The director’s cut: how CAG repeats change the editing of genetic messages

Long repetitive sequences of C-A-G letters in the DNA code are associated with at least 12 genetic diseases, including Huntington’s disease (HD). A group of scientists in Massachusetts, USA, have recently developed a new genetic strategy to study how CAG repeats can lead to harmful proteins being made in cells, causing cells to become unhealthy. Their findings showed that expanded CAG repeats can interfere with a process called ‘splicingsplicing the cutting up of RNA messages, to remove non-coding regions and join together coding regions.’, which chops up and organises genetic message molecules before they are turned into proteins.

CAG repetition

Our DNA is a genetic code that holds instructions for making thousands of different proteins, the molecular machines that run our cells. This code is made of four building blocks or ‘bases’: C, A, G, and T. DNA is arranged like a twisted ladder with two DNA strands bound together in a helix, each made of a string of bases. The bases on one DNA strand pair with bases on the opposite DNA strand to form the ‘rungs’ of the ladder.

HD is known as a ‘CAG repeatCAG repeat The stretch of DNA at the beginning of the HD gene, which contains the sequence CAG repeated many times, and is abnormally long in people who will develop HD expansion disease’. Everyone has a repetitive sequence of C-A-G DNA letters in their huntingtin gene, but people who go on to develop HD have over 36 C-A-G repeats. The number of CAG repeats can increase over time, called repeat expansion, and this seems to happen mainly in cells that get the most unhealthy in HD such as brain cells.

If we can understand exactly how a longer CAG repeatCAG repeat The stretch of DNA at the beginning of the HD gene, which contains the sequence CAG repeated many times, and is abnormally long in people who will develop HD itself makes cells sick, we may be able to keep brain cells healthy and delay when HD symptoms appear. There are also other diseases caused by expansions in CAG repeats, including spinocerebellar ataxias and myotonic dystrophies. Trying to find similarities between what happens in cells affected by these other diseases may help us learn more about what goes on in HD.

Cutting scenes in the genetic script

When a cell wants to make a protein coded by a certain gene, the two DNA strands unwind and separate from each other. Cellular machinery then reads the opened-up DNA base code and makes a copy of it, called an RNARNA the chemical, similar to DNA, that makes up the 'message' molecules that cells use as working copies of genes, when manufacturing proteins. message molecule, a bit like making a photocopy of a recipe from a cookery book.

However, before any RNARNA the chemical, similar to DNA, that makes up the 'message' molecules that cells use as working copies of genes, when manufacturing proteins. message molecules are read by the next set of cellular machinery to make the corresponding protein, an essential process needs to take place. Much like editing out unnecessary scenes from a film to make a final polished version, this process involves editing the RNARNA the chemical, similar to DNA, that makes up the 'message' molecules that cells use as working copies of genes, when manufacturing proteins. message to remove all of the waffly bits of genetic code copied from DNA which aren’t actually needed to make a protein. The process of going from the unedited RNARNA the chemical, similar to DNA, that makes up the 'message' molecules that cells use as working copies of genes, when manufacturing proteins. message molecule to a shorter more succinct message is called ‘splicingsplicing the cutting up of RNA messages, to remove non-coding regions and join together coding regions.’. During splicingsplicing the cutting up of RNA messages, to remove non-coding regions and join together coding regions., non-essential sections of the unedited message are cut out and the important sections that remain are pasted together to produce what is known as ‘mature’ RNARNA the chemical, similar to DNA, that makes up the 'message' molecules that cells use as working copies of genes, when manufacturing proteins.. This final mature RNARNA the chemical, similar to DNA, that makes up the 'message' molecules that cells use as working copies of genes, when manufacturing proteins. product has only the necessary instructions that the cell needs to make proteins.

Expanded CAG repeats can cause genetic plot twists

In diseases caused by expanding CAGs, the CAG repeatCAG repeat The stretch of DNA at the beginning of the HD gene, which contains the sequence CAG repeated many times, and is abnormally long in people who will develop HD in the DNA is copied into the RNARNA the chemical, similar to DNA, that makes up the 'message' molecules that cells use as working copies of genes, when manufacturing proteins. message, which can cause abnormal proteins to be made. In the case of HD, an extra-long version of the huntingtin proteinhuntingtin protein The protein produced by the HD gene. is made. A group of scientists led by Dr Jain in Cambridge, Massachusetts, previously found that repeat-containing RNARNA the chemical, similar to DNA, that makes up the 'message' molecules that cells use as working copies of genes, when manufacturing proteins. messages, along with the proteins made from them, combine to form toxic clumps in cells which can cause serious damage.

To find out exactly how longer CAG repeats cause the production of harmful RNARNA the chemical, similar to DNA, that makes up the 'message' molecules that cells use as working copies of genes, when manufacturing proteins. and proteins, Rachel Anderson and colleagues within the Jain team recently developed a clever new method to look in detail at the precise genetic message in RNARNA the chemical, similar to DNA, that makes up the 'message' molecules that cells use as working copies of genes, when manufacturing proteins. molecules containing large CAG repeats. Interestingly, they found that CAG repeats in RNARNA the chemical, similar to DNA, that makes up the 'message' molecules that cells use as working copies of genes, when manufacturing proteins. cause mistakes to be made during splicingsplicing the cutting up of RNA messages, to remove non-coding regions and join together coding regions. of that RNARNA the chemical, similar to DNA, that makes up the 'message' molecules that cells use as working copies of genes, when manufacturing proteins. message molecule. Expanded CAG repeats in RNARNA the chemical, similar to DNA, that makes up the 'message' molecules that cells use as working copies of genes, when manufacturing proteins. cause other sections of the message molecule, sometimes far away from the CAG repeatCAG repeat The stretch of DNA at the beginning of the HD gene, which contains the sequence CAG repeated many times, and is abnormally long in people who will develop HD itself, to be cut and pasted into or next to the repeat during splicingsplicing the cutting up of RNA messages, to remove non-coding regions and join together coding regions..

