A genetic editing system similar to CRISPR-Cas9 has been uncovered for the first time in eukaryotes – the group of organisms that include fungi, plants, and animals. The system, based on a protein called Fanzor, can be guided to precisely target and edit sections of DNA, and that could open up the possibility of its use as a human genome editing tool.
The research team, led by Professor Feng Zhang at the McGovern Institute for Brain Research at MIT and the Broad Institute of MIT and Harvard, began to suspect that Fanzor proteins might act as nucleases – enzymes that can chop up nucleic acids, like DNA – during a previous investigation.
They were looking into the origins of proteins like Cas9. This is the enzyme part of the CRISPR-Cas9 system. CRISPR (short for clustered regularly interspaced short palindromic repeats) sequences are the guide to particular regions of DNA, and Cas9 makes the cut. We hear a lot about CRISPR-Cas systems and their applications in medicine and biotechnology, but you may not be aware that they originate in bacteria, where they play a key role in immunity against viruses.
When studying Cas9 and other related proteins, Zhang’s team discovered their ancestors, a class of proteins they named OMEGA proteins. One of these, TnpB, bore a remarkable resemblance to a protein found in eukaryotes: Fanzor.
“Because of the conservation between TnpB and Fanzor, we had a good reason to think that Fanzor is most likely also an RNA-guided OMEGA nuclease. So after we finished the OMEGA study on IscB [another OMEGA protein] and TnpB, we focused on studying Fanzor,” Zhang told IFLScience.
In this latest study, the team isolated Fanzors from fungi, algae, amoebae, and the northern quahog clam. With the leadership of co-first author Makoto Saito, the function of the proteins was characterized, showing that they were, as suspected, DNA-cutting enzymes. Just as Cas9 is guided by CRISPR fragments, Fanzors are guided by sections of RNA called ωRNAs.
Co-first author Peiyu Xu led another set of experiments to look at the molecular structure of the Fanzor-ωRNA complex, to show precisely how the proteins interact and attach themselves to the DNA sequence that is to be snipped.
How does Fanzor compare to CRISPR-Cas systems?
“The Fanzor systems are more compact than CRISPR proteins and therefore have the potential to be more easily delivered to cells and tissues. Fanzor enzymes are also encoded in the eukaryotic genome within transposable elements,” Zhang explained to IFLScience. “Unlike CRISPR systems, which are adaptive immune systems, the function of Fanzor is still not clear.”
Another key difference with Fanzor is the lack of “collateral damage”. With some CRISPR systems and the TnpB OMEGA protein, there is a risk of off-target effects, where the enzyme cleaves not only the desired portion of DNA but also damages nearby sections of the molecule. This does not seem to be the case with the fungal Fanzor protein that the team studied in detail.
On top of this, although Fanzor initially seemed to be less efficient than CRISPR-Cas systems, the team was able to engineer the protein to achieve a 10-fold increase in activity.
It took many years and a huge amount of research for scientists to begin to harness the potential of CRISPR-Cas. Whilst this new work is an exciting development, it is still too early to fully understand what the impact will be.
“We are excited to see how the trajectory unfolds, and we are continuing to work to develop Fanzor into a valuable new technology for human genome editing,” Zhang told IFLScience. “Additionally, it is quite exciting to see the existence of CRISPR-like proteins in animal cells.”
“Going forward, we are continuing to study the biology of Fanzor proteins and exploring ways that we can engineer them for use as molecular technologies. We still need to engineer the enzyme further so that it will match the efficacy of the Cas9 gold standard.”
“Aside from the potential offered by the small size of Fanzor, this work really underscores that there are likely more RNA-guided systems out there in nature that hold future promise for gene editing,” Zhang added. “This is another example of the power of studying biodiversity. There are likely many more interesting and potentially useful systems waiting to be discovered and harnessed.”
The study is published in Nature.