Troy+ brain stem cells cycle through quiescence and regulate their number by sensing niche occupancy

The adult mouse subependymal zone (SEZ) provides a niche for mammalian neural stem cells (NSCs). However, the molecular signature, self-renewal potential and fate behavior of NSCs remain poorly defined. Here we propose a model in which the fate of active NSCs is coupled to the total number of neighboring NSCs in a shared niche. Using knock-in reporter alleles and single-cell RNA sequencing, we show that the Wnt target Tnfrsf19/Troy identifies both active and quiescent NSCs. Quantitative analysis of genetic lineage tracing of individual NSCs under homeostasis or in response to injury reveals rapid expansion of stem cell number before some return to quiescence. This behavior is best explained by stochastic fate decisions, where stem cell number within a shared niche fluctuates over time. Fate-mapping proliferating cells using a novel Ki67iresCreER allele confirms that active NSCs reversibly return to quiescence, achieving long-term self-renewal. Our findings suggest a novel niche-based mechanism for the regulation of NSC fate and number. Overall design: We analysed the adult mouse subependymal zone (SEZ), the largest neruogenic niche in the brain, at a single cell level. For this, we used SORT-seq, a combination of FACS and single cell RNA-sequencing that resulted in high quality transcriptome data from ~1500 single cells (out of ~2500 cells sequenced). To identify different cell types, we used knock-in mouse models, surface labeling using antibdoies as well as lineage tracing. Troy-GFPiresCreER +/ki mice were used to identify NSCs. Ki67-tagRFP ki/ki mice were used to identify dividing cells. Ki67-iresCreER +/ki Rosa-tdTomato +/ki mice were administered a single injection of 5mg of Tamoxifen to label dividing cells (day2 - d2) and follow their progeny (day60 - d60) in the SEZ as well as the olfactory bulb (for d60 only). We also included published markers; combinations of anti-GLAST antibody (to identify NSCs) and EGF binding ability using alexa647-conjugated EGF (to identify dividing cells), combinations of anti-Prom1 (Cell contacting the ventricles) and anti-CD24 antibodies (high in neuroblasts), as well as anti-O4 antibody (oligodendrocytes) to create an atlas of NSCs.