Fanzor: First CRISPR-Like System Found in Eukaryotes

GEN BiotechnologyVol. 2, No. 4 News FeaturesFree AccessFanzor: First CRISPR-Like System Found in EukaryotesGEN Staff WriterGEN Staff WriterPublished Online:17 Aug 2023https://doi.org/10.1089/genbio.2023.29110.gswAboutSectionsPDF/EPUB Permissions & CitationsPermissionsDownload CitationsTrack CitationsAdd to favorites Back To Publication ShareShare onFacebookTwitterLinked InRedditEmail Feng Zhang's team at the Broad Institute discovers RNA-guided Fanzor enzymes from fungi and amoeba that show promise for genome editing applications.A cryo-EM map of a Fanzor protein (gray, yellow, light blue, and pink) in complex with ωRNA (purple) and its target DNA (red). Non-target DNA strand in blue.(Credit: Zhang lab, Broad Institute of MIT and Harvard/McGovern Institute for Brain Research at MIT).Another potentially game-changing discovery in the world of genome editing has come out of the Broad Institute lab of Feng Zhang. In a study published in Nature, Zhang and colleagues have uncovered the first programmable RNA-guided system in eukaryotes.1 Earlier this year, the Zhang lab adapted a contractile injection system, found naturally in bacteria, that delivers protein payloads to target human cells.2In the latest study, Zhang's team describes how the eukaryotic system—based on the Fanzor protein—uses RNA as a guide to target DNA precisely and shows that Fanzor can be reprogrammed to edit the genome of human cells. In addition, the compact Fanzor systems have the potential to be more easily delivered to cells and tissues as therapeutics than CRISPR-Cas systems. Further refinements to improve their targeting efficiency could make them a valuable new technology for human genome editing.Scientists have long wondered whether systems similar to the well-known bacterial CRISPR system exist in eukaryotes. In a recent analysis published in The CRISPR Journal (a sister journal to GEN Biotechnology), Blake Wiedenheft's group at Montana State University studied repetitive elements that had previously been identified as putative CRISPR loci in the human genome. They showed that these repeats do not contain the repeat-spacer-repeat architecture nor the cas nuclease genes characteristic of functional CRISPR systems.3“Overall, the evidence for functional CRISPR-Cas systems in the human genome and other eukaryotic genomes remains unsupported,” the authors wrote. “Although the phylogenetic and functional diversity of CRISPRs continue to expand, (to date) CRISPR-Cas adaptive immune systems remain restricted to prokaryotes.”3Present and CorrectNevertheless, the new Zhang lab study demonstrates that RNA-guided DNA-cutting mechanisms are present across all kingdoms of life.“CRISPR-based systems are widely used and powerful because they can be easily reprogrammed to target different sites in the genome,” said Zhang, a core institute member at the Broad Institute. “This new system is another way to make precise changes in human cells, complementing the genome editing tools we already have.”A decade ago, in January 2013, Zhang and his former supervisor, George Church, independently provided the first demonstrations that the bacterial CRISPR-Cas9 editing system worked in eukaryotic cells.A major aim of the Zhang lab is to develop genetic medicines using systems that can modulate human cells by targeting specific genes and processes. “A number of years ago, we started to ask, ‘What is there beyond CRISPR and are there other RNA-programmable systems out there in nature?’” Zhang recalled.Two years ago, Zhang lab members discovered a class of RNA-programmable systems in prokaryotes called OMEGAs, which are often linked with transposable elements in bacterial genomes and likely gave rise to CRISPR-Cas systems. That study also highlighted similarities between prokaryotic OMEGA systems and Fanzor proteins in eukaryotes, suggesting that the Fanzor enzymes might also use an RNA-guided mechanism to target and cut DNA.In the latest study, the researchers continued their study of RNA-guided systems by isolating Fanzor from fungi, algae, and amoeba species, in addition to a clam—the Northern Quahog.The biochemical characterization of the Fanzor proteins showed that they are DNA-cutting endonuclease enzymes that use nearby noncoding RNAs (ωRNAs) to target particular sites in the genome. It is the first time this mechanism has been found in eukaryotes.Unlike CRISPR proteins, Fanzor enzymes are encoded in the eukaryotic genome within transposable elements. Zhang's team's phylogenetic analysis suggests that the Fanzor genes have migrated from bacteria to eukaryotes through horizontal gene transfer.The researchers then demonstrated that Fanzor can generate insertions and deletions at targeted genome sites within human cells. They found the Fanzor system initially to be less efficient at snipping DNA than CRISPR-Cas systems, but genetically engineered the protein to increase its activity 10-fold. In addition, unlike some CRISPR systems and the OMEGA protein TnpB, Zhang's team found that a fungal-derived Fanzor protein did not exhibit “collateral activity,” where an RNA-guided enzyme cleaves its DNA target as well as degrading nearby DNA or RNA. The results suggest that Fanzor could potentially be developed as efficient genome editors.The group also analyzed the molecular structure of the Fanzor/ωRNA complex. They found that Fanzor shares structural similarities with CRISPR-Cas12, but the interaction between the ωRNA and the catalytic domains of Fanzor is more extensive, suggesting that the ωRNA might play a role in the catalytic reactions.Similar to CRISPR-based systems, the Fanzor system can be easily reprogrammed to target specific genome sites. Zhang says it could one day be developed into a powerful new genome editing technology for research and therapeutic applications. The abundance of RNA-guided endonucleases such as Fanzor further expands the number of OMEGA systems known across kingdoms of life and suggests that there are more yet to be found.“Nature is amazing. There's so much diversity!” said Zhang. “There are probably more RNA-programmable systems out there, and we're continuing to explore and will hopefully discover more.”References1. Saito M, Xu P, Faure G, et al. Fanzor is a eukaryotic programmable RNA-guided endonuclease. Nature 2023; doi: 10.1038/s41586-023-06356-2 Google Scholar2. Grinstein J. Nature's needles: Adapting bacterial “syringes” into programmable protein delivery devices. GEN Biotechnol 2023;2:162–166; doi: 10.1089/genbio.2023.29098.jdg Link, Google Scholar3. Buyukyoruk M, Henriques WS, Wiedenheft B. Clarifying CRISPR: Why repeats identified in the human genome should not be considered CRISPRs. CRISPR J 2023;6:216–221; doi: 10.1089/crispr.2022.0106 Link, Google ScholarFiguresReferencesRelatedDetails Volume 2Issue 4Aug 2023 InformationCopyright 2023, Mary Ann Liebert, Inc., publishersTo cite this article:GEN Staff Writer.Fanzor: First CRISPR-Like System Found in Eukaryotes.GEN Biotechnology.Aug 2023.276-277.http://doi.org/10.1089/genbio.2023.29110.gswPublished in Volume: 2 Issue 4: August 17, 2023PDF download