Ontology highlight
ABSTRACT:
PROVIDER: PRJNA991924 | ENA |
REPOSITORIES: ENA
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Project description:As the ancestor of CRISPR-Cas12 nucleases, TnpB represents the most compact gene editing tool currently available. Recent studies have identified multiple TnpB systems with gene editing activity in mammalian cells, and the potential of TnpB in treating diseases has been demonstrated in animal models. However, the editing characteristics of various TnpB systems, comparable to CRISPR tools, require more extensive investigation. Using a standardized evaluation framework, we conducted a thorough analysis of the editing properties of four TnpB variants alongside representative Cas12 and Cas9 tools. Overall, TnpBs exhibit intermediate editing activity and safety profiles among all tested systems, with ISYmu1 TnpB demonstrating a good performance in both editing activity and specificity. Considering its compact size, potent editing efficiency and high specificity, ISYmu1 TnpB represents a promising candidate for in vivo gene therapy applications.
2025-01-01 | GSE264389 | GEO
Project description:Virus-induced genome editing (VIGE) using compact RNA-guided endonucleases is a transformational new approach in plant biotechnology, enabling tissue-culture-independent and transgene-free genome editing. We recently established a transgene-free VIGE approach for heritable editing at single loci in Arabidopsis by delivering ISYmu1 TnpB (Ymu1) and its guide RNA (gRNA) via Tobacco Rattle Virus (TRV). Here, we greatly improved this system by devising a multiple gRNA expression system and by utilizing an engineered high-activity Ymu1 variant (Ymu1-WFR) to develop an efficient multiplexed genome editing approach.
2026-01-15 | GSE316183 | GEO
Project description:TnpB is a compact RNA-guided endonuclease and evolutionary ancestor of CRISPR-Cas12 that offers a promising platform for genome engineering. However, the genome-editing activity of TnpBs remains limited and its underlying determinants are poorly understood. Here, we used biochemical and single-molecule assays to examine the DNA-unwinding mechanism of Youngiibacter multivorans TnpB (Ymu1 TnpB). DNA unwinding proceeds through formation of a partially unwound intermediate state to a fully unwound open state. The open state forms inefficiently and collapses readily in the absence of negative supercoiling. An optimized variant, Ymu1-WFR, stabilizes formation of both the intermediate and open states, resulting in enhanced DNA cleavage in vitro and increased genome editing in vivo. These findings identify the physical basis for the observed minimal activities of natural TnpBs, revealing how stabilizing specific unwinding states enables efficient DNA targeting.
2026-04-17 | GSE327273 | GEO
Project description:RNA-guided endonucleases form the crux of diverse biological processes and technologies, including adaptive immunity, transposition, and genome editing. Some of these enzymes are components of insertion sequences (IS) in the IS200/IS605 and IS607 transposon families. Both IS families encode a TnpA transposase and TnpB nuclease, an RNA-guided enzyme ancestral to CRISPR-Cas12. In eukaryotes and their viruses, TnpB homologs occur as two distinct types, Fanzor1 and Fanzor2. We analyzed the evolutionary relationships between prokaryotic TnpBs and eukaryotic Fanzors, revealing that a clade of IS607 TnpBs with unusual active site arrangement found primarily in cyanobacteria likely gave rise to both types of Fanzors. The widespread nature of Fanzors imply that the properties of this particular group of IS607 TnpBs were particularly suited to adaptation and evolution in eukaryotes and their viruses. Biochemical analysis of a prokaryotic IS607 TnpB and virally encoded Fanzor1s revealed features that may have fostered co-evolution between TnpBs/Fanzors and their cognate transposases. These results provide insight into the evolutionary origins of a ubiquitous family of RNA-guided proteins that shows remarkable conservation across the three domains of life.
2023-11-18 | GSE246134 | GEO
Project description:Fanzor proteins are recently identified from eukaryotic cells as RNA-guided nuclease homologs of prokaryotic TnpB. However, Fanzor-ωRNA system has not been well characterized in terms of gene editing activity, off-target profile and in vivo efficacy for disease correction. In the present study, Fanzor and other TnpB-ωRNA systems were reconstituted in mammalian cells for gene editing and off-target analysis. Furthermore, we identified several Fanzor variants with enhanced activity via structure-guided protein engineering. Additionally, we demonstrated the efficacy of engineered Fanzor-ωRNA system in embryos to generate mutant mice via embryonic injection of Fanzor messenger RNA (mRNA) and ωRNA. Notably, single adeno-associated virus (AAV) delivery of engineered Fanzor-ωRNA system in a mouse model of Duchenne muscular dystrophy (DMD) achieved sufficient gene editing of DMD gene mutations to effectively induce mutant exons skipping and restore dystrophin expression, indicating the applications potential for single AAV gene editing therapy with Fanzor-ωRNA system.
2026-09-05 | GSE345606 | GEO
Project description:Stepwise DNA unwinding gates TnpB genome-editing activity
| PRJNA1450085 | ENA