An innovative study led by a team of scientists at the Broad Institute of MIT and Harvard and the McGovern Institute for Brain Research at MIT has revealed the first programmable RNA-guided system in eukaryotes, which includes organisms like fungi, plants, and animals. Published in Nature, the study introduces a system based on a protein called Fanzor, demonstrating its ability to precisely target DNA using RNA as a guide. Moreover, Fanzor proteins can be reprogrammed to edit the genome of human cells. The compact nature of the Fanzor system presents the potential for efficient delivery to cells and tissues, making it a promising alternative to CRISPR/Cas systems for therapeutic applications. Refinements to enhance its targeting efficiency could position it as a valuable tool for human genome editing.
Until now, CRISPR/Cas systems were primarily discovered in prokaryotes (single-cell organisms without nuclei), leaving scientists curious about the existence of similar systems in eukaryotes. This study establishes that RNA-guided DNA-cutting mechanisms are ubiquitous across all domains of life.
The research team, led by Feng Zhang, has been dedicated to developing genetic medicines that can modulate human cells by targeting specific genes and processes. Zhang's team embarked on a quest to explore RNA-programmable systems beyond CRISPR. Two years ago, they discovered a class of RNA-programmable systems called OMEGAs in prokaryotes, which are associated with transposable elements or "jumping genes" in bacterial genomes and likely gave rise to CRISPR/Cas systems. Notably, the researchers observed similarities between prokaryotic OMEGA systems and Fanzor proteins in eukaryotes, suggesting that Fanzor enzymes may employ an RNA-guided mechanism to target and cleave DNA.
In their latest study, the team isolated Fanzors from various species, including fungi, algae, amoebas, and a clam called the Northern Quahog. The researchers conducted biochemical characterizations of Fanzor proteins and demonstrated that they function as DNA-cutting endonuclease enzymes, utilizing non-coding RNAs known as ωRNAs to precisely target specific genomic sites. This RNA-guided mechanism represents a novel discovery in eukaryotes, including animals.
Unlike CRISPR proteins, Fanzor enzymes are encoded within transposable elements of the eukaryotic genome, suggesting their horizontal transfer from bacteria to eukaryotes.
To investigate Fanzor's potential as a genome editing tool, the researchers successfully generated insertions and deletions at targeted sites within human cells using the Fanzor system. Although initially less efficient at cleaving DNA compared to CRISPR/Cas systems, the team engineered specific mutations into the protein, resulting in a tenfold increase in its activity. Importantly, unlike certain CRISPR systems and the OMEGA protein TnpB, a fungal-derived Fanzor protein exhibited no collateral activity, indicating its potential as an efficient genome editor.
Structural analysis of the Fanzor/ωRNA complex, led by co-first author Peiyu Xu, revealed insights into how Fanzor binds to DNA for cleavage. Fanzor shares structural similarities with its prokaryotic counterpart, CRISPR-Cas12 protein, but exhibits more extensive interactions between the ωRNA and catalytic domains of Fanzor. This suggests a potential role for ωRNA in catalytic reactions. The team believes that these structural insights will contribute to further engineering and optimization of Fanzor, enhancing its efficiency and precision as a genome editor.
Similar to CRISPR-based systems, the Fanzor system can be easily reprogrammed to target specific genome sites. Zhang envisions its development into a powerful genome editing technology for research and therapeutic applications. The abundance of RNA-guided endonucleases like Fanzors expands our knowledge of OMEGA systems across different kingdoms of life, hinting at the existence of many more systems yet to be discovered.
Zhang expressed excitement about the vast diversity found in nature, stating, "There are probably more RNA-programmable systems out there, and we're continuing to explore and will hopefully discover more." With this pioneering research, the scientific community eagerly anticipates the future applications and potential breakthroughs that could arise from this newly uncovered programmable RNA-guided system in eukaryotes.
Until now, CRISPR/Cas systems were primarily discovered in prokaryotes (single-cell organisms without nuclei), leaving scientists curious about the existence of similar systems in eukaryotes. This study establishes that RNA-guided DNA-cutting mechanisms are ubiquitous across all domains of life.
The research team, led by Feng Zhang, has been dedicated to developing genetic medicines that can modulate human cells by targeting specific genes and processes. Zhang's team embarked on a quest to explore RNA-programmable systems beyond CRISPR. Two years ago, they discovered a class of RNA-programmable systems called OMEGAs in prokaryotes, which are associated with transposable elements or "jumping genes" in bacterial genomes and likely gave rise to CRISPR/Cas systems. Notably, the researchers observed similarities between prokaryotic OMEGA systems and Fanzor proteins in eukaryotes, suggesting that Fanzor enzymes may employ an RNA-guided mechanism to target and cleave DNA.
In their latest study, the team isolated Fanzors from various species, including fungi, algae, amoebas, and a clam called the Northern Quahog. The researchers conducted biochemical characterizations of Fanzor proteins and demonstrated that they function as DNA-cutting endonuclease enzymes, utilizing non-coding RNAs known as ωRNAs to precisely target specific genomic sites. This RNA-guided mechanism represents a novel discovery in eukaryotes, including animals.
Unlike CRISPR proteins, Fanzor enzymes are encoded within transposable elements of the eukaryotic genome, suggesting their horizontal transfer from bacteria to eukaryotes.
To investigate Fanzor's potential as a genome editing tool, the researchers successfully generated insertions and deletions at targeted sites within human cells using the Fanzor system. Although initially less efficient at cleaving DNA compared to CRISPR/Cas systems, the team engineered specific mutations into the protein, resulting in a tenfold increase in its activity. Importantly, unlike certain CRISPR systems and the OMEGA protein TnpB, a fungal-derived Fanzor protein exhibited no collateral activity, indicating its potential as an efficient genome editor.
Structural analysis of the Fanzor/ωRNA complex, led by co-first author Peiyu Xu, revealed insights into how Fanzor binds to DNA for cleavage. Fanzor shares structural similarities with its prokaryotic counterpart, CRISPR-Cas12 protein, but exhibits more extensive interactions between the ωRNA and catalytic domains of Fanzor. This suggests a potential role for ωRNA in catalytic reactions. The team believes that these structural insights will contribute to further engineering and optimization of Fanzor, enhancing its efficiency and precision as a genome editor.
Similar to CRISPR-based systems, the Fanzor system can be easily reprogrammed to target specific genome sites. Zhang envisions its development into a powerful genome editing technology for research and therapeutic applications. The abundance of RNA-guided endonucleases like Fanzors expands our knowledge of OMEGA systems across different kingdoms of life, hinting at the existence of many more systems yet to be discovered.
Zhang expressed excitement about the vast diversity found in nature, stating, "There are probably more RNA-programmable systems out there, and we're continuing to explore and will hopefully discover more." With this pioneering research, the scientific community eagerly anticipates the future applications and potential breakthroughs that could arise from this newly uncovered programmable RNA-guided system in eukaryotes.