Abstract
The three-dimensional (3D) structure of chromatin is intrinsically associated with gene regulation and cell function1–3. Methods based on chromatin conformation capture have mapped chromatin structures in neuronal systems such as in vitro differentiated neurons, neurons isolated through fluorescence-activated cell sorting from cortical tissues pooled from different animals and from dissociated whole hippocampi4–6. However, changes in chromatin organization captured by imaging, such as the relocation of Bdnf away from the nuclear periphery after activation7, are invisible with such approaches8. Here we developed immunoGAM, an extension of genome architecture mapping (GAM)2,9, to map 3D chromatin topology genome-wide in specific brain cell types, without tissue disruption, from single animals. GAM is a ligation-free technology that maps genome topology by sequencing the DNA content from thin (about 220 nm) nuclear cryosections. Chromatin interactions are identified from the increased probability of co-segregation of contacting loci across a collection of nuclear slices. ImmunoGAM expands the scope of GAM to enable the selection of specific cell types using low cell numbers (approximately 1,000 cells) within a complex tissue and avoids tissue dissociation2,10. We report cell-type specialized 3D chromatin structures at multiple genomic scales that relate to patterns of gene expression. We discover extensive ‘melting’ of long genes when they are highly expressed and/or have high chromatin accessibility. The contacts most specific of neuron subtypes contain genes associated with specialized processes, such as addiction and synaptic plasticity, which harbour putative binding sites for neuronal transcription factors within accessible chromatin regions. Moreover, sensory receptor genes are preferentially found in heterochromatic compartments in brain cells, which establish strong contacts across tens of megabases. Our results demonstrate that highly specific chromatin conformations in brain cells are tightly related to gene regulation mechanisms and specialized functions.
Highlights
Consolidation[12], and DNs are activated during cue-guided reward-based learning[13]
A detailed flowchart of immunoGAM quality control (QC) measures and normalization is shown in Extended Data Fig. 1a–d and Supplementary Table 2
Calculated local contact densities and topological domains using the insulation square method[14], and calculated compartments associated with open chromatin and closed chromatin using principal component analysis (PCA)[2] (Supplementary Tables 3–5)
Summary
Synaptic plasticity (e.g., Egr[1], Neurod[1], Ctcf Dlg[4], Grik2) Neurod[2], Egr. PGN-expressed gene DN-expressed gene RNA transcript and other expressed genes. e, Differential contacts with the most abundant TF feature pairs in DNs contain differentially expressed genes (top) with DN-specific functions (middle; one-sided Fisher’s exact permuted P < 0.01). Egr[1] was highly upregulated in PGNs (log2(fold-change) = 3, PGNs compared to DNs) and gained contacts with its adjacent TAD It contained accessible chromatin peaks rich in TF motifs belonging to the Neurod group that are not seen in DNs. Binding of EGR1 protein to its own promoter is confirmed in published chromatin immunoprecipitation with sequencing (ChIP-seq) data from the cortex[31]. Our strategy identifies hubs of chromatin contacts specific for different neuron types that contain putative binding sites for differentially expressed TFs (Fig. 4g) These interconnected hubs bring together distal genes with specialized neuronal functions, such as synaptic plasticity in PGNs or drug addiction in DNs. Last, we found broad changes in A/B compartmentalization between all cell types (Extended Data Fig. 11a, b), with lowest Pearson’s correlations of compartment eigenvector values between brain cells and mES cells and highest correlations between neuronal replicates (Extended Data Fig. 11c). This result suggests that sensory genes are more likely to belong to heterochromatic B compartments and to more strongly contact other B compartments in brain cells
Sign-in/Register to view highlights & summary Sign-in/Register to access full text options 
Talk to us
Join us for a 30 min session where you can share your feedback and ask us any queries you have
Schedule a call
