。特别是，有越来越多的证据表明，缺少CHD8（和其他自闭症风险基因）副本的影响取决于受影响的细胞类型。我们将使用转基因技术从兴奋性神经元或不同类型的抑制性神经元中选择性删除CHD8的副本，并确切地找出突触功能和/或形成如何影响。然后，我们将收集每种类型的单个细胞 - 兴奋性和抑制性 - 并使用单核测序发现当该细胞类型缺少CHD8的副本时，哪些基因显示出增加或降低的表达。尽管这种方法通常会导致许多不同的基因显示出变化，但我们将重点介绍我们已经知道的那些与突触功能 /形成有关的基因。我们将对潜在靶标进行广泛的验证，包括测试菜肴中皮质神经元中候选者的表达或降低导致突触表型的预期改变。最后，我们将利用CRISPR/CAS9技术的最新进展（所谓的遗传“剪刀”，使研究人员能够编辑DNA）使候选基因的表达水平恢复到活体大脑中，并查看这是否可以挽救突触的表型，这是否可以使我们能够弥补分子和遗传突变，从而使我们能够弥补遗传突变，从而弥补了遗传突变，从而在元素中弥补了突变，从而在元素中变化了，该突变是在元素中的变化，使其在功能中逐渐变化，并在其上弥补了构成的功能，使其在功能中逐渐变化，并在其上弥补了良好的范围，该杂物是在范围内的变化，使我们能够在范围内进行综合，从而使我们能够链接出来。这个疾病。这些分子和途径可能为治疗干预提供新的目标，并有可能在开发新疗法的努力中开放新的途径。