Cas13 RNA editing flips the central dogma of CRISPR on its head. Where Cas9 and Cas12 cut DNA, Cas13 enzymes cut single-stranded RNA — and that single biochemical pivot has produced two of the most consequential CRISPR applications of the last decade: programmable RNA knockdown without permanent genome changes, and the SHERLOCK diagnostics platform that helped power COVID-19 testing. If your goal is to silence a gene without ever touching DNA, or to detect a virus from a single molecule of nucleic acid, Cas13 is the tool.
What Is Cas13?
Cas13 was discovered by Omar Abudayyeh, Jonathan Gootenberg, Feng Zhang, and colleagues, reported in Science in June 2016 ("C2c2 is a single-component programmable RNA-guided RNA-targeting CRISPR effector"). The enzyme — initially called C2c2, later renamed Cas13 — is a Class 2, Type VI CRISPR effector that uses a single crRNA to find and cleave complementary RNA. Unlike Cas9 and Cas12, Cas13 has nothing to do with DNA at all.
Several variants have since been characterized:
- Cas13a (LwaCas13a) — the original C2c2; powers most SHERLOCK diagnostics.
- Cas13b (PspCas13b) — high knockdown efficiency in mammalian cells; the basis of REPAIR (RNA editing by ADAR fusions).
- Cas13c — less commonly used.
- Cas13d (RfxCas13d, also called CasRx) — the smallest and currently most efficient for therapeutic RNA knockdown.
How Cas13 Works at the Molecular Level
Cas13 has two HEPN ribonuclease domains that come together upon target binding. The mechanism is unusual:
1. Target recognition. A crRNA guides Cas13 to a complementary RNA sequence. There is no PAM in the strict sense, but Cas13a/b have a "protospacer flanking site" preference. Cas13d (CasRx) has essentially no flanking-site requirement, which makes it the most flexible.
2. Activation by binding. Once Cas13 binds its target RNA, the two HEPN domains assemble into a single active site on the protein's surface — not buried inside it like Cas9's nuclease.
3. Collateral cleavage. Activated Cas13 cleaves not only the target RNA but any single-stranded RNA in solution. In a test tube with a fluorescent reporter RNA, this collateral activity produces a sensitive signal. In living human cells, it is still debated whether and how much collateral activity occurs — early concerns from Wessels et al. (2020) have been partly tempered by later optimization (Kelley et al., 2022; Tong et al., 2023).
4. Catalytically dead Cas13 (dCas13) for editing. Mutating the HEPN residues produces a dead Cas13 that binds RNA without cutting. Fused to ADAR2 deaminase domains, dCas13 becomes REPAIR (Cox et al., 2017) for A-to-I edits and RESCUE (Abudayyeh et al., 2019) for C-to-U edits — making it possible to rewrite individual RNA bases without ever modifying the genome.
Key Papers and Milestones
- Abudayyeh et al., 2016 (Science). Discovery of C2c2/Cas13a.
- Gootenberg et al., 2017 (Science). SHERLOCK — Cas13a-based diagnostics with attomolar sensitivity.
- Cox et al., 2017 (Science). REPAIR — programmable A-to-I RNA editing using dCas13b-ADAR2.
- Konermann et al., 2018 (Cell). Discovery and characterization of Cas13d / CasRx — smaller, more efficient mammalian RNA knockdown.
- Gootenberg et al., 2018 (Science). SHERLOCKv2 — multiplexed and quantitative diagnostics.
- Abudayyeh et al., 2019 (Science). RESCUE — C-to-U RNA editing.
- Wessels et al., 2020 (Nature Biotechnology). Genome-wide CasRx screen design rules.
- Broughton et al., 2020 (Nature Biotechnology). SHERLOCK and DETECTR for SARS-CoV-2.
- Kushawah et al., 2020 (Developmental Cell). CasRx for embryonic RNA knockdown.
Applications and Use Cases
Therapeutic RNA knockdown. This is the area moving fastest. Locanabio (acquired by Vertex in 2024) developed RNA-targeting CRISPR therapies for Huntington's disease and myotonic dystrophy using CasRx-based knockdown of toxic repeat-containing RNAs. Editas Medicine has also disclosed an RNA-targeting program. The appeal is straightforward: Cas13 silences disease RNAs reversibly and never touches the genome — no off-target indels, no large deletions, no p53 activation.
