Scientists determine the structure of the key factor in RNA quality control – ScienceDaily

In biology, it can be just as important as getting rid of things. Accumulation of cells, proteins, or other molecules that are no longer needed can cause problems, so living things have developed a variety of ways to clean your home.

A prime example is the RNA exosome. RNA molecules perform many functions in cells. Some of them go back to protein; others are the machinery for creating the proteins in a cell. RNA is a cellular machine that degrades defective, harmful, or no longer needed RNA molecules. Without this microscopic Marie Kondo to pruning what does not spark joy, our cells would become dysfunctional collectors, unable to function.

“RNA surveillance and degradation pathways are present in all walks of life,” explains Christopher Lima, president of the Sloan Kettering Institute’s Structural Biology Program. “From bacteria to humans, all living things have mechanisms in place to control and deliberately degrade the quality of RNA.”

For a long time, Dr. Lima says, these paths, like homework, were considered boring. But it turns out that these degradation pathways are highly regulated and control everything from embryonic development to the progression of the cell cycle.

In addition, defects in these pathways can lead to many types of disease, from cancer to neurodegeneration.

In a new article published on June 9, 2022 Cell, Dr. Lima and M. Rhyan Puno, a postdoctoral fellow at the Lima Laboratory, present findings that help explain how the RNA exosome locates RNA that needs to be degraded. With the help of cryogenic-electron microscopy (cryo-EM), scientists were able to decipher the structure of a set of proteins called the Nuclear Exosome Targeting (NEXT) Complex, which is a key part of degradation. machinery.

“We knew that NEXT targets and delivers RNA to the exosome, but we didn’t know what it looks like or how it works biochemically and structurally,” says Dr. Puno.

Now, with cryo-EM, scientists have obtained the first clear images of RNA-related NEXT. These images, along with biochemical and biological experiments, provide guidance on how to destroy RNA molecules in the exosome.

Approaching a structure

A few years ago, Dr. Puno began studying the structure of NEXT using an hour-long gold view of X-ray crystallography. In this method, the proteins are first converted to crystals, with all the proteins aligned in the same way. The X-rays are then passed through the crystals and the X-ray pattern played on a detector can be interpreted to determine the structure of the protein.

While Dr. Puno was able to crystallize the NEXT protein, the resulting X-ray diffraction images were not good enough to see the details of the structure.

“But then came the cryo-EM revolution,” he says. “It helped us see what this cryo-EM protein looks like and how it binds to its RNA substrates.”

Display of moving proteins

Cryo-EM works by capturing many different images of a frozen but non-crystallized sample of a protein, and then using computational methods to align it into a finite sharp image.

“It’s almost like taking a bunch of pictures of a bird on the flight,” says Dr. Lima. “There are all kinds of confusing movements and the bird’s wings can be blurred. But if we find parts of the wing in these different images, then we can align the images to reconstruct what the bird’s wings look like and how they are.”

From the cryo-EM images, the scientists were able to see that NEXT proteins form a very flexible dimer, meaning that two copies of NEXT proteins come together as functional units.

“It was really, really amazing,” says Dr. Puno, noting that the formation of dimers for these types of proteins has not been seen.

“From the biochemical experiments we’ve done, we know that dimerization is somehow important for degradation,” he continues. “But it’s still a mystery to us what role dimer plays in driving RNA into the exosome.”

To help solve the mystery, they hope to capture the NEXT complex interacting at different stages of the degradation process and then visualize these conformations via cryo-EM.

RNA degradation and disease

There are big bets involved. The extent of RNA degradation is indicative of the long list of defective or poorly controlled diseases. Perhaps the most well-known example is cystic fibrosis. In this case, the messenger RNA encoding a protein that travels ions in the cell membranes is degraded by RNA decay pathways. As a result, the protein is not present in the mucous membranes of the lungs, which accumulates mucus in it and severely interferes with respiration.

“It is a famous example of RNA quality control with poor results,” says Dr. Lima.

But defects in RNA degradation pathways also affect a variety of cancers. In fact, MSK’s genetic testing platform, MSK-IMPACT®, tests two genetic mutations that are found in genes related to the RNA exosome pathway, including a NEXT protein.

And it’s not just messenger RNA that needs proper quality control, Dr. Lima explained.

“The reality is that if you have faulty ways to control RNA quality, your ribosomes don’t work, your transfer RNAs don’t work, your spliceosomes don’t work.” The list goes on.

The breadth of functions that RNA performs explains why faulty RNA degradation pathways can have adverse effects on the cascade that causes disease.

To make sense of these effects, we need to understand not only the RNA exosome itself, but also the upstream proteins that help preserve RNA and help decide whether an RNA is defective or not.

“The dream is to start the RNA degradation reaction, put the sample in cryo-EM, and actually see all the possible assertions as it is doing its job,” says Dr. Lima. “As structural biologists, we want to be able to see the processes up and running and then reassemble them.”

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