A cell nucleus produces significantly more non-functional RNA than functional RNA. A Danish research group is delving deep into the engine room of evolution to try and find the reason why.
Your cells are full of RNA with no apparent function. In fact, researchers can only account for the function of 2–3 % of the RNA produced by the genome. The function of the remaining RNA, if any, is unknown.
Even though the apparently useless RNA has no immediate function, it can still affect our health. An ingenious system of different protein complexes protects and degrades both the functional RNA and the apparently useless RNA. If mutations arise in some of these protein complexes, this can lead to the development of cancer, disorders of the nervous system or other diseases.
This is why researchers are extremely interested in discovering how this non-coding RNA that does not translate into proteins affects the cells and, ultimately, the well-being of the organism and how the body’s cells regulate what RNA to keep and what to destroy.
“Researching these cellular processes means researching the fundamental processes of how our genome works. It is the lifeblood of evolution and biology, and it is easy to imagine that if these processes do not function properly, the cells will be flooded with non-coding RNA that can cause all kinds of problems,” says Torben Heick Jensen, Professor, Department of Molecular Biology and Genetics, Aarhus University, who leads an extensive new research project in this field.
The Novo Nordisk Foundation recently awarded Torben Heick Jensen a major Challenge Programme grant. For the next 6 years, he and colleagues from the University of Southern Denmark in Odense and the Max Planck Institute of Biochemistry in Martinsried, Germany will be busy performing experiments and delving deep into the engine room of biology.
Only a few percent of the function of the body’s RNA is accounted for
Torben Heick Jensen and his research group will basically be trying to determine how cells decide whether RNA should be kept or destroyed.
RNA enables DNA to function. Each of the genome’s genes is an architectural blueprint of a protein, and the body’s cells create pieces of RNA from the gene – messenger RNA (mRNA) – that can be translated into proteins.
Other pieces of RNA, such as transfer RNA (tRNA) and ribosomal RNA (rRNA), have structural roles and support the translation process even though they are not translated into protein.
And then there is the remaining RNA, the function of which still baffles researchers.
“Many types of non-coding RNA have various functions. However, this non-coding RNA is still important. If we add up all the RNA of which we know the functions, we can only account for a mere 2–3% of our genome,” explains Torben Heick Jensen.
Errors in regulating RNA can lead to cancer
The body’s cells are constantly performing a delicate balancing act to determine which RNA can be used and which cannot, and if the body’s cells do not maintain good RNA hygiene, then things can go wrong very quickly.
For this purpose, the cells are equipped with various protein complexes that can preserve or degrade pieces of RNA in the cell nucleus. If these protein complexes are defective, such as from a mutation, the cell almost immediately loses its ability to function properly.
The reason is that RNA – both the functional and the non-coding RNA – are continually produced, and if they are not sorted, the apparently useless RNA binds to proteins that it should not bind to, and the most basic cellular functions cease.
If this happens, a cascade effect begins that will ultimately affect people’s health. This is why mutations in the proteins that ensure good RNA hygiene are also linked to various diseases, such as cancer and disorders of the nervous system.
“RNA biology plays a big role in neurons. This is where we often see diseases in connection with mutations in the RNA degradation complexes, but we cannot say for certain that there is 1:1 relationship between mutations and specific diseases,” says Torben Heick Jensen.
Nevertheless, research into the cellular RNA degradation complexes opens up a variety of pharmaceutical opportunities.
Torben Heick Jensen envisages that using the gene editing technology CRISPR could, for example, correct errors in the complexes that degrade RNA, thereby preventing people from developing disorders of the nervous system and cancer.
“We are not researching this ourselves, but these are the prospects, and other people are investigating the potential of this field,” he says.
A pioneer in discovering new RNA
Torben Heick Jensen and his team are researching the protein complexes that degrade the RNA with no apparent function.
As of 10–15 years ago, researchers believed that most of the genome was not transcribed into anything useful, and certainly not to non-coding RNA, but Torben Heick Jensen’s research and that of other researchers show that this happens.
In 2008, the research team at Aarhus University published an article in Science outlining an experiment in which they shut down part of the RNA degradation machinery. The cell produced enormous amounts of RNA, which was previously unknown, since it had been degraded as rapidly as it was being produced.
“There was an explosion of new RNA that had never been seen before. We were co-pioneers in discovering that cells contain much more RNA that we had previously imagined. The main challenge in this field now is to determine exactly why the cells transcribe the genome into RNA that has no apparent function,” says Torben Heick Jensen.
RNA can make evolutionary sense
So why does evolution enable a cell to use its valuable resources to create RNA that has no function and is immediately degraded? This seems like such a waste.
But it actually makes sense to Torben Heick Jensen from an evolutionary viewpoint.
According to Torben Heick Jensen, the constant production of RNA brings life to the genome. This makes the genome become a dynamic toolbox that contains many potential new tools that could have a function in the future or tools that had a function in the past.
Some pieces of RNA could one day be used to encode a protein that benefits the cell or the organism in some way. Unless the RNA is expressed, the cell can never test these potential possibilities. In this case, it would only have access to the genome, which is like having a drawing of a hammer rather than an actual hammer. Determining whether or not a hammer is useful is difficult unless you test it in the real world.
“Having a dynamic genome with which to test things makes perfect sense in evolutionary terms. It does, however, require you to have the necessary machinery to maintain a certain level of hygiene,” says Torben Heick Jensen.
Protein complex degrades RNA
The specific degradation complex that Torben Heick Jensen previously deactivated in his research is called the RNA exosome.
