Cracking the chromatin code
The genetic material of a single human cell is about two meters long. For it to fit inside the cell’s nucleus, DNA and its associated proteins fold together into nanometer-sized clumps called chromatin. This process brings sections of genetic material that are located far apart on the DNA strand into direct contact with each other, which is how individual genes can be switched on and off. This chromatin “origami” is regulated and controlled by so-called chromatin factors – and their failure to work properly can have serious consequences.
“Mutations in certain chromatin factors are often associated with developmental and mental disorders,” explains Dr. Alexander Kukalev, a senior scientist in Professor Ana Pombo’s Epigenetic Regulation and Chromatin Architecture Lab at the Berlin Institute for Medical Systems Biology of the Max Delbrück Center (MDC-BIMSB). Mutated chromatin factors are found, for example, in patients with learning disabilities, schizophrenia, and autism. Mice whose brain cells do not produce some important chromatin factors are not able to learn nor to form long-term memory. “Our core hypothesis is that by comparing how chromatin changes in different models of learning disability, we may be able to zoom in on specific mechanisms or pathways towards developing novel therapeutic interventions,” says Pombo. To put this to the test, the team wants to examine genome function after the loss of three chromatin factors in brain cells. This work is being supported by almost €400,000 in grant funding from the German Research Foundation (DFG) between now and 2026.
The scientists are focusing their studies on the chromatin factors CTCF, ATRX, and SATB2. The effects on learning had been previously modeled in mice by the groups of Professor Bong-Kiun Kaang in South Korea, Dr Nathalie Berubé in Canada and Professor Georg Dechant in Austria. The three research groups genetically suppressed these factors in mice in specific neurons of the hippocampus that “control” memory formation and learning. They independently found that the genetically modified rodents failed to learn and memorize. “It is therefore clear that the three chromatin factors are important for neuronal function, and now we would like to understand how they work in memory and learning mechanisms” says Kukalev. To find this out, the team now wants to explore the consequences of the presence or absence of the chromatin factors in the same hippocampal neurons. “Our goal is to understand the role that CTCF, SATB2, and ATRX play in gene expression and chromatin regulation, and to identify mechanisms that link chromatin structure and the activity of specific neurons in learning and memory processes,” explains Kukalev.
High-tech mapping of a cell’s genetic makeup
To do so, they will be using state-of-the-art techniques – including genome architecture mapping (GAM). Pombo and her colleagues developed GAM themselves as a way to reconstruct three-dimensional maps of a genome from extremely thin sections of nuclei within intact tissues. They have built on this technique to create immunoGAM: “Labeling certain cell types with fluorescent dye enables us to analyze only specific cell types of interest within a complex tissue without disrupting the tissue structure,” explains Izabela-Cezara Harabula, who is also involved in developing the project.
They will also use 10x Genomics Multiome technology to detect gene expression and chromatin changes at the single-cell level. “With this two-pronged approach, we can create cell-type-specific maps of gene expression and regulatory chromatin regions,” says Harabula.
By combining GAM with 10x multiome data, the team wants to find out which regions on the DNA have altered structures in the three mutants, and identify genes and pathways affected in all mutants. The researchers will also examine the regulatory mechanisms that lead to altered gene expression in the genetically modified mice. “The affected genes and pathways could serve as targets for developing new diagnostic and therapeutic strategies for the neurologically impaired,” says Kukalev. Pombo adds that the studies could eventually also be carried out on samples from human patients. “We hope the findings will help to develop effective therapies for people with learning disabilities.”
Text: Janosch Deeg