RNA base editing. REPAIR (A-to-I) and RESCUE (C-to-U) provide a complementary approach to DNA base editing for diseases caused by point mutations — particularly attractive when permanent DNA changes are too risky.
Diagnostics. SHERLOCK (Sherlock Biosciences) and DETECTR (Mammoth Biosciences) achieved FDA emergency use authorization for SARS-CoV-2 detection in 2020. Both platforms can detect a few molecules per microliter with simple paper-strip readouts. They have since been adapted for tuberculosis, HPV, and a growing list of other pathogens.
Functional genomics. CasRx has become a popular alternative to RNAi for transcriptome-wide loss-of-function screens — especially for non-coding RNAs that RNAi handles poorly.
Live-cell RNA imaging. dCas13 fused to fluorescent proteins enables tracking of specific RNAs in living cells.
Cas13 vs DNA-Targeting CRISPR vs ADAR-Based RNA Editing
| Feature | Cas13 (CasRx, RfxCas13d) | Cas9 / Cas12 (DNA) | ADAR-based RNA editing (no Cas13) |
|---|---|---|---|
| Substrate | Single-stranded RNA | Double-stranded DNA | Double-stranded RNA |
| Permanent? | No (RNA turns over) | Yes | No |
| Off-target genomic edits | None | Possible | None |
| Reversible | Yes | No | Yes |
| Delivery | Editor + crRNA | Editor + sgRNA (+ HDR template) | Antisense oligo or vector |
| Best use | Toxic gain-of-function RNAs | Loss-of-function or correction | Single A-to-I edits |
Connection to the Broader Gene Editing Ecosystem
Cas13 is the RNA-world counterpart to the DNA-targeting CRISPR and Cas12 systems. Like CRISPRi, it offers reversible silencing — but at the post-transcriptional rather than transcriptional level. Like base editing, it can change individual letters — but in RNA rather than DNA. The larger story is that gene editing has fragmented into a toolkit, each tool optimized for a different biological substrate and a different therapeutic risk profile — a development tied closely to David Liu and Feng Zhang. Cas13 also shares the delivery-system constraints of every CRISPR therapy and is being explored with both LNP mRNA and AAV approaches.
Current Limitations and Challenges
- Collateral activity in cells. The biggest open question. Some studies report robust trans-cleavage in mammalian cells; others report negligible effects with current variants. Cell type and target abundance matter.
- Delivery. CasRx is small (~970 aa) and fits AAV, but the editor must be sustained for ongoing knockdown — creating long-term immunogenicity questions.
- Immunogenicity of bacterial Cas13. As with Cas9, anti-Cas13 antibodies and T-cell responses are likely in many adults.
- Off-target hybridization. Cas13 crRNAs are short and can mismatch tolerantly, producing knockdown of unintended transcripts.
- No genomic permanence. A double-edged sword — safer, but the patient needs continuous expression for chronic indications.
FAQ
Does Cas13 cut DNA?
No. Cas13 cleaves only single-stranded RNA. The genome is not touched.
What is CasRx?
CasRx is RfxCas13d — a Cas13 variant from Ruminococcus flavefaciens discovered by the Hsu and Konermann labs in 2018. It is the smallest and most efficient Cas13 for mammalian RNA knockdown.
How is Cas13 different from RNAi?
RNAi uses small RNAs and the endogenous Argonaute machinery, which can be saturated and produces seed-based off-targets. Cas13 brings its own RNase, can be guided more specifically, and silences non-coding RNAs more readily.
Is collateral cleavage a problem in human cells?
It depends on the variant, the target, and the cell type. Engineered Cas13 variants and high-fidelity crRNAs have substantially reduced — but not eliminated — the concern. It remains an active research area.
Are Cas13 therapies in clinical trials?
Locanabio had Cas13-based programs in late preclinical development before its 2024 acquisition by Vertex. Several academic groups are pursuing IND-enabling studies. As of 2026, no Cas13 therapy has reached Phase 1.
What is SHERLOCK?
SHERLOCK (Specific High-sensitivity Enzymatic Reporter unLOCKing) is the Cas13a-based diagnostic platform from the Zhang lab and Sherlock Biosciences, capable of detecting nucleic acids at single-molecule sensitivity using a paper-strip readout.
