Abstract
Here we report the generation of a multimodal cell census and atlas of the mammalian primary motor cortex as the initial product of the BRAIN Initiative Cell Census Network (BICCN). This was achieved by coordinated large-scale analyses of single-cell transcriptomes, chromatin accessibility, DNA methylomes, spatially resolved single-cell transcriptomes, morphological and electrophysiological properties and cellular resolution input–output mapping, integrated through cross-modal computational analysis. Our results advance the collective knowledge and understanding of brain cell-type organization1–5. First, our study reveals a unified molecular genetic landscape of cortical cell types that integrates their transcriptome, open chromatin and DNA methylation maps. Second, cross-species analysis achieves a consensus taxonomy of transcriptomic types and their hierarchical organization that is conserved from mouse to marmoset and human. Third, in situ single-cell transcriptomics provides a spatially resolved cell-type atlas of the motor cortex. Fourth, cross-modal analysis provides compelling evidence for the transcriptomic, epigenomic and gene regulatory basis of neuronal phenotypes such as their physiological and anatomical properties, demonstrating the biological validity and genomic underpinning of neuron types. We further present an extensive genetic toolset for targeting glutamatergic neuron types towards linking their molecular and developmental identity to their circuit function. Together, our results establish a unifying and mechanistic framework of neuronal cell-type organization that integrates multi-layered molecular genetic and spatial information with multi-faceted phenotypic properties.
Highlights
Molecular definition of cell types in MOp A mouse MOp molecular taxonomy was derived from seven scRNA-seq and snRNA-seq datasets and single-nucleus methylcytosine sequencing and single-nucleus assay for transposase-accessible chromatin using sequencing datasets[37]
The combined sc/snRNA-seq datasets contained a large number of cells profiled using both droplet-based and deep full-length sequencing methods (Extended Data Table 1), resulting in a consensus transcriptomic taxonomy with the greatest resolution compared with other data types, including 90 neuronal and 116 total clusters or transcriptomic types (t-types)[37]
We further applied two computational approaches, SingleCellFusion (SCF) and LIGER, to combine the transcriptomic and epigenomic datasets and derive an integrated molecular taxonomy consisting of 56 neuronal cell types[37] (Fig. 1a)
Summary
Molecular definition of cell types in MOp A mouse MOp molecular taxonomy was derived from seven scRNA-seq and snRNA-seq (sc/snRNA-seq) datasets and single-nucleus methylcytosine sequencing (snmC-seq2) and single-nucleus assay for transposase-accessible chromatin using sequencing (snATAC-seq) datasets[37]. We generated a fine-grained areal and laminar distribution map of multiple MOp-ul projection neuron populations using retrograde tracing[44] (Extended Data Fig. 4a) In parallel with these tracer-labelled, projection- and layer-defined cell populations, we characterized the distribution patterns in MOp-ul of neuronal populations labelled in 28 Cre driver lines, including those from different IT (for example, Cux[2], Plxnd[1] and Tlx[3] driver lines), ET (Rbp[4], Sim[1] and Fezf2) and CT (Ntsr[1] and Tle4) subclasses with distinct laminar distributions[47,62]. We identified multiple full-morphology reconstructions of MOp L5 ET neurons from fMOST imaging of Fezf2-CreER;Ai166 and Pvalb-T2A-CreERT2;Ai166 transgenic mice, which were clustered into MY-projecting and non-MY projecting morphological types and exhibited extensive morphological and projectional variability among individual cells[46] (Fig. 8e), this was not directly linked a Mouse consensus transcriptomic taxonomy. We identified transcription factors showing cell-type specificity supported by both RNA expression and DNA-binding motif enrichment in cell subclasses[37,39] (Methods) (Extended Data Fig. 7)
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