DNA-guided CRISPR

DNA-guided CRISPR refers to engineered CRISPR-Cas systems that utilize synthetic DNA guides, rather than canonical RNA guides, to recognize and target RNA molecules.[1][2] These systems primarily utilize Cas12a and variants such as Cas12i1 to separate protein activation from target recognition.[3]

Mechanism of action

In native bacterial defense systems, Cas12 activation requires a CRISPR RNA scaffold that forms a pseudoknot structure. Engineered systems replace this with a synthetic single-stranded DNA molecule, referred to as ΨDNA or crDNA.[4]

Activation requires a Protospacer Adjacent Motif (PAM). The synthetic DNA guide mimics a crRNA framework and includes a 3' or 5' extension that binds the PAM-detecting region of Cas12. This interaction forms a stable deoxyribonucleoprotein (DNP) complex capable of binding complementary RNA strands.[3] Structural analyses have detailed the molecular basis of this ternary complex.[5] Target binding is enabled through molecular alignment.

Upon binding to target RNA, the Cas12-DNA complex undergoes a structural shift that activates the collateral cleavage of nearby single-stranded DNA. Depending on the Cas12 variant, the complex can degrade mRNA directly or physically block ribosomes to induce translational repression via no-go decay.[1] DNA-guided Cas12 systems preferentially bind RNA and rarely interact with single- or double-stranded DNA targets.[4]

Development

The foundation and system for DNA-guided Cas12 RNA targeting was first established publicly in a November 2024 pre-print by the Jain laboratory.[6] In 2026, the Jain laboratory and the Hsing laboratory (Wu et al.) published peer-reviewed validations of these guide architectures in Nature Biotechnology.[1][3] These parallel developments were highlighted in an independent review.[4] Subsequent optimizations of the guide backbones and spatial positioning enhanced binding affinity across orthologs such as AsCas12a and Cas12i1.

See also

References

  1. 1 2 3 Orosco, Carlos; Huang, Boyu; Rananaware, Santosh R.; Bodin, August P.; Browning, Ivy; Fang, Anne; Baugh, Michael P.; Lange, Ian H.; Elhabashy, Yasmin B.; Balaraju, Mahika; Lewis, Jordan G.; Shah, Nayan H.; Hanna, Michael P.; Flannery, Sarah J.; Meister, Katelyn S.; Karalkar, Vedant; Jain, Piyush K. (2026). "DNA-guided CRISPR/Cas12 for RNA targeting". Nature Biotechnology: 1–12. doi:10.1038/s41587-026-03129-w. PMID 42141118.
  2. ↑ Wiegel, Savannah. "DNA-guided CRISPR Suggests a New Direction for RNA Editing". Genetic Engineering & Biotechnology News.
  3. 1 2 3 Wu, Xiaolong; Lam, Wai Hei; Zhao, Zibin; Cao, Yumeng; Lin, Haosi; Feng, Xianzhen; Zhai, Yuanliang; Hsing, I-Ming (2026). "DNA-guided CRISPR-Cas12a effectors for programmable RNA recognition and cleavage". Nature Biotechnology: 1–11. doi:10.1038/s41587-026-03120-5. PMID 42067668.
  4. 1 2 3 Beisel, Chase L. (2026). "DNA-guided CRISPR". Nature Biotechnology: 1–2. doi:10.1038/s41587-026-03138-9. PMID 42141117.
  5. ↑ Ocampo, Rodrigo Fregoso; Orosco, Carlos; Huang, Boyu; West, Madeline S.; Jain, Piyush K.; Taylor, David W. (2026). "Architecture of a DNA-guided Cas12a". bioRxiv 10.64898/2026.03.19.712971.
  6. ↑ Orosco, Carlos; Rananaware, Santosh R.; Huang, Boyu; Hanna, Michael P.; Ahmadimashhadi, M. Reza; Lewis, Jordan G.; Baugh, Michael P.; Bodin, August P.; Flannery, Sarah J.; Lange, Ian H.; Fang, Zoe R.; Karalkar, Vedant N.; Meister, Katelyn S.; Jain, Piyush K. (2024). "DNA-guided CRISPR/Cas12 for RNA targeting". medRxiv 10.1101/2024.11.21.24317744.
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