ArticleNature structural & molecular biology2024
Single-base tiled screen unveils design principles of PspCas13b for potent and off-target-free RNA silencing.
Article in Nature structural & molecular biology, 2024. The graph could read no effect estimate from its abstract, so it casts no vote on the map. Cited by 14 papers.
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Who cites it
14 citing papers in PubMed.
- Programmable and selective silencing of oncogenic fusion transcripts by CRISPR-Cas13b via breakpoint sequence recognition.Science advances · 2026Article
- CRISPR in clinical oncology: translational advances from molecular diagnostics to therapeutics.Nature reviews. Clinical oncology · 2026Review
- Single-Molecule Imaging of Endogenous RNA Using a Miniaturized and Circularly Permuted CRISPR-Cas13 Degron-Masking System CDegSR.Advanced science (Weinheim, Baden-Wurttemberg, Germany) · 2026Article
- Craspase Protease Activation Is Sensitive to Oncogenic Single-Nucleotide RNA Mismatches.ACS chemical biology · 2026Article
- DUSP11 is an RNA triphosphatase that limits PspCas13b activity by destabilizing gRNA abundance in mammalian cells.Nucleic acids research · 2026Article
- Molecular basis of target RNA cleavage by Cas13.Nature communications · 2026Article
- Characterization of gRNA-dependent and gRNA-independent off-target binding sites of PspCas13b and RfxCas13d in mammalian cells.Nucleic acids research · 2026Article
- Role of Non-Coding RNAs in Regulating PD-L1 Expression in Breast Cancer: Emerging Insights and Implications.Current cancer drug targets · 2026Review
- Systematic evaluation of CrRNA design parameters for optimized Cas13d-mediated RNA targeting in chicken cells.Functional & integrative genomics · 2025Article
- Structural insights into RNA-guided RNA editing by the Cas13b-ADAR2 complex.Nature structural & molecular biology · 2025Article
- Article
- A Manual for Genome and Transcriptome Engineering.IEEE reviews in biomedical engineering · 2025Review
- Single-base tiled screen unveils design principles of PspCas13b for potent and off-target-free RNA silencing.Nature structural & molecular biology · 2024Article
- Article
Corrections and comments
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Authors and funding
14 authors.
Funding
No grant is acknowledged in the PubMed record.
Abstract
The development of precise RNA-editing tools is essential for the advancement of RNA therapeutics. CRISPR (clustered regularly interspaced short palindromic repeats) PspCas13b is a programmable RNA nuclease predicted to offer superior specificity because of its 30-nucleotide spacer sequence. However, its design principles and its on-target, off-target and collateral activities remain poorly characterized. Here, we present single-base tiled screening and computational analyses that identify key design principles for potent and highly selective RNA recognition and cleavage in human cells. We show that the de novo design of spacers containing guanosine bases at precise positions can greatly enhance the catalytic activity of inefficient CRISPR RNAs (crRNAs). These validated design principles (integrated into an online tool, https://cas13target.azurewebsites.net/ ) can predict highly effective crRNAs with ~90% accuracy. Furthermore, the comprehensive spacer-target mutagenesis revealed that PspCas13b can tolerate only up to four mismatches and requires ~26-nucleotide base pairing with the target to activate its nuclease domains, highlighting its superior specificity compared to other RNA or DNA interference tools. On the basis of this targeting resolution, we predict an extremely low probability of PspCas13b having off-target effects on other cellular transcripts. Proteomic analysis validated this prediction and showed that, unlike other Cas13 orthologs, PspCas13b exhibits potent on-target activity and lacks collateral effects.
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