Before a cell can divide into two, it must first copy all of its chromosomes so each recent cell inherits a whole set of genetic material. For years, researchers believed that as this process unfolded, the complex three-dimensional shape of the genome was temporarily lost.
After division, scientists believed, DNA would step by step rebuild its complex, folded structure, which is vital to regulating which genes are energetic in a given cell.
Now, recent research from MIT shows that this long-accepted model isn't quite right. Using a strong, high-resolution genome-mapping method, the team found that small 3D loops—contacts between regulatory DNA elements and genes—are maintained even during cell division, a stage generally known as mitosis.
“This study really helps clarify how we should think about mitosis. In the past, mitosis was thought of as a blank slate, with no transcription and no structure related to gene activity. And now we know that's not quite the case,” says Anders Sager Hansen, an associate professor of biological engineering at MIT. “What we see is that there is always structure. It never goes away.”
DNA loops that help cells “remember”.
The researchers also discovered that these DNA loops actually strengthen the chromosomes as they shorten in preparation for division. This stiffness brings distant regulatory elements closer together, encouraging them to remain. According to the team, this might allow cells to “remember” which genetic interactions were present before division and re-establish them afterwards.
“These findings help link the structure of the genome to its function in how genes are turned on and off, which has been a great challenge in the field for decades,” says the study's lead writer, Virat Goyal PhD '25.
Henson and Edward Bannigan, research scientists at MIT's Institute for Medical Engineering and Science, are senior authors of the study, which is published Co-authors include Professors Leonard Mirny of MIT and Gerd Buble of the University of Pennsylvania's Perelman School of Medicine.
Mapping the hidden architecture of DNA
Over the past twenty years, scientists have discovered that DNA within the cell nucleus organizes itself into 3D loops. Many of those loops allow genes to interact with distant regulatory regions—sometimes thousands and thousands of base pairs apart—while others form to tightly pack chromosomes during mitosis.
Much of this mapping has relied on a tool called HiC, a method developed in collaboration with MIT researchers and Job Dekker on the University of Massachusetts Chen Medical School. HI-C works by cutting DNA into small pieces and joining those which might be close to one another within the cell nucleus before sequencing them to discover which regions interact.
Although efficient, HiC lacks the resolution needed to detect fine-scale interactions between genes and regulatory sequences generally known as enhancers. Enhancers are short DNA segments that activate genes by binding to promoters, regions where transcription begins.
A breakthrough tool: region capture micro-C
In 2023, Hansen and his colleagues developed a next-generation technique that may map genome structure with 1,000 times more precision. This method, called region-capture microC (RC-MC), uses a unique enzyme to chop DNA into pieces of uniform size and is concentrated on a small a part of the genome. This allows researchers to create highly detailed 3D maps of targeted DNA regions.
Using RC-MC, the team identified a brand new structural feature called “microcompartments”. These are small, densely connected loops that connect nearby enhancers and promoters.
Previous work had shown that the microcompartments were formed by different mechanisms than the larger 3D genome structure, however the team didn’t yet understand how. To explore this, they decided to look at what happens to those structures as cells undergo mitosis. During this phase, chromosomes compact dramatically to be sure that they may be duplicated and distributed equally between daughter cells. As this happens, large genome domains generally known as A/B compartments and topologically associating domains (TADs) normally disappear.
Unexpected stability during cell division
The researchers expected that the microcompartments would also disappear. To test this, they monitored cells throughout the division cycle to see how these loops behaved before and after mitosis.
“During mitosis, it's thought that almost all gene transcription is turned off. And before our paper, it was also thought that all of the 3D structure related to gene regulation is lost and replaced by compression. This completely resets each cell cycle,” Hanson says.
Surprisingly, the loops didn't disappear. In fact, they became much more pronounced as they divided into cells.
“We went into this study thinking, well, one thing we know for sure is that there is no regulatory structure in mitosis, and then we accidentally found structure in mitosis,” Hanson says.
Using their technique, the researchers also confirmed that enormous structures similar to the A/B compartment and TDS disappear during mitosis, as previously observed.
“This study takes advantage of the unprecedented genomic resolution of the RC-MC assay to reveal new and surprising aspects of mitotic chromatin organization, which we have overlooked in the past using traditional 3C-based assays. The authors revealed that regulatory elements—are maintained or even transiently strengthened,” at Weill Cornell Medicine. says Effie Apostolo, an associate professor of molecular biology in Medicine, who was not involved within the study.
Explaining mysterious bursts of gene activity
This finding may explain the long-term burst in gene transcription that happens near the tip of mitosis. Since the Sixties, scientists believed that replication stopped completely during cell division. However, studies in 2016 and 2017 revealed a transient increase in gene activity before it turned off again.
In their recent study, the MIT team found that in mitosis, microcompartments usually tend to be found near genes that increase during cell division. They also discovered that these loops appear because of this of genome compression that happens during mitosis. This compression brings enhancers and promoters closer together, allowing them to stay together to form microcompartments.
Once formed, loops forming microcompartments can partially activate gene transcription by accident, which is then turned off by the cell. When the cell stops dividing, entering a state generally known as G1, a lot of these cells weaken or disappear.
“This transcriptional spiking in mitosis appears to be an unwanted accident that arises from creating a uniquely favorable environment for the mycomas compartment to form during mitosis,” Hanson says. “Then, when it enters G1 the cell quickly eliminates and filters out many of these loops.”
Because chromosome compression may also be affected by cell size and shape, researchers at the moment are exploring how variations in these properties affect genome structure and, in turn, gene regulation.
“We're thinking about some of the natural biological settings where cells change shape and size, and whether we might be able to explain some of the 3D genome changes that have previously lacked explanation,” Hanson says. “Another important question is how does the cell then choose what microcompartments to keep and when you enter G1, what are the microcompartments to ensure fidelity of gene expression?”
This research was supported partly by the National Institutes of Health, a National Science Foundation CAREER Award, the Broad Institute's Gene Regulation Observatory, a Pew Steward Scholar Award for Cancer Research, the Mathers Foundation, the MIT Westway Fund, the Bridge Project of the Koch Institute and the Dana-Ferber/Harvard Cancer Center, and Koch Institute support. (Core) was funded by a grant.