Understanding the mechanism of SPl1 dependent Alzheimer disease risk

阿尔茨海默病（AD）是美国十大杀手中唯一没有疾病修饰疗法的疾病。因此，它也是唯一一种流行率正在上升的疾病。人类遗传学研究提供了一种强有力的手段来鉴定与疾病病因学有因果关系的基因和途径，并为药物发现产生新的治疗假设。过去几年的技术进步使大规模全基因组关联研究（GWAS）能够识别调节AD风险的常见变异，并通过全基因组/全外显子组测序识别与AD相关的罕见突变。这项工作导致发现了20多个与AD有因果关系的基因座（除APOE外）。我们在GWAS和测序研究中对与AD相关的遗传变异的系统水平分析表明，髓样细胞对细胞碎片的吞噬清除缺陷（巨噬细胞增多症）是AD病因学的重要组成部分，其他研究者对健康和AD人脑中的基因调控网络进行了分析。这些GWAS基因座之一的精细定位使我们确定了SPI 1中的一个常见变体（rs 1057233），该变体可降低SPI 1表达和AD风险。与ABCA 7和TREM 2（在红细胞增多症中起关键作用的另外两种确定的AD风险因子）一样，SPI 1在髓系免疫细胞中表达（例如，单核细胞、巨噬细胞和小胶质细胞）。SPI 1是一种转录因子（PU.1），对小胶质细胞的发育至关重要，并调节许多与AD相关基因的表达。我们推测，SPI 1表达的调节通过小胶质细胞内基因表达的整体变化，导致红细胞增多症的改变，从而影响AD的风险。我们将整合计算和实验方法来定义SPI 1功能变异降低SPI 1表达和AD风险的机制。为了研究这种保护作用，我们将首先寻求使用来自具有不同rs 1057233基因型的同基因人类iPSC细胞系的小胶质细胞在体外复制它（Aim 1）。我们还将在遗传上减少/增加小鼠大脑小胶质细胞中的SPI 1表达，并测量这些干预措施对体内分子、细胞和AD相关表型的影响（分别为目标2和3）。为了实现这些研究，我们最近开发了一种新的小鼠模型，该模型可用于分析大脑中小胶质细胞的核糖体结合转录组，同时也有条件地和特异性地下调或上调小胶质细胞中感兴趣的基因如SPI 1的表达。使用与AD小鼠模型交叉的相同模型，我们将在小胶质细胞中SPI 1表达减少或增加的背景下研究AD相关的结果，如小胶质细胞增生和β-淀粉样蛋白沉积。

Alzheimer's disease (AD) is the only disease among the top ten killers in the U.S. without a disease modifying therapy. As a result it is also the only one that is increasing in prevalence. Human genetic studies provide a powerful means to identify genes and pathways that are causally linked to the etiology of disease, and to generate new therapeutic hypotheses for drug discovery. Technological advances in the last few years have enabled large-scale genome-wide association studies (GWAS) to identify common variants that modulate AD risk, and whole genome/whole exome sequencing to identify rare mutations associated with AD. This work has led to the discovery of more than 20 loci (in addition to APOE) that are causally linked to AD. Our systems- level analysis of genetic variants associated with AD in GWAS and sequencing studies implicate defective phagocytic clearance of cellular debris by myeloid cells (efferocytosis), as an important component of the etiology of AD as have analyses of gene regulatory networks in healthy and AD human brains by other investigators. Fine mapping of one of these GWAS loci led us to identify a common variant (rs1057233) in SPI1, which reduces SPI1 expression and risk for AD. Like ABCA7 and TREM2, two other established AD risk factors that play key roles in efferocytosis, SPI1 is expressed in immune cells of the myeloid lineage (e.g., monocytes, macrophages and microglia). SPI1 is a transcription factor (PU.1) that is critical for microglial development and regulates expression of many of the AD-associated genes implicated in efferocytosis. We hypothesize that modulation of SPI1 expression influences AD risk through global changes in gene expression within microglia that lead to altered efferocytosis. We will integrate computational and experimental approaches to define the mechanism(s) by which functional variation in SPI1 reduces SPI1 expression and risk for AD. To investigate this protective effect, we will first seek to replicate it in vitro using microglial cells derived from isogenic human iPSC cell lines with different rs1057233 genotypes (Aim 1). We will also genetically decrease/increase SPI1 expression in microglial cells of the mouse brain and measure the effect of these interventions on molecular, cellular and AD-related phenotypes in vivo (Aim 2 and 3 respectively). To enable these studies, we have recently developed a novel mouse model that can be used to profile the ribosome-bound transcriptome of microglial cells in the brain while also conditionally and specifically down- or up-regulating the expression of a gene of interest like SPI1 in microglia. Using the same model crossed with an AD mouse model, we will investigate AD-related outcomes like micro-gliosis and ß-amyloid deposition in the context of reduced or increased SPI1 expression in microglia.