An epigenome atlas of neural progenitors within the embryonic mouse forebrain

Abstract

A comprehensive characterization of epigenomic organization in the embryonic mouse forebrain will enhance our understanding of neurodevelopment and provide insight into mechanisms of neurological disease. Here we collected single-cell chromatin accessibility profiles from four distinct neurogenic regions of the embryonic mouse forebrain using single nuclei ATAC-Seq (snATAC-Seq). We identified thousands of differentially accessible peaks, many restricted to distinct progenitor cell types or brain regions. We integrated snATAC-Seq and single cell transcriptome data to characterize changes of chromatin accessibility at enhancers and promoters with associated transcript abundance. Multi-modal integration of histone modifications (CUT&Tag and CUT&RUN), promoter-enhancer interactions (Capture-C) and high-order chromatin structure (Hi-C) extended these initial observations. This dataset reveals a diverse chromatin landscape with region-specific regulatory mechanisms and genomic interactions in distinct neurogenic regions of the embryonic mouse brain and represents an extensive public resource of a ‘ground truth’ epigenomic landscape at this critical stage of neurogenesis.

Spatial enhancer activation determines inhibitory neuron identity

“…To select enhancers active in the developing forebrain, we intersected MEIS1/2-DLX5 co-binding sites from ChIP-seq data with the VISTA in vivo enhancer database (Visel et al, 2007) (Figure S3e). Additionally, we confirmed the accessibility of the respective genomic regions, utilizing published scATAC-seq data of the LGE and MGE (Rhodes et al, 2022) (Figure 4a). First, we chose two enhancers (hs1080 and hs956) of the TF Foxp2 , which both contained MEIS/DLX motifs with a spacing of 3 bps (Visel et al, 2007; Visel et al, 2013) (Figure 4a, Figure S4a, b, d, e).…”

“… a , Representative profiles of MEIS1/2 (red) and DLX5 (blue) ChIP-seq at E14.5 and E13.5 respectively, as well as scATAC-seq from LGE (dark gray) and MGE (gray) at E12.5 are shown at the Foxp2 gene locus. DLX5 ChIP-seq data from (Lindtner et al, 2019); scATAC-seq data from (Rhodes et al, 2022). b , Luciferase activity driven by the enhancer hs1080, co-transfected with Meis2 and Dlx5 expression vectors in Neuro2a cells.…”

“…b-c , LacZ expression in the LGE of E12.5 embryos driven by the enhancers hs1041 and hs956 (Visel et al, 2007). d-f , Representative tracks of MEIS1/2 ChIP-seq in the GE at E14.5 (red), DLX5 ChIP-seq in the GE at E13.5 (blue) (Lindtner et al, 2019), LHX6 ChIP-seq in the GE at E13.5 (purple) (Sandberg et al, 2016) and scATAC-seq in LGE (dark gray) and MGE (gray) at E12.5 (Rhodes et al, 2022). g-i , Luciferase activity driven by hs1041, hs956, and hs748 enhancers co-transfected with Meis2 , Dlx5 , and Lhx6 expression vectors in Neuro2a cells.…”

Generation of single-cell and single-nuclei suspensions from embryonic and adult mouse brains

“…Notably, these single-cell and single-nuclei preparations can be used for a variety of downstream applications such as ChIP-seq, CUT&Tag and chromatin confirmation technologies. 2 These preparations may also be useful for a number of recently developed single-cell multimodal assays that are compatible with the 10x Genomics platform such as Perturb-seq, 10 , 11 CITE-seq, 12 ASAP-seq 13 and scCUT&Tag-pro, 14 although we have not tested these protocols on these specific assays. Fluorescence-activated cell sorting (FACS) can (and in many cases should) be performed to harvest fluorescent cells or nuclei from suspensions to enrich for specific neuronal subtypes and/or eliminate debris.…”

“…In this protocol, we describe procedures to generate single-cell or single-nuclei suspensions from both embryonic and juvenile/adult mouse brains. 1 , 2 , 3 , 4 , 5 While our focus is the forebrain, we performed this procedure in the spinal cord as well. 9 We provide additional detail to prepare the samples for sequencing on the 10x Genomics platform, specifically with the single-cell/nuclei RNA-seq, single-nuclei ATAC-seq (chromatin accessibility) and single-nuclei Multiome (RNA-seq and chromatin accessibility) kits.…”