Fate restriction governs regional astrocyte allocation during brain development

Astrocytes within specific brain regions contribute uniquely to regional circuits for higher-order brain function through interactions with local neurons. The regional diversification of astrocytes is dictated by their embryonic origin, yet the mechanisms governing their regional allocation remain unknown. Here we show that allocation of astrocytes to specific brain regions requires the transcription factor 4 (Tcf4) mediated fate restriction during brain development. Loss of Tcf4 in ventral telencephalic neural progenitors alters the fate of oligodendrocyte precursors to transient intermediate astrocyte precursor cells, resulting in mislocated astrocytes in the dorsal neocortex. These ectopic astrocytes originated from the ventral telencephalon engage with neurons and acquire features reminiscent of local neocortical astrocytes. Furthermore, Tcf4 functions as a suppressor of astrocyte fate during differentiation of oligodendrocyte precursors, thereby restricting the fate to oligodendrocyte lineage. Our study reveals that fate restriction governs regional astrocyte allocation, contributing to astrocyte diversification across brain regions. For all experiments, cells from the Nkx2.1 lineage in the neocortex from control and Tcf4 cKO mice were collected by FACS sorting. After cell variablity checking, single cell sequencing library were prepared using 10x Genomics Chromium Single Cell 3`v3 assays, or ATACseq and cut and tag were performed.特定脑区中的星形胶质细胞通过与局部神经元的相互作用，对介导高阶脑功能的区域环路发挥独特调控作用。星形胶质细胞的区域多样性由其胚胎起源决定，但其区域分配的调控机制仍不明朗。 本研究表明，在脑发育过程中，星形胶质细胞向特定脑区的分配依赖于转录因子4（transcription factor 4, Tcf4）介导的命运限制。腹侧端脑神经前体细胞中Tcf4的缺失，会将少突胶质细胞前体细胞的命运重定向为短暂中间态星形胶质细胞前体细胞，最终导致背侧新皮层中出现异位分布的星形胶质细胞。这些源自腹侧端脑的异位星形胶质细胞可与神经元相互作用，并获得与局部新皮层星形胶质细胞相似的特征。此外，在少突胶质细胞前体细胞的分化过程中，Tcf4可作为星形胶质细胞命运的抑制因子，从而将细胞命运限制在少突胶质细胞谱系内。 本研究揭示，命运限制机制调控星形胶质细胞的区域分配，进而促成脑区内星形胶质细胞的多样性形成。 所有实验均采用荧光激活细胞分选（Fluorescence-Activated Cell Sorting, FACS）技术，收集对照组与Tcf4条件性敲除（conditional knockout, cKO）小鼠新皮层内源自Nkx2.1谱系的细胞。在完成细胞异质性检测后，使用10x Genomics Chromium单细胞3'端测序v3实验体系构建单细胞测序文库，或开展ATAC测序（Assay for Transposase-Accessible Chromatin using sequencing, ATAC-seq）与切割与标签测序（Cleavage Under Targets and Tagmentation, CUT&Tag）实验。