The RNA exosome destroys RNA by indiscriminately degrading the RNA building blocks. Most RNA exosomes are present inside the cell nucleus, where the genome is translated into RNA, so if the RNA is to become the template for creating a protein, it must exit the cell nucleus as quickly as possible before an RNA exosome gets hold of it and degrades it.
The cell nucleus is a harsh environment for the RNA, but the cytoplasm outside the nucleus is a safe haven.
To assist the functional pieces of RNA in escaping the hellish environment of the cell nucleus and to prevent the non-coding RNA from exiting, evolution has equipped the cells with a series of protein complexes with various functions.
In addition to RNA exosomes, other protein complexes bind to the exosomes and help them to identify the correct RNA to break apart: adaptor complexes.
Cells also have protein complexes that bind to specific RNAs and protect them from exosomes while guiding the RNA out of the cell nucleus. These are the pieces of RNA that will be translated into functional proteins later.
The interaction between the protein complexes therefore determines which pieces of RNA escape from the cell nucleus and which ones are degraded as quickly as they are produced.
“The complexity of the whole system is enhanced by the fact that some RNA has a function in some cells and not in others. Each cell must therefore be equipped with specific protein complexes that ensure that the right pieces of RNA are expressed as proteins in the cytoplasm of a specific type of cell whereas others are not,” explains Torben Heick Jensen.
Discovering two protein complexes that decide the RNA’s fate
In relation to the present project, Torben Heick Jensen has had two major highlights in his research career. In 2011 and 2016, respectively, his research group identified two adaptor protein complexes: the proteins that help exosomes recognize the pieces of RNA to be degraded.
These adaptor complexes sort through the RNA for the exosomes and hand over the pieces of RNA to be degraded.
Torben Heick Jensen has just been awarded a grant to perform further research into identifying and mapping adaptor complexes.
In addition to mapping the precise structure of the complexes that have already been identified, Torben Heick Jensen hopes that his research team will also discover new adaptor complexes.
“There may be 5–10 adaptors in all. This depends somewhat on how broadly one looks. There may be some in human cell lines, and others may be in stem cells. This is because stem cells have a slightly different biology and need a different RNA expression to remain pluripotent and to continue to have the ability to differentiate into different types of cells. Finding one or two new adaptor complexes during the 6-year project would be a major achievement,” says Torben Heick Jensen.
Mapping adaptor complexes in minute detail
If the researchers discover new adaptor complexes, they will work on mapping their structures. They will start off by doing this with the two adaptor complexes they have already discovered.
When the researchers map these structures, they first find the genes that create the proteins in the protein complexes. Then they express the genes in a bacteria, yeast or mammalian cell, so they can produce enough proteins to do further studies.
When Torben Heick Jensen and his colleagues have successfully produced large amounts of adaptor proteins, they must try to get them in crystal form so they can X-ray them.
This technique is known as X-ray crystallography. How the proteins scatter the X-rays reflects the structure of the specific protein.
Parallel to this, the researchers will also study the proteins by taking hundreds of images with an electron microscope.
This technique is called cryoelectron microscopy, in which researchers take the many indistinct images of a protein and then consolidate the images into one sharp image of the protein using advanced imaging software.
Using the protein to discover more adaptor complexes
The research project is already underway, and the researchers from Aarhus University, the University of Southern Denmark and the Max Planck Institute of Biochemistry have mapped some very interesting structural aspects of the two adaptor complex structures that they can potentially use to find other ones.
Both protein complexes have a part that is important to the functioning of the exosome. This is called MTR4, and Torben Heick Jensen envisages that, if two adaptor complexes have this and therefore can optimally interact with the exosomes, then other potential adaptor protein complexes may also have them.
“If this is the case, then we can use MTR4 to find other adaptor complexes. This is the approach we are taking,” says Torben Heick Jensen.
RNA structure determines whether it is destroyed
Mapping the adaptor complexes that have already been discovered and possible discoveries of new ones will give researchers a much clearer idea of what happens inside cell nuclei when RNA is being created, degraded or protected.
Torben Heick Jensen envisages a very dynamic fate for all the pieces of RNA.
When RNA is being produced, the adaptor complexes recognize it within milliseconds, but the interaction with an adaptor complex is not static.
Adaptor complexes continually bind and release the RNA, maintaining an equilibrium between the exosomes that want to destroy the RNA and the proteins that want to help the RNA safely exit the cell nucleus.
“The RNAs that exit the cell nucleus generally contain fewer nucleotide sequences, known as introns, than the pieces of RNA that are not exported from the cell nucleus. This points to a mechanism that cells use to differentiate between the functional pieces of RNA and the non-coding pieces. We do not completely comprehend the entire recognition process, but we will get there,” explains Torben Heick Jensen.
RNA may have an unknown effect
Overall, the research should improve understanding of whether the apparently useless RNA has other functions than being a dynamic toolbox for evolution.
Torben Heick Jensen’s research has shown that RNA has a very short lifespan inside the cell nucleus, but part of the RNA may still have a function in the short time it exists before it is degraded.
“Right now, we think that much of it is just produced so it can be degraded again, but some of it may well have a transient effect that we would also like to characterize,” says Torben Heick Jensen.
“The MTR4 helicase recruits nuclear adaptors of the human RNA exosome using distinct arch-interacting motifs” has been published in Nature Communications. “Escaping nuclear decay: the significance of mRNA export for gene expression” has been published in Current Genetics. “Controlling nuclear RNA levels” has been published in Nature Reviews Genetics. In 2018, the Novo Nordisk Foundation awarded Torben Heick Jensen a Challenge Programme grant for the project Function, Structure, Regulation and Targeting of Exosome Adaptor Complexes.