Study identifies how twin proteins work together to patch up DNA damage

Cutting-edge technology has provided highly valuable new insights into how a pair of molecules, which occur naturally in the body, play an essential role in the DNA repair process.

The findings increase our understanding of what goes wrong in the cells of some cancer patients and could potentially inform the development of cancer treatments in the future.

While healthy DNA carries genetic instructions telling our cells how to function, if it is damaged and mutated, it can cause those cells to behave abnormally, potentially leading to diseases such as cancer.

To help repair DNA damage, the body produces twin molecules known as Rad51 filaments and Rad51 paralogs, which are part of a family of proteins that play a vital role in DNA repair. These proteins are already known to be important as people who inherit a faulty version of the genes that make them have a higher risk of developing breast and ovarian cancers.  The discovery of a mutation in one of these genes often leads people to take preventative action - Angelina Jolie famously underwent a double mastectomy in 2013 after discovering that she had a defect in the gene that produces BRCA1, a protein intimately related to Rad51. 

In a pioneering study in 2015 using nematode worms to study the function of the twin Rad51 molecules, it was discovered that they work together to repair DNA, but, until now, the mechanism of exactly how they do this has been a mystery that has foxed the scientific community.

In order to repair breaks in strands of DNA (a process known as homologous recombination), Rad51 filaments first wrap around it. They then hunt around the genome looking for a section of intact DNA similar to the part that has been damaged. When they find a match, they copy and paste that sequence, patching up the gap where damage has occurred.

Rad51 filaments, however, cannot act alone. To do their job effectively, they need the help of their twin molecules Rad51 paralogs. It has been observed that paralogs have three beneficial effects on filaments: They cause filaments to become less tightly wrapped, they enable them to become more flexible, and slow down the process by which the filaments release themselves from the DNA. The first two effects allow the filaments to search the genome more effectively; the third makes the repair process more stable overall.  

Now, in a new study, published in Molecular Cell, Martin Taylor, a Research Associate at St John’s College, University of Cambridge, who carried out much of the initial research at the Francis Crick Institute, has discovered more about how this partnership between filaments and paralogs functions at a molecular level.

Teaming up with researchers in the Czech Republic, including co-authors Mario Spirek and Lumir Krejci, Taylor and colleagues used a cutting-edge technique called stopped flow analysis to look at Rad51 proteins harvested from nematode worms. This technology allowed the scientists to monitor how the DNA repair process evolves in real time and watch what happens to Rad51 filaments when Rad51 paralogs are present.

Looking along the DNA strand, which was made visible using florescent molecules, they noticed that the influence paralogs had on the filaments was different from one end to the other. At one end there was a stronger stabilising effect, which got weaker towards the middle. By the other end there was no effect at all.

A technique called electron microscopy, which uses a powerful microscope and particles of gold to make proteins visible, provided evidence that paralogs tend to accumulate at one end of a filament strand.

Taylor said: “We think this means that paralogs dock on one end of the filaments, and propagate structural changes towards the other, which help the filaments to repair DNA damage. This helps to explain why we can observe their effects working better at one end.

“This is the first time that anyone has proposed how such a mechanism might work and it opens up a range of possibilities in the field for discovering more about how similar proteins are regulated. The next step is to find out if Rad51 and the paralogs work in the same way in humans as in worms, as having a better idea of what these proteins are doing and what causes them to work more efficiently to repair DNA could help us to understand more about what goes wrong in the case of cancer.”