Project description:Developmental signaling inputs are fundamental for shaping cell fates and behavior. However, traditional fluorescent-based signaling reporters have limitations in scalability and molecular resolution of cell types. We present SABER-seq, a CRISPR-Cas molecular recorder that stores transient developmental signaling cues as permanent mutations in cellular genomes for deconstruction at later stages via single-cell transcriptomics. We applied SABER-seq to record Notch signaling in developing zebrafish brains. SABER-seq has two components: a signaling sensor and a barcode recorder. The sensor activates Cas9 in a Notch-dependent manner with inducible control while the recorder obtains mutations in ancestral cells where Notch is active. We combine SABER-seq with an expanded juvenile brain atlas to identify cell types derived from Notch-active founders. Our data reveals rare examples where differential Notch activity in ancestral progenitors is detected in terminally differentiated neuronal subtypes. SABER-seq is a novel platform for rapid, scalable and high-resolution mapping of signaling activity during development.
Project description:Gene expression is finely regulated during development, and deregulation can lead to disease. In pediatric brain tumors (PBT), disruption of neurodevelopmental gene regulation programs are suspected to drive oncogenesis. However, the transcriptional landscape and genetic regulation processes of the healthy developing brain are not fully characterized, limiting our investigation of these tumors. We used single-cell RNA-sequencing to generate a transcriptomic atlas of >65,000 cells in the developing forebrain and pons in human and mouse, two regions where PBT commonly arise. We projected bulk RNA-seq profiles for a cohort of 198 PBT onto these cell types, followed by focused analysis of three PBT subtypes by single-cell profiling: WNT medulloblastoma, embryonal tumors with multilayered rosettes (ETMR), and atypical teratoid/rhabdoid tumors (ATRT). Altogether, we pinpoint stalled differentiation during developmental programs as a common etiological mechanism of PBT, providing a valuable resource to aid modeling and therapeutics.
Project description:Illustrating the cellular architecture of the mammalian brain is critical to understanding its diverse functions and complex animal behaviors. With support from NIMH and NINDS as part of the BRAIN Initiative Cell Census Network (BICCN), the Center for Epigenomics of the Mouse Brain Atlas (CEMBA) applied single nucleus methylation sequencing in 118 (45 in the current release) brain functional regions across the whole adult male P56 C57BL/6J mouse brain. In each major brain regions, we identified distinct cell clusters and formed them in a hierarchical taxonomy. These clusters include known primary brain cell types and possible sub-types. We use these data to identify the epigenomic characteristics and to define specific regulatory elements for each cell cluster. We integrate the single-cell methylome with single-cell gene expression and chromatin accessibility profiles from the same brain region to further identify potential enhancers and their corresponding genes for several neuronal cell types. By brain region spacial comparisons, we found spatial specificity among excitatory neuronal clusters within the same cortical layer. We identify brain-region-specific transcription factors that may regulate cortical region development of excitatory neurons. These data define the landscape of cell type and spatial heterogeneity in the mouse brain and the underlining regulatory epigenomic basis.