Single cell transcriptomics to investigate monoallelic expression as a potential trigger of Alzheimer’s disease

阿尔茨海默病（AD）是与衰老相关的主要疾病之一，目前影响全球超过3500万人，并且预计发病率将增加。虽然AD的发病机制正在被深入研究，但仍不完全清楚。一些脑区域，如颞叶，比其他区域更早受到疾病的影响，这表明某些神经元对AD病理学的敏感性存在差异。这一观察结果可以解释为神经元之间的基因表达差异导致不同的表型表达。因此，单细胞水平的研究可能揭示AD发病的触发因素。随机单等位基因表达（RMAE）是一种机制，其中只有一个基因的等位基因表达。由于等位基因是随机选择的，这种基因表达模式可以在相同细胞类型的细胞之间产生变异性，并且可能是使一些神经元比其他神经元对发展AD更敏感的一种机制。RMAE已被证明涉及其他认知疾病，如精神分裂症和自闭症，以及神经发育障碍。AD相关基因在RMAE基因中显著富集，表明AD和RMAE之间存在联系。此外，AD特征性淀粉样前体蛋白（APP）是单等位基因表达的，可能导致不同细胞中APP的量不同。然而，在人类神经元中的RMAE的程度，如何在细胞中建立RMAE，以及在RMAE失调的后果几乎还没有被研究you.To调查的作用，RMAE在晚发性AD，我们建议序列的第一次约2000个单神经元的全长转录从5个健康和5个AD影响的个人。另外对相应个体的外显子组进行测序将使我们能够发现杂合SNP和RMAE。我们将建立一个计算管道，用于RMAE分析和RMAE变化的功能后果。利用大约2000个神经元的转录组数据，我们将描述这些细胞中的转录变异性和RMAE的程度。我们将测试这一假设，即在AD患者的神经元中，RMAE的模式发生了改变。使用最先进的比较转录组和共表达网络分析，我们的目标是揭示可能与AD相关的RMAE变化的功能后果。重要的是，我们将研究不同神经元类型的网络差异，以识别神经元类型特异性改变。此外，通过整合表观基因组、转录因子和长链非编码RNA的信息，我们还旨在为建立RMAE并可能导致其在AD中失调的机制提供重要见解。通过我们的单神经元分辨率，我们希望解开与AD发病和进展有关的新的候选基因和调控机制，到目前为止，通过对脑组织RNA进行批量测序无法找到这些基因和机制。

Alzheimer’s disease (AD) is one of the major diseases related to aging, currently affecting more than 35 million people worldwide, and expected to increase in incidences. Although being intensely investigated, the mechanisms of the onset of AD are still not fully understood. Some brain regions, such as the temporal lobe, are affected earlier by the disease than others, suggesting differences in sensitivity of certain neurons towards AD pathology. This observation might be explained by differences in gene expression between neurons leading to varying expressivity of the phenotype. Thus, investigations at the single cell level might reveal the trigger for the onset of AD. Random monoallelic expression (RMAE) is a mechanism, in which only one allele of a gene is expressed. Since the allele is randomly chosen, this gene expression mode can create variability between cells of the same cell type and might be one mechanism to render some neurons more sensitive than others to developing AD. RMAE has been shown to be involved in other cognitive diseases, such as schizophrenia and autism, and in neurodevelopmental disorders. AD-associated genes are significantly enriched among RMAE genes, suggesting a link between AD and RMAE. Moreover, the AD-characteristic amyloid precursor protein (APP) is expressed monoallelically, potentially leading to different amounts of APP in different cells. However, the extent of RMAE in human neurons, how RMAE is established in the cell, and consequences of dysregulation in RMAE have almost not been studied yet.To investigate the role of RMAE in late-onset AD, we propose to sequence for the first time the full-length transcripts of about 2000 single neurons from five healthy and five AD-affected individuals. Sequencing additionally the exomes of the respective individuals will allow us to discover heterozygous SNPs and RMAE. We will establish a computational pipeline for RMAE analysis and functional consequences of changes in RMAE. Utilizing the transcriptome data of about 2000 neurons, we will describe the transcriptional variability and the extent of RMAE in these cells. We will test the hypothesis, that patterns of RMAE are altered in neurons of individuals with AD. Using state-of-the-art comparative transcriptome and co-expression network analyses we aim to uncover functional consequences of changes in RMAE that might be related to AD. Importantly, we will investigate network differences for different neuron types to identify neuron-type specific alterations. In addition, integrating information of the epigenome, of transcription factors, and long non-coding RNAs, we also aim to provide important insights into mechanisms that establish RMAE and might lead to its dysregulation in AD. With our single neuron resolution, we expect to unravel new candidate genes and regulatory mechanisms involved in AD onset and progression, which so far were impossible to find by bulk sequencing of RNA from brain tissue.