To further our quantitative understanding of cellular decision-making, Dr. Gregory Reeves and his team in the chemical engineering department worked to interpret how a transcription factor dictates the change in gene expression in cells.
The team’s work, recently published in Advances in Sciencefocuses on a protein called Dorsal, which is a version of nuclear factor-κB (NF-κB)—a critical transcription factor that controls cellular processes and decision-making while regulating cellular immunity and growth.
NF-κB is involved in various medical cellular behaviors such as inflammation, innate immunity, and wound healing. This level of understanding could lead to the ability to control these cellular processes ourselves, because errors in NF-κB activity can lead to disease states such as cancer.”
Dr. Gregory Reeves
NF-κB has several states within the cell nucleus. It has the ability to bind to DNA, accumulate, and be active or inactive. Reeves’ team has shown that gene regulation occurs at this level.
“We can distinguish between molecules that move slowly versus those that move fast, as well as those that don’t move at all,” Reeves said. “We can do this using a fluctuation spectroscopy method that shows us how much Dorsal is moving.”
The goal is to create a map that correlates how much backbone is in the nucleus and how much of it is bound to DNA. Understanding this map of how Dorsal binds to DNA leads to a predictive level of understanding and may reveal how to manipulate this pathway for therapeutic purposes.
Using special imaging techniques to identify the different states of Dorsal in the cell, the team was able to obtain mathematical models that reflect an accurate picture of how much Dorsal binds to DNA, as well as how much Dorsal is clustered together.
In previous studies, Reeves’ team only took snapshots with the imaging technique. They decided to look at the cells for a longer time.
The work spans multiple length and time scales, allowing them to gain a core-wide picture of the mechanism that connects Dorsal to DNA.
“When we looked at how much Dorsal was freely circulating, it seemed to be independent of how much was in the core,” Reeves said. “This map would reveal the free amount of Dorsal in the nucleus. Once we created it, others would be free to use it so they could advance their understanding of gene regulation.”
The relationship between how much NF-κB is in the nucleus versus how much of it is doing its job on DNA becomes clear when applying these methods. This allowed Reeves and the team to evaluate different parts of the embryos.
The team found that the amount of NF-κB that is free to move is constant in different parts of the embryo. The amount bound to DNA, however, is not constant. This means that the relationship between the two is not linear. “With this knowledge of how Dorsal interacts with DNA, we better understand how much we would need to activate the NF-κB pathway if we needed to intervene for therapeutic purposes,” Reeves said.
