Single-cell dissection of neural stem cell heterogeneity in vivo

 描述(申请人提供)：成人神经干细胞(NSC)在大脑有限的部分持续产生神经元，这些神经元在功能上整合到神经回路中。脑室-室下区(V/SVZ)是成人脑内最大的生发区，可产生不同亚型的嗅球神经元和少突胶质细胞。最近的研究还表明，成年神经干细胞在其后代方面表现出区域异质性。然而，目前对神经干细胞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，人们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实验室定义了一种标记的组合，可以直接从体内的利基中分离纯化静止和激活的成年神经干细胞。我将把这种纯化策略与来自V/SVZ不同区域的NSCs的单细胞转录图谱结合起来。最终，我建议：1)通过聚类从V/SVZ不同区域分离的单个纯化的静止和激活的成人神经干细胞的单细胞转录谱来确定成体NSC的异质性；2)验证新的成体NSC亚群的体内分布，并推断不同NSC身份的细胞调控网络。基于单细胞基因表达谱的NSC分类将成为未来利用这些细胞进行细胞学研究的巨大资源 重新编程，并与来自其他脑区的神经干细胞或星形胶质细胞进行功能比较。这种方法提供的单细胞分辨率将为成人神经干细胞的分子和功能异质性提供急需的关键洞察力。

 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.