RNA molecules are masters of their own destiny

At any given time in the human body, in about 30 trillion cells, “DNA” is read into molecules of messenger RNA, the intermediate stage between DNA and proteins, in the process of is called transcription.

Scientists have a good idea of ​​how transcription begins: proteins called RNA polymerases are recruited to specific regions of the DNA molecules and begin to spray down the strand, producing synthesis of mRNA molecules as they go. But part of this process is not understood: how does the cell know when to stop transcribing?

Now, there is new work from the laboratory of Whitehead Institute Member Richard Young, who is also a professor of biology at the Massachusetts Institute of Technology (MIT), and Arup K. Chakraborty, professor of chemical engineering, physics and chemistry at MIT, suggesting that RNA molecules themselves are responsible for regulating their formation through a feedback loop. Too many RNA molecules, and the cell begins to transcribe to create more. Then, at a certain threshold, too many RNA molecules cause transcription to be stopped.

The research, published in Cell on 16 December, representing a collaboration between biologists and physicists, and giving some insights into the potential roles of thousands of RNAs that are not translated into any proteins, added called non-coding RNAs, which are common in mammals and have been kept secret by scientists for decades.

Previous work in Young’s laboratory has focused on transcriptional condensates, tiny cell droplets that bring together the molecules needed to rewrite DNA into RNA. Laboratory scientists discovered the transcripts in 2018, noting that they were usually formed when transcription began and transmitted a few seconds or later when the process was complete.

The researchers questioned whether the force controlling the release of the transcription condensates could be related to the chemical properties of the RNA they produced – in particular, its negative cost. If this were the case, it would be the latest example of cell processes regulated through a feedback mechanism – an efficient cellular system to control biological functions such as red blood cell production and DNA repair.

As a first test, the researchers used an in vitro test to determine if the level of RNA had an effect on condensate formation. They found that, within the range of physiological levels observed in cells, low levels of RNA stimulated droplet formation and high levels of RNA depolarized.

With these results in mind, Young Lab postdocs and co-authors Ozgur Oksuz and Jon Henninger teamed up with physics and co-author Krishna Shrinivas, a graduate student in Arup Chakraborty’s lab, to study the physical forces who was playing.

Shrinivas suggested that the team build a computer model to study the physical and chemical interactions between actively transcribed RNA and condensates formed by transcription proteins. The goal of the model was not to replicate the existing results, but to create a platform to test different scenarios.

“The way most people study these types of problems is to take a mixture of molecules in a test tube, shake it and see what happens,” Shrinivas said. “That’s as far removed from what happens in a cell as he can imagine. Our thought was, ‘Can we try to study this problem in its biological context, which is like this complex and unfair process? “

Studying the problem from a physics perspective allowed the researchers to take a step back from traditional methods of biology. “As a biologist, it ‘s hard to come up with new ideas, new ways of understanding how things work from the available data,” said Henninger. “You can make screens, you can make new players, identify new proteins, new RNAs that may be involved in a process, but you are still limited by our classical understanding of how all of these things interact. But when you talk about physics, you are in this theoretical space that extends beyond what the data can currently give you. Physicists like to think about how something would behave, with certain parameters. “

When the model was ready, the researchers could ask him questions about possible conditions in cells – for example, what happens to concentrations when RNAs of different lengths are extracted at different levels as time comes? – and then proceeded with a test at the laboratory bench. “We ended up with a really good collection of model and test,” Henninger said. “For me, it’s as if the model helps to extract the simplest features of this type of system, and then you can do more predictive tests in cells to see if it respond to that model. “

Through a series of modeling and experiments at the bench bench, the researchers were able to validate their hypothesis that the effect of RNA on transcription is due to the very negative molecular cost of RNAs. In addition, initial low levels of RNA were expected to increase and subsequently higher levels of release of condensates formed by transcription proteins. Because the cost is borne by backbone phosphate RNAs, the cost-effective molecular weight of RNA is directly proportional to its length.

To test this finding in a living cell, the researchers invented the stem cells of mouse cells with glorious condensates, then treated them with a chemical to inhibit the elongation transcription rate. Consistent with the model prediction, the scarcity of condensate-releasing RNA molecules increased the size and life of condensates in the cell. On the other hand, when the researchers engineered cells to secrete additional RNAs, transcriptional concentrations spread at these sites. “These results highlight the importance of understanding how unbalanced feedback mechanisms regulate the actions of the biomolecular condensates present in cells,” Chakraborty said.

Demonstration of this feedback mechanism could help answer long-term secretions of the mammalian genome: the cause of uncoded RNAs, which make up a large proportion of genetic material. “While we know a lot about how proteins work, there are tens of thousands of non-coding RNA species, and we don’t know the functions of most of these molecules,” Young said. “The discovery that RNA molecules can regulate transcriptional condensates makes us wonder if many of the noncoding species are just working locally to modulate gene expression throughout the genome. a possible solution to this great mystery of what all these RNAs do. “

The researchers hope that understanding this new role for RNA in the cell could inform treatments for a wide range of diseases. “Some diseases are actually caused by more or less a single gene mutation,” said Oksuz, co-author. “We know now that if you change RNA levels, you have a predictive effect on condensates. So you could raise or lower the expression of a disease gene to restore the expression – and maybe bring back the phenotype – that you want, to treat an infection. “

Young said a deeper understanding of RNA behavior could inform pharmacology in general. In the last 10 years, several drugs have been developed that target RNA directly. “RNA is an important target,” Young said. “Understanding mechanically how RNA molecules regulate gene expression closes the gap between gene dysregulation in disease and new RNA-focused therapeutic approaches.”

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