Single-cell dissection of neural stem cell heterogeneity in vivo

 描述（由申请人提供）：成人神经干细胞（NSC）在大脑的有限部位不断产生神经元，这些神经元在功能上整合到神经回路中。在成年人大脑中最大的生发区--脑室-脑室下区（V/SVZ）中可以发现大量的神经干细胞聚集，该区域产生不同亚型的嗅球神经元以及少突胶质细胞。最近的工作也表明，成人神经干细胞表现出区域异质性，在其后代。然而，目前对NSC的理解依赖于群体水平的研究，这些研究平均了不同的单细胞行为。因此，对体内单个神经干细胞的分子程序和多样性知之甚少。最近，Doetsch实验室已经定义了一种标记物的组合，该标记物允许直接从体内小生境中前瞻性地纯化静止和活化的成体NSC。我将结合联合收割机这种纯化策略与单细胞转录谱的神经干细胞从不同地区的V/SVZ。最后，我提议：1）通过聚类分离自V/SVZ不同区域的单个纯化的静止和活化的成体神经干细胞的单细胞转录谱来定义成体NSC的异质性，和2）验证新的成体NSC亚群的体内分布并推断不同NSC身份背后的细胞调节网络。基于单细胞基因表达谱的NSC分类将为未来利用这些细胞进行细胞免疫的努力提供巨大的资源。 重编程以及与来自其他脑区域的NSC或星形胶质细胞的功能比较。这种方法提供的单细胞分辨率将产生急需的关键洞察成人神经干细胞的分子和功能异质性。

 DESCRIPTION (provided by applicant): Adult neural stem cells (NSC) continuously generate neurons in restricted parts of the brain that functionally integrate into neural circuits. Significnt clusters of NSCs can be found in the ventricular-subventricular zone (V/SVZ), the largest germinal region in the adult brain, which generates different subtypes of olfactory bulb neurons, as well as oligodendrocytes. Recent work has also suggested that adult neural stem cells exhibit regional heterogeneity in terms of their progeny. However, the current understanding of NSCs has relied on population-level studies that average diverse single-cell behaviors. Therefore, little is known about the molecular programs and diversity of individual neural stem cells in vivo. Recently, the Doetsch lab has defined a combination of markers that allows the prospective purification of quiescent and activated adult NSCs directly from the in vivo niche. I will combine this purification strategy with single cell transcriptional profiling of NSCs from distinct regions of the V/SVZ. Ultimately, I propose to: 1) define the heterogeneity of adult NSCs by clustering single-cell transcriptional profiles of individual purified quiescent and activated adult neural stem cells isolated from different regions of the V/SVZ, and 2) validate the in vivo distribution of the novel adult NSC subpopulations and infer cellular regulatory networks underlying different NSC identities. The NSC classification based on single-cell gene expression profiles will serve as a tremendous resource for future efforts to harness these cells for cellular reprogramming and for functional comparisons with NSCs or astrocytes from other brain regions. The single cell resolution provided by this approach will yield much needed key insight into the molecular and functional heterogeneity of adult neural stem cells.