3D Multiomics Adds Depth to Study of Human Brain Development

An international research team has provided an unprecedented look at how gene regulation evolves during human brain development, showing how the 3D structure of chromatin—DNA and proteins—plays a critical role. Using single-cell profiling and multimodal imaging techniques, the team headed by Chongyuan Luo, PhD, at the University of California, Los Angeles (UCLA), and Mercedes Paredes, MD, PhD, at UC San Francisco (UCSF), created the first map of DNA modification in the hippocampus (HPC) and prefrontal cortex (PFC)—two regions of the brain critical to learning, memory, and emotional regulation. These areas are also frequently involved in disorders such as autism and schizophrenia. The study offers new insights into how early brain development may shape lifelong mental health.

“Neuropsychiatric disorders, even those emerging in adulthood, often stem from genetic factors disrupting early brain development,” said Luo, a member of the Eli and Edythe Broad Center of Regenerative Medicine and Stem Cell Research at UCLA. “Our map offers a baseline to compare against genetic studies of diseased-affected brains and pinpoint when and where molecular changes occur.”

The researchers hope their data resource, which they’ve made publicly available through an online platform, will prove to be a valuable tool that scientists can use to connect genetic variants associated with such conditions to the genes, cells and developmental periods that are most sensitive to their effects.

Luo, Paredes and collaborating scientists at the Salk Institute, UC San Diego, and Seoul National University, report on their studies in Nature, in a paper titled “Temporally distinct 3D multi-omic dynamics in the developing human brain.” In their paper they concluded, “Our data provide multimodal resources for studying gene regulatory dynamics in brain development and demonstrate that single-cell three-dimensional multi-omics is a powerful approach for dissecting neuropsychiatric risk loci.”

The adult human brain contains hundreds of cell types that exhibit what the authors described as “an extraordinary diversity of molecular, morphological, anatomic and functional characteristics.” And while the human hippocampus and prefrontal cortex play critical roles in learning and cognition, they continued, “the dynamic molecular characteristics of their development remain enigmatic.”

To produce their map the research team used a cutting-edge sequencing approach Luo developed and scaled with support from the UCLA Broad Stem Cell Research Center Flow Cytometry Core, called single nucleus methyl-seq and chromatin conformation capture (snm3C-seq).

This technique enables researchers to simultaneously analyze two epigenetic mechanisms that control gene expression on a single-cell basis: chemical changes to DNA known as methylation, and chromatin conformation, the 3D structure of how chromosomes are tightly folded to fit into nuclei.

Figuring out how these two regulatory elements act on genes that affect development is a critical step to understanding how errors in this process lead to neuropsychiatric conditions. Yet, as the researchers pointed out, to date, “the dynamic trajectory of DNA methylation and chromatin conformation changes have not been characterized with single-cell resolution in prenatal human brain tissues and compared to those of postnatal development using infant and adult samples.”

For their newly reported study, the research team analyzed more than 53,000 brain cells from donors spanning mid-gestation to adulthood, revealing significant changes in gene regulation during critical developmental windows. In capturing such a broad spectrum of developmental phases, the researchers were able to assemble a remarkably comprehensive picture of the massive genetic rewiring that occurs during critical timepoints in human brain development.

“The vast majority of disease-causing variants we’ve identified are located between genes on the chromosome, so it’s challenging to know which genes they regulate,” said Luo, who is also an assistant professor of human genetics at the David Geffen School of Medicine at UCLA. “By studying how DNA is folded inside of individual cells, we can see where genetic variants connect with certain genes, which can help us pinpoint the cell types and developmental periods most vulnerable to these conditions.”

For example, autism spectrum disorder is commonly diagnosed in children aged two and over. However, if researchers can gain a better understanding of the genetic risk of autism and how it impacts development, they can potentially develop intervention strategies to help alleviate the symptoms of autism, like communication challenges, while the brain is developing.

One of the most dynamic periods comes around the midpoint of pregnancy. At this time, neural stem cells called radial glia, which have produced billions of neurons during the first and second trimesters, stop producing neurons and begin generating glial cells, which support and protect neurons. At the same time, the newly formed neurons mature, gaining the characteristics they need to fulfill specific functions and forming the synaptic connections that enable them to communicate.

This stage of development has been overlooked in previous studies, the researchers suggest, due to the limited availability of brain tissue from this period. They commented “Together, the findings of our imaging analysis of the mid-gestational HPC demonstrated spatially distinct chromatin conformation signatures that marked transitions from neural progenitors to mature neurons,” they wrote. “Our study underscores the dynamic shifts from progenitors to neuronal and glial populations in the second and third trimesters to the neonatal period, highlighting the importance of using primary brain specimens in studies of perinatal development.”

Paredes, an associate professor of neurology at UCSF, added, “Our study tackles the complex relationship between DNA organization and gene expression in developing human brain at ages typically not interrogated: the third trimester and infancy. The connections we’ve identified across different cell types through this work could untangle the current challenges in identifying meaningful genetic risk factors for neurodevelopmental and neuropsychiatric conditions.” The authors added, “The pervasive remodeling of the neuronal methylome and chromatin conformation during perinatal development suggests that the human brain is particularly vulnerable to genetic and environmental perturbations that affect these developmental stages…This work provides a data resource to understand the genetic and epigenetic mechanisms of brain diseases.”

The findings also have implications for improving stem cell-based models, such as brain organoids, which are used to study brain development and diseases. The new map offers a benchmark for scientists to ensure these models accurately replicate human brain development.

“Growing a healthy human brain is a tremendous feat,” said co-author Joseph Ecker, PhD, professor at the Salk Institute and Howard Hughes Medical Institute investigator. “Our study establishes an important database that captures key epigenetic changes that occur during brain development, in turn bringing us closer to understanding where and when failures arise in this development that can lead to neurodevelopmental disorders like autism.”