Here, the expanded CAG repeatCAG repeat The stretch of DNA at the beginning of the HD gene, which contains the sequence CAG repeated many times, and is abnormally long in people who will develop HD can act like the opening credits of a film, into which the final scenes of the film get mistakenly inserted out of order. When this happens, the plot of the film no longer makes sense. Similarly, the final RNARNA the chemical, similar to DNA, that makes up the 'message' molecules that cells use as working copies of genes, when manufacturing proteins. message doesn’t make much sense when other sections of genetic information are inserted into the CAG repeatCAG repeat The stretch of DNA at the beginning of the HD gene, which contains the sequence CAG repeated many times, and is abnormally long in people who will develop HD during splicingsplicing the cutting up of RNA messages, to remove non-coding regions and join together coding regions.. This leads to the creation of many different repeat-containing mature RNAs with unexpected sequences.

The researchers found that the longer the CAG repeatCAG repeat The stretch of DNA at the beginning of the HD gene, which contains the sequence CAG repeated many times, and is abnormally long in people who will develop HD in the RNARNA the chemical, similar to DNA, that makes up the 'message' molecules that cells use as working copies of genes, when manufacturing proteins. message, the more faulty splicingsplicing the cutting up of RNA messages, to remove non-coding regions and join together coding regions. events that occurred. This is interesting as the CAG number in HD tracks with the age at which symptoms start and the rate at which they progress. The researchers showed that when they stopped all splicingsplicing the cutting up of RNA messages, to remove non-coding regions and join together coding regions. events in cells using a chemical, repeat-containing RNARNA the chemical, similar to DNA, that makes up the 'message' molecules that cells use as working copies of genes, when manufacturing proteins. messages did not form clumps in cells and so did not cause cell toxicity.

Protein production glitches

So far, these results explain how expanded CAG repeats lead to abnormal and incorrectly spliced mature RNARNA the chemical, similar to DNA, that makes up the 'message' molecules that cells use as working copies of genes, when manufacturing proteins. messages, but what happens when these messages are read to make proteins? Any mature RNAs that are ready to be read by cellular machinery to make a protein contain a ‘start’ signal, like a green traffic light. The researchers found that sometimes when repeat-containing RNAs are incorrectly spliced, more of these start signals are found before the repeat, causing many different proteins to be made from a single RNARNA the chemical, similar to DNA, that makes up the 'message' molecules that cells use as working copies of genes, when manufacturing proteins. message than normal. The researchers altered these start signals in the CAG repeatCAG repeat The stretch of DNA at the beginning of the HD gene, which contains the sequence CAG repeated many times, and is abnormally long in people who will develop HD-containing RNAs to turn them off and found that this stopped abnormal proteins from being made.

The researchers also studied the RNARNA the chemical, similar to DNA, that makes up the 'message' molecules that cells use as working copies of genes, when manufacturing proteins. messages containing CAGs that were copied from genes associated with CAG repeatCAG repeat The stretch of DNA at the beginning of the HD gene, which contains the sequence CAG repeated many times, and is abnormally long in people who will develop HD expansion diseases, including spinocerebellar ataxiaspinocerebellar ataxia A family of diseases which result in characteristic movement disorders. Many types of spinocerebellar ataxia are caused by the same type of mutation as HD – a CAG expansion. and myotonic dystrophy. The researchers showed that expanded CAGs copied from these genes also caused abnormal splicingsplicing the cutting up of RNA messages, to remove non-coding regions and join together coding regions. into the repeat, which again contained more protein reading start signals which may cause more abnormal proteins to be made.

What does this mean for CAG repeatCAG repeat The stretch of DNA at the beginning of the HD gene, which contains the sequence CAG repeated many times, and is abnormally long in people who will develop HD expansion diseases?

Understanding how important processes in cells are impacted by long CAG repeats can help researchers piece together exactly how cells become unhealthy in CAG repeatCAG repeat The stretch of DNA at the beginning of the HD gene, which contains the sequence CAG repeated many times, and is abnormally long in people who will develop HD expansion diseases and point to which processes can be targeted with therapeuticstherapeutics treatments. The findings from this study add another piece to the puzzle of what happens in cells, suggesting expanded CAG repeats in RNARNA the chemical, similar to DNA, that makes up the 'message' molecules that cells use as working copies of genes, when manufacturing proteins. interfere with splicingsplicing the cutting up of RNA messages, to remove non-coding regions and join together coding regions., which can lead to damaging proteins being made.

Importantly, these experiments were performed in cell types, such as kidney cells, which are easy to grow and manage in the lab but are not most affected by HD. Therefore, these cells may not accurately reflect what causes cells to become sick in HD. A lot more work is needed looking at how expanded repeats alter RNARNA the chemical, similar to DNA, that makes up the 'message' molecules that cells use as working copies of genes, when manufacturing proteins. splicingsplicing the cutting up of RNA messages, to remove non-coding regions and join together coding regions. and protein production in cell and animal models of HD. Nonetheless, targeting splicingsplicing the cutting up of RNA messages, to remove non-coding regions and join together coding regions. may be a potentially exciting avenue that researchers can pursue to develop medicines for HD and other repeat expansion diseases.

Learn more

• CAG repeat expansions create splicing acceptor sites and produce aberrant repeat-containing RNAs (full article requires payment or subscription)