A study published today (Jan. 8) in eLife, led by University of Wisconsin–Madison Professor of Biochemistry and Bacteriology Robert Landick and his research team, reveals for the first time the elemental mechanism behind transcriptional pausing, which underlies the control of gene expression in all living organisms.
It also provides new understanding of the enzyme RNA polymerase, an important drug target for treating conditions such as Clostridium difficile infections and tuberculosis. The findings could ultimately improve our understanding of how certain drugs work against the enzyme and aid in actively targeting it.
Gene expression is the process by which DNA is translated into all the proteins and other molecules living organisms need. Though it’s a process introductory biology students everywhere learn about, scientists are still far from fully understanding it.
The process occurs in two steps. Transcription is the first, where RNA polymerase reads the information on a strand of DNA, which is then copied into a new molecule of messenger RNA (mRNA). In the second stage, the mRNA moves on to be processed or translated into proteins.
To help control gene expression levels, transcriptional pausing by RNA polymerase can occur between the two stages, providing a kind of ‘roadblock’ where transcription may be terminated or modulated by the cell if need be.
“A sequence that causes pausing of RNA polymerases in all organisms, from bacteria to mammals, halts the enzyme in a paused state from which longer-lived pauses can arise,” explains Landick. “As the fundamental mechanism of this elemental pause is not well defined, we decided to explore this using a variety of biochemical and biophysical approaches.”
The team’s analyses first revealed that the elemental pause process involves several biological players, which together create a barrier to prevent escape from paused states. The process also causes a modest conformational shift that makes RNA polymerase ‘stumble’ when feeding DNA into its reaction center, temporarily stopping it from making RNA.
“We also found that transcriptional pausing makes RNA polymerase loosen its grip and backtrack on the DNA while paused,” says Landick. “Together, these results provide a framework to understand how the process is controlled by certain conditions and regulators within cells.”
He adds that these insights could aid future efforts to design synthetic genes, for example to direct the pausing behavior of RNA polymerase in a way that yields desired outputs from genes. It could also help our understanding of how certain drugs, known as RNA polymerase inhibitors, target the enzyme.
“For now, we would like to try and generate structures of paused transcription complexes obtained at a series of time intervals,” Landick concludes. “This would allow us to see exactly how parts of the enzyme move as it enters and leaves the paused state.”