Broad Institute researchers have developed a technology that provides new insight into how disruptions in the nucleus of the cell can impact health and disease.

The approach, called expansion in situ genome sequencing, allows scientists to sequence DNA and map its location relative to proteins within cell nuclei. The method uses a gel to expand cells while keeping them intact, enabling both sequencing and high-resolution imaging within the same cells. The research team applied their technique to cells from patients with progeria, a disorder marked by accelerated aging. The scientists found that mutated proteins in the nucleus may suppress the expression of certain genes, which may play a role in the disease and the aging process. 

The researchers suggest that these types of changes in the cell nucleus could be at the root of other diseases, and can now be studied in greater detail using their new method.

The findings appear today in Science and come from the labs of institute member Jason Buenrostro and core institute member Fei Chen at the Broad. Buenrostro and Chen are also associate professors at Harvard University, and Buenrostro is a co-leader of Broad’s Gene Regulation Observatory.

“With this work we can now sequence the genomes of cells directly within cells,” said Buenrostro. “This has inspired us to dream up new biological questions connecting genome structure to function.”

Postdoctoral researchers Ajay Labade and Zachary Chiang and graduate student Caroline Comenho, all in Buenrostro’s lab, are co-first authors on the work.

“When we think of cell biology, we usually think of imaging and sequencing as two very different modalities,” said Chiang. “This technology is a way of connecting the types of images clinicians use to diagnose disease with high-resolution molecular readouts — and we hope it will allow scientists to ask new types of questions about disease.”

An expanding view of cells

Buenrostro and Chen have been collaborating since 2015, when they were both Schmidt Fellows at the Broad. By 2020, in collaboration with Ed Boyden’s group at MIT, the pair had developed in situ sequencing, which sequences DNA within intact cells, providing a direct view of the genome’s structure in 3D. But the approach was best suited for analyzing cells with large nuclei, such as embryos. 

To push its resolution to the nanoscale, Labade, Chiang, and later Comenho spent years integrating the technique with a method developed by Chen in 2015 called expansion microscopy, which expands tissues so that typical light microscopes can capture high-resolution images of biological structures.

“This took a lot of refining, from both experiments to computational analysis, and we’ve shared some tips and tricks with the field,” said Labade. “We hope this work will be a resource for people who want to apply expansion genomics to their own experiments.”

Gene repression

In the new study, the team first examined skin cells from patients with progeria, a disorder caused by mutations in lamin proteins. These proteins form a mesh-like structure that supports the nucleus in healthy cells but is distorted in progeria, forming invaginations inside nuclei that are difficult to observe with standard methods. 

Using their approach, the team observed not only these invaginations but also the DNA sequences near them — and found that genes critical to cell function were repressed in those areas, and had fewer RNA synthesis enzymes nearby. The team also noticed invaginations in cells from a 92-year old individual without the condition, suggesting that these changes in lamin structure disrupt gene expression in both progeria and the aging process.

Gene repression in the periphery of the nucleus has been well studied, but is less well understood near the center of the nucleus, where the invaginations form. So the team thinks that lamin and the spatial organization of the genome deep in the nucleus could be underappreciated factors controlling gene expression throughout a person’s lifetime. They add that their approach could reveal other connections between nuclear abnormalities and disease.

“When you’re studying biology, you learn that a nucleus is a circle, but when you look at cells under the microscope, they all have unique sizes and shapes,” Comenho said. “I think we’re a part of a new era, where we can start asking how these structures translate to cell function.”

With that aim, the team plans to expand the technique to include other capabilities such as sequencing RNA and chromatin accessibility and measuring cell behavior. Incorporating different microscopes would also allow researchers to examine tissue sections, making the approach more practical as a possible diagnostic tool.