RiboSwitches: Working smarter, not harder

What are riboswitches?

Are you one of those people who gets really annoyed when lights are on unnecessarily during daylight? What if we told you that the concept of saving energy is something present at a molecular level in all organisms, and that by using it we can get an insight into things like genetic structure or protein expression; and even develop novel systems to detect all sorts of molecules, from metabolic compounds to pollutants?

But in order to do that, we must get into the world of microbiology. Most microorganisms have an array of possible responses to their environment, which vary in the presence of, let’s say, nutrients, when compared to competitors. Under some conditions, some bacterial colonies reach a state known as quorum sensing, where all the elements of the colony behave in a specific, collective way. These responses only occur in certain circumstances, as doing so in others will not be as efficient for the colony, and could even have detrimental effects.

One way bacteria can regulate the activation of these states is via an element called a riboswitch. Riboswitches are short mRNA (messenger RNA) sequences, which are able to change their spatial conformation in the presence of a specific molecule of interest, or ligand. This change in its conformation is shown as a change in the gene expression of the sequences downstream from the riboswitch.

By doing this, bacteria can turn on specific genes ONLY when they are needed, and save the energy and resources by stopping these molecules from being produced constantly.

 

Image showing a riboswitch before and after ligand binding
The change in conformation of a riboswitch in the absence and presence of the ligand that binds to the RNA. Without the ligand, the ribosome is free to recognise the ribosome binding sire (RBS) and begin translation. When the ligand is present, the change in conformation hides the RBS and the protein cannot be made. Diagram from iGEM team Ribonostics from Exeter.

 

Riboswitches are extensively found in both bacteria and archaea, with only a few found in eukaryotes. This supports the theory that these elements have been conserved throughout evolution, since the early stages of life; and may even be remnants of what’s known as the RNA world, a theoretical stage of life where self-replicating RNA molecules existed before the appearance of DNA or proteins.

Nowadays, improved riboswitches have been developed based on pre-existing natural ones with increased affinity for their substrate, a higher specificity to discern between similar compounds, or even a different compound affinity altogether. At the same time, fully synthetic riboswitches have been developed, by coupling a synthetic aptamer (the element whose conformation changes, ie the RNA) with an expression platform (the element that makes use of that conformational change to regulate gene expression, ie the ribosome). This theoretically means that a riboswitch can be developed for any small molecule, but sadly, it is not that easy.

How to make riboswitches

The current process by which aptamers are developed is known as SELEX (Systematic Evolution of Ligands by Exponential enrichment), a process where a library of random oligonucleotides is exposed to the ligand of interest. Those members of the library which successfully bind the ligand with a certain degree of affinity will then be selected, amplified, and then undergo a second, more stringent selection process. This will be iterated until a sequence with the desired properties is obtained.

SELEX, however, has a big problem: that it is done in vitro. This means that even the best SELEX-produced aptamer may not work in vivo. Several technologies are being developed nowadays to work around this caveat, such as the directed evolution of whole riboswitches, a process that can be done in vivo and has shown promising results in other instances.

Why make riboswitches?

All of this matters because nowadays riboswitches are highly important in synthetic biology, for many reasons. They can be used to induce or inhibit the expression of genes of interest in research settings, or for industrial purposes. So new riboswitches could be developed and implemented for their use in disease treatments, pollutant detection, or bioremediation, and that is only the beginning.

We all know about the current antibiotic crisis. With the SELEX process, riboswitches could be developed to detect specific by-products of certain antibiotic-resistant microorganisms, and then cloned into specific bacteria or even viruses, so that the moment the by-product is detected, a pathway is activated which ends with the killing of the infecting microorganisms. Once the infection has been dealt with, it can be made so that the riboswitch carrier would not be able to survive any longer, and so it is eliminated from the organism.

That same philosophy can be applied to bioremediation, creating bacteria that degrade the pollutants and then die off, eliminating possible environmental spread.

There is still a lot of work to do in this area, but the prospects are certainly encouraging, and it could mean a new way of understanding industrial procedures, or battle toxic compounds.

Remember, work smarter, not harder.

A more detailed description of riboswitches and the mechanism behind them can be found in Riboswitches: Structures and Mechanisms and a review of some of the applications of synthetic riboswitches can be found in Synthetic riboswitches — A tool comes of age. Finally, just a few weeks ago, researchers from Okinawa Institute of Science and Technology Graduate University  used riboswitches to understand more about the ‘living antibiotic’ B. bacteriovorus and how it controls its predatory lifecycle.

 

Thanks for reading!

Do you have any possible ideas where a system such as a riboswitch could be implemented?

Tell us!

Today’s piece was written by Eduardo Goicoechea Serrano, a synthetic biology PhD student at Warwick. For any questions about riboswitches, or being a PhD student, tweet Eduardo @EduGoico

Leave a Reply

Your email address will not be published. Required fields are marked *