With nearly six feet of DNA packed inside each human cell, the question of how chromosomes are organized within the nucleus has intrigued scientists for decades. Increasingly, it is understood that chromosomes are not dispersed randomly in the nucleus – that in fact, location within the nucleus has functional consequences.
Mapping the Spatial Organization of Chromosomes
“Specific stretches of DNA are anchored to distinct physical structures in the nucleus, and this aspect of genome architecture helps regulate gene function,” said UCSF neurosurgeon and neuroscientist Daniel Lim, MD, PhD (Professor of Neurological Surgery). “However, we really had no idea how this is organized in the human brain.”
To address this question, a team of researchers led by Lim developed a novel technique for mapping the genome to specific nuclear compartments. The new method, called GO-CaRT (Genome Organization with CUT and RUN Technology), uses antibodies to target specific nuclear compartments, such that DNA fragments associated with those structures can be isolated and analyzed using high-throughput sequencing methods.
Published in Nature Neuroscience, Lim and colleagues use GO-CaRT technology to generate the first maps of how the genome is spatially organized in the nucleus of brain cells. Specifically, they mapped chromosomal regions that interact with two nuclear compartments: the lamina, which is considered to be transcriptionally repressive, and nuclear speckles, which are thought to be involved in RNA processing.
By identifying these lamina-associated domains (LADs) and speckle-associated domains (SPADs), Lim and colleagues have mapped the spatial genome organization inside brain cells, which has implications for our understanding of both brain development and disease.
The localization of chromosomal regions to certain nuclear compartments could well underlie the differences in gene expression that impact a cell’s identity and how it functions. In fact, the Lim Lab found that a majority of LADs – the chromosomal regions located near the lamina – are conserved across the brains of humans, macaques, and mice.
“We have been able to give physical addresses to the entire genome,” said co-first author and postdoctoral fellow Sajad Ahanger, PhD. “Now we can map these addresses in the context of different cell types, or even different diseases, for example to see whether certain genes have lost their addresses, and whether that contributes to how the disease manifests.”
Indeed, Ahanger and Lim discover that in the developing brain, SPADs – the chromosomal regions located in the nuclear speckles – are highly enriched for genetic risk variants of schizophrenia. Although it remains unclear how this contributes to schizophrenia pathology, this finding lays the groundwork for investigation of the relationship between genome architecture and genetic causes of a wide range of diseases.
A New Tool for In Vivo Investigation
Ahanger and Lim’s innovative GO-CaRT tool also offers the unprecedented ability to study genome architecture more easily than before. “Prior studies were in mammalian cultured cells and required millions and millions of cells,” said Ahanger. “Here we have done it using 75,000 cells that were directly isolated from the human brain.”
Often, cultured cells do not fully recapitulate the conditions found in animal models and tissues, so the ability to more accurately study what occurs in vivo, in living organisms, is critical. “Almost everything that we’ve previously known about genome organization was derived from cell culture studies,” said Lim. “This is the only study, to date, focused on in vivo tissues.”
Lim and colleagues found significant differences in the chromosomal regions reported to be located near the lamina in cultured neural progenitor cells, compared to the LADs they identified in the developing brain – underscoring the need for a tool that is compatible with in vivo investigation.
“We are really starting to explore how the genome is organized relative to these structures during development or during disease,” said Ahanger. “I think this opens up a lot of exciting opportunities not only in the brain, but different organs and systems.”
Pioneering New Concepts in Genomics Research
While the field of spatial chromosomal organization is relatively new, Lim is no stranger to challenging convention when it comes to our understanding of genome biology. Lim is well known for his work studying the non-coding regions of DNA, which had long been considered “junk DNA”. It is now increasingly evident, in part due to the Lim Lab, that long non-coding RNAs play a critical role in both development and disease.
Ahanger and Lim’s current publication follows in a similar tradition of exploring new, and sometimes unexpected, concepts. “The bigger question that inspired all this is: How is the genome organized inside the nucleus and what is the importance of that?” said Lim.
The idea that genome architecture impacts gene expression – and ultimately, identity and function of the cell – opens new opportunities for future investigation. “As people start to understand genetic variation and disease,” said Lim, “we’re able to start mapping it to a 3D spatial dataset that we can now easily generate.”
Ahanger SH, Delgado, RN, Gil E, Cole MA, Zhao J, Hong SJ, Kriegstein AR, Nowakowski TJ, Pollen AA, Lim DA. Distinct nuclear compartment-associated genome architecture in the developing mammalian brain. Nat Neurosci (2021). https://doi.org/10.1038/s41593-021-00879-5
