For the first time, researchers describe how raw proteins actually block gene expression.
New research has identified and described the cellular process, which, despite what textbooks say, remains elusive for scientists – precisely how the genetic material was copied, once started, properly Is closed.
This discovery concerns the key process necessary for life: the transcription phase of gene expression, which enables cells to live and do their job.
During transcription, an enzyme is called RNA The polymerase is wrapped around its double helix DNA, Using a single strand to match nucleotides to mimic genetic material – resulting in a new synthesized strand of RNA that breaks down when transcription is complete. It enables the production of RNA proteins, which are essential for all life and do most of the work within cells.
With any consistent message, the RNA It is necessary to start and stop at the right place to understand. A bacterial protein called RHO was discovered ho0 years ago due to its ability to inhibit or terminate transcription. In each textbook, the RHO is used as a model terminator that, using its very strong motor force, binds to the RNA and pulls it through the RNA polymerase. But a closer look at these scientists showed that the RHO could not find the RNA needed to release it using the textbook mechanism.
“We started studying RHO, and realized that people might tell us that it might not work the way it works,” said Irina Artisimovich, co-lead author of the study at The Ohio State University and professor of microbiology.
This research, published online by the journal Science Today, November 26, 2020, instead of joining a specific part of the RNA near the end of the transcription and helping to unravel it from the DNA, RNA actually “moves” on the RNA polymerase for the duration of the transcription. RNA cooperates with other proteins to eventually coax the enzyme into a series of structural changes that end up enabling the release of the dormant state RNA.
The team used a sophisticated microscope to reveal how a complete transcription complex made up of RNA polymerase and the two supporting proteins that travel with it during transcription work.
“This is the first structure of the termination complex in any system, and it was considered impossible to obtain because it breaks down so quickly,” Artisimovich said.
“It answers a basic question – transcription is fundamental to life, but nothing will work if it is not controlled. The RNA polymerase itself should be completely neutral. It should be able to make any RNA, including those that have damaged or damaged the cell. When traveling with RNA polymerase, the RHO can tell if the synthesized RNA is fit to form – and if not, the RHO releases it. “
Artisimovich has made many important discoveries about how RNA polymerase successfully completes transcription. While working on the heredity project no undergraduate student in his lab could cope with years of understanding about Ronnie’s role in ending it until he identified a surprising change in RH.
Rowe is known to mute the expression of viral genes in bacteria, essentially keeping them inactive until they need to be infected. But these genes do not contain any RNA sequences that RHOs are known to bind primarily to. Because of this, Artisimovich said, he never realized that RHOs only look for specific RNA sequences, without knowing whether they are connected to RNA polymerase only.
In fact, the scientific understanding of the RHO mechanism was established with the help of simple biochemical experiments that repeatedly left RNA polymerase – in short, defining how the process ends without factoring in the process.
In this work, the researchers used cryo-electron microscopy to obtain images of RNA polymerase acting on a DNA sample in their model system Escherichia coli. This high-resolution visualization, coupled with high-end calculations, made accurate modeling of transcription terminations possible.
“RNA polymerase advances, matching thousands of nucleotides in bacteria. The complex is extremely stable because it is – if the RNA. If released, it is lost, “said Artisimovich.” However, RH can collapse in seconds, if not seconds, in minutes. You can see it, but you can’t get a stable complex to analyze. “
Using a clever method of trapping them just before separating the complexes, the scientists were able to visualize seven complexes, representing a sequential step in the finished path, starting with the RHO’s engagement with the RNA polymerase and ending with the fully inactive RNA polymerase. The team created models based on what they saw, and then made sure that these models were appropriate using genetic and biochemical methods.
Although the study was conducted in bacteria, Artisimovich said the termination process is likely to occur in other forms of life.
“It seems to be normal,” he said. “In general, cells use the same functional mechanisms as the common ancestor. He learned all the tricks until these tricks were useful. ”
Ref: 26 November 2020, Science.
Artisimovich, working with an international research team of collaborators, now co-leads the study with Marcus Wahl, a former Ohio State graduate student at Free University Berlin.
This work was supported by a grant from the German Research Foundation; German of the Ministry of Education and Research; Indian Medical Research Council; Department of Biotechnology, Government of India; National health organizations; And the Sigrid Jusalius Foundation.
