3D Epigenome of Fetal Human Cerebral Cortex Astrocytes Unveils Links Between Neuropsychiatric Disorders and Human Evolution
Background
The human cerebral cortex rapidly expands during fetal development as neural stem cells proliferate and differentiate into multiple lineages. At the core of this process are ventricular radial glia (vRG) and outer radial glia (oRG). Particularly in primates, the significantly increased oRG is considered a key cell type responsible for the uniquely broad and gyrencephalic human cortex, although the cell-specific gene regulatory mechanisms remain poorly understood.
Previous single-cell studies have revealed which genes are expressed and which chromatin regions are open, but have struggled to precisely trace long-range regulatory contacts in 3D space. This limits the interpretation of disease-associated variants, which often regulate genes tens of kilobases away rather than the nearest gene.
Researchers isolated vRG, oRG, oligodendrocyte precursor cells (OPC), and microglia (MG) from the human cerebral cortex at 15–24 weeks of gestation and performed an integrated analysis of gene expression, chromatin accessibility, DNA methylation, and 3D chromatin interactions. The different developmental stages were chosen because the peak appearances of RG and OPC·MG differ. Nature paper
Key Findings
The team identified 69,141 candidate cis-regulatory elements (cCREs) in vRG, 72,450 in oRG, 65,295 in OPC, and 69,508 in MG with high accessibility and low methylation. Over 60% of these were located outside promoters. Using promoter-linked chromatin interaction analysis by sequencing (PLAC-seq) with H3K4me3, they captured 135,000–144,000 significant interactions per cell type at 2-kilobase resolution, detecting approximately four times more contacts than the previous 5-kilobase analysis. The average interaction distance ranged from 188,000 to 233,000 base pairs.
In mouse embryo reporter experiments, 14 out of 18 RG and intermediate progenitor regulatory sequences showed enhancer activity in the embryonic brain. Comparing vRG and oRG revealed 7,941 differentially accessible regions but only 756 differentially methylated regions, indicating that differences between the two cells were more pronounced in chromatin accessibility and 3D contacts. Transcription factor LHX2 was linked to the oRG regulatory network, while ASCL1 was associated with vRG. When LHX2 was inhibited using short hairpin RNA, oRG self-renewal decreased and oligodendrocyte-like differentiation increased.
Machine learning analysis also narrowed the scope of disease variants. Among 11,360 schizophrenia-associated variants identified by DeepGWAS, 929 were located in open chromatin, and 112 were classified as candidates for altering accessibility. The risk allele T of rs4449074 was predicted to reduce the activity of the vRG enhancer hs3134, and in mouse forebrain, it showed weaker signals than the non-risk allele C. Alzheimer’s disease risk was enriched only in MG regulatory elements, while autism spectrum disorder risk was prominent in vRG and oRG.
Implications and Outlook
Evolutionary analysis showed that 72 of the differentially accessible regions in oRG overlapped with human accelerated regions (HAR), compared to only 13 in vRG, indicating a 2.43-fold higher concentration in oRG. The team evaluated 565 open HARs and 3,447 human-chimpanzee sequence differences in oRG, identifying 76 candidates where human sequences increased accessibility and 73 where chimpanzee sequences remained more open.
HARsv2_1313 was connected to the promoter of ROCK2 through 3D contact. Inhibiting this region using CRISPR interference (CRISPRi) in induced pluripotent stem cell-derived neural progenitors reduced ROCK2 expression and increased Ki67-positive cells. Additionally, the human-specific sequence of HARsv2_1602 lost a binding site for the repressive transcription factor ZBT18, potentially enhancing EPHA4-related regulatory activity. This suggests that human cortical expansion and neuropsychiatric risk may share some non-coding regulatory circuits.
However, the analysis values represent averages of cell populations isolated by fluorescence-activated cell sorting. Rare progenitor subtypes and spatial heterogeneity were not captured, and the accessibility differences among rs4449074 carriers were not statistically significant. Results from mouse embryos and cultured neural progenitors should not be directly extrapolated to human fetal development or patient pathology. Spatial and single-cell epigenomic analyses, along with precise editing experiments in human brain organoids, are needed to confirm causal relationships.
Nature, Published online: 02 September 2026; doi:10.1038/s41586-026-10987-6Integrative 3D epigenomic profiling of four glial cell types from the mid-gestation human cortex reveals cell-type-specific regulatory elements and chromatin interactions that illuminate the roles of non-coding variants in neuropsychiatric disease and human-specific cortical evolution.
This map can serve as a selective tool to connect non-coding variants identified in genome-wide association studies to the actual operating cells and target genes. For example, if a schizophrenia variant reduces vRG enhancer activity and interacts with specific developmental genes, pharmaceutical companies could evaluate candidate compounds using human cortical organoids that replicate the regulatory axis. Alzheimer’s disease variants could be prioritized for MG, while autism and schizophrenia variants could be assigned to RG and OPC, reducing the cost of cell model selection and functional validation. However, the data alone are not sufficient to determine diagnostic risk scores or therapeutic targets, and reproducibility must be confirmed in more donors with diverse genetic backgrounds.