Katia GeorgopolousKatia Georgopoulos, PhD, an investigator in the Cutaneous Biology Research Center at Massachusetts General Hospital, professor of Dermatology at Harvard Medical School, and Jean and Terry de Gunzberg MGH Research Scholar 2014-2019, is the corresponding author of a new study in Cell, Lineage-Specific 3D Genome Organization is Assembled at Multiple Scales by IKAROS.

What Question Were You Investigating with this Study?

A cell nucleus is a busy place with cellular proteins twisting and pulling DNA to fold the genome into intricate 3D structures that allow critical parts to touch each other to turn genes on and off.

This choreography is essential for cell development with each cell type finding its own way to craft an organization that is optimal for its chores.

Establishing proper communication between genes and far-away control switches at the right time in the right cell is not a small feat. Very few proteins possess the combination of the right features to control such organization.

So how does a a protein called IKAROS help "weave" the genome into the correct structure required for B cell differentiation and generation of a life-saving repertoire of antibodies?

What Approach Did You Use?

Dr. Georgopoulos has pioneered previous studies showing that IKAROS is critical for immune cell development.

These studies showed that IKAROS-loss-of function mutations caused lymphoid malignancies in animal models and partially explained why IKAROS loss is associated with poor prognosis in children and young adults with B cell precursor leukemias. 

Previous research by Dr. Georgopoulos also demonstrated that IKAROS was required to turn on the genes that are necessary to make a B cell.

To better understand how this happens, her lab teamed up with Ferhat Ay, PhD, an expert in genomic architecture at La Jolla Institute for Immunology. He investigates how parts of the genome interact and his lab develops bioinformatics tools to create and analyze 3D maps of the genome.

Yeguang Hu, PhD, an instructor in the Georgopoulos lab, performed technically demanding experiments that identified which parts of the genome touch each other in normal B cell precursors as well as where IKAROS and many other proteins were bound to the DNA, and also mapped modifications of the proteins that identify the functional state of DNA in different regions.

Dr. Hu then repeated the same experiments on B cell precursors after removing IKAROS, and on skin cells that don’t express IKAROS and skin cell with IKAROS turned on.

Daniela Salgado Figueroa, a graduate student in Dr. Ay’s lab, and her colleagues took these terabytes of data, generated integrated maps of all the data, and identified the critical contacts between regions of the DNA that allow the correct genes to be turned on or off.

What Were the Results?

The researchers found that IKAROS binds to specific parts of the genome and controls the formation of very useful loops using these sites as anchors.

This looping brings far-away genes together with their control elements, leading to the activation and expression of the genes needed for proper B cell development and antibody production.

Just as important, this folding of the genome keeps other control elements away from genes that should not be expressed in a B cell.

A striking example of this function is observed at the immunoglobulin light chain gene. A B cell’s main function is to generate antibodies that allows the body to attack different pathogens.

To make antibodies that will recognize different pathogens, the B cells chooses one of hundreds of “variable” regions and connects it to the “constant region” of the antibody gene.

These many hundreds of choices are next to the constant region on the DNA, but their sheer number means that some are as far as 3 million bases away.

Hundreds of loops anchored by IKAROS pull these distant regions together in space to allow both near and far variable regions to be chosen.

This compaction of the DNA is so dramatic, it can be seen under the microscope, and the two ends of the regions are superimposed in normal B cell precursors but separated when IKAROS is removed.

In a follow-up experiment, the researchers introduced IKAROS into human skin cells. They tested whether IKAROS can organize the genome in a cell type that does not produce any IKAROS of its own.

And IKAROS prevailed! It, indeed, changed the 3D organization of a substantial part of the skin cell genome and led to formation of new loops, some of which resembled those from their B cell analysis.

Research into how chromatin is organized in the 3D space can help us understand how healthy cells develop—and how an improperly folded genome causes disease such as immunodeficiencies and cancer. Going forward, the researchers are interested in learning more about IKAROS disruption and disease development.

Additional authors of the study, "Lineage-specific 3D genome organization is assembled at multiple scales by IKAROS," included, Zhihong Zhang, Margaret Veselits, Sourya Bhattacharyya, Mariko Kashiwagi, Marcus R. Clark, and Bruce A. Morgan. This research was supported by the National Institutes of Health (grants R01HL140622, R01AR069132, R56AR082402, T32AI007512, R35GM128938, R21AR074748, and and R01AI150860).

Paper Cited:

Hu, Y., Salgado Figueroa, D., Zhang, Z., Veselits, M., Bhattacharyya, S., Kashiwagi, M., Clark, M. R., Morgan, B. A., Ay, F., & Georgopoulos, K. (2023). Lineage-specific 3D genome organization is assembled at multiple scales by IKAROS. Cell, 186(24), 5269–5289.e22. https://doi.org/10.1016/j.cell.2023.10.023

About the Massachusetts General Hospital

Massachusetts General Hospital, founded in 1811, is the original and largest teaching hospital of Harvard Medical School. The Mass General Research Institute conducts the largest hospital-based research program in the nation, with annual research operations of more than $1 billion and comprises more than 9,500 researchers working across more than 30 institutes, centers and departments.