A tale of two measurements: protein-DNA interactions and gene expression in single cells. Protein-DNA interactions and gene expression in single cells

Abstract

Our bodies are composed of trillions of cells which are the building blocks that cooperate with each other. A neuron has a characteristic tree-like appearance and is specialized in transmitting electrical pulses to signal. Fat cells on the other hand, are round and their primary function is fat storage. These two cell types, as well as the rest of the 200 different cell types in the human body, contain the same genetic information, yet are functionally and morphologically different. How does the same DNA code result in different outcomes? It is all about exposure.

Not all DNA code is available for the cell to read and act upon at all times. Some parts of the DNA can be tightly folded, while others are more lose and are therefore easier for the cell to read. When the cell “reads” the genetic code, it copies this region, creating so-called messenger RNA (or mRNA) transcripts which are used as blueprints to produce proteins. A neuron needs different proteins than an adipocyte, which means that the accessible DNA parts and the resulting mRNA transcripts are different between the two cell types. Therefore, by measuring the mRNA transcripts of a cell, a PhD student can understand what cell type she is looking at.

The DNA is stored in a spherical compartment at the center of the cell called the nucleus. Near the center of the nucleus, the DNA is more flexible and open, while at its periphery, the DNA is more densely packed. Whether the DNA is open or packed is important for cell function and morphology. Firstly, because the regions that are located at the periphery and the ones residing at the nuclear interior can differ between cell types. Secondly, because the DNA code that is needed by the cell is often located in the center of the nucleus where it is “read”. In contrast, code that is not useful for the cell’s tasks is “stored” at the periphery of the nucleus, and mostly goes unused.

This thesis focuses on relating DNA folding and mRNA production. In general, DNA at the periphery of the nucleus is not read by the cell and produces almost no mRNA. However, until now, this was not confirmed because of the lack of molecular tools. To link the DNA folding with the mRNA production, one has to measure both things in the same cell. We developed the method scDam&T-seq that measures both DNA folding and mRNA in the same cell. We confirmed that most DNA regions are not read by the cell when they are at the periphery of the nucleus. We also found that some regions are more prone to be “shut off” at the nuclear periphery than other regions. We also applied scDam&T-seq to the brain of developing mice. We found that the cells which produce the neurons in the brain show differences in DNA folding even though their mRNA seems similar. In conclusion, this thesis presents mainly technical solutions to the field of DNA architecture.