Abstract
BACKGROUND: RNA-targeting CRISPR-Cas can provide potential advantages over DNA editing, such as avoiding pleiotropic effects of genome editing, providing precise spatiotemporal regulation, and expanded function including antiviral immunity. RESULTS: Here, we report the use of CRISPR-Cas13 in plants to reduce both viral and endogenous RNA. Unexpectedly, we observe that crRNA designed to guide Cas13 could, in the absence of the Cas13 protein, cause substantial reduction in RNA levels as well. We demonstrate Cas13-independent guide-induced gene silencing (GIGS) in three plant species, including stable transgenic Arabidopsis. Small RNA sequencing during GIGS identifies the production of small RNA that extend beyond the crRNA expressed sequence in samples expressing multi-guide crRNA. Additionally, we demonstrate that mismatches in guide sequences at position 10 and 11 abolish GIGS. Finally, we show that GIGS is elicited by guides that lack the Cas13 direct repeat and can extend to Cas9 designed crRNA of at least 28 base pairs, indicating that GIGS can be elicited through a variety of guide designs and is not dependent on Cas13 crRNA sequences or design. CONCLUSIONS: Collectively, our results suggest that GIGS utilizes endogenous RNAi machinery despite the fact that crRNA are unlike canonical triggers of RNAi such as miRNA, hairpins, or long double-stranded RNA. Given similar evidence of Cas13-independent silencing in an insect system, it is likely GIGS is active across many eukaryotes. Our results show that GIGS offers a novel and flexible approach to RNA reduction with potential benefits over existing technologies for crop improvement and functional genomics.
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Claims 8
Claim · model-written summary
CRISPR-Cas13 reduced viral and endogenous RNA in plants.
- Taxon
- plantsViridiplantaee0
- Treatment
- CRISPR-Cas13e0
- Outcome
- viral and endogenous RNAe0
- Direction
- reducee0
Not reportedComparator
Verbatim
e0“the use of CRISPR-Cas13 in plants to reduce both viral and endogenous RNA”
Claim · model-written summary
Cas13 guide RNA substantially reduced RNA levels without Cas13 protein.
Verbatim
e0“crRNA designed to guide Cas13 could, in the absence of the Cas13 protein, cause substantial reduction in RNA levels as well”
e1“in plants”
Claim · model-written summary
Cas13-independent guide-induced gene silencing occurred in three plant species including transgenic Arabidopsis.
- Taxon
- Arabidopsis
Arabidopsise0 - Outcome
- Cas13-independent guide-induced gene silencinge0
- Value
- three plant speciese0
Organisms pictured · Representative pictures of the matched species, not figures from the paper.
Verbatim
e0“Cas13-independent guide-induced gene silencing (GIGS) in three plant species, including stable transgenic Arabidopsis”
Claim · model-written summary
Multi-guide crRNA expression produced small RNAs beyond the expressed crRNA sequence.
- Taxon
- plantsViridiplantaee1
- Treatment
- multi-guide crRNAe0
- Outcome
- small RNA that extend beyond the crRNA expressed sequencee0
Verbatim
e0“production of small RNA that extend beyond the crRNA expressed sequence in samples expressing multi-guide crRNA”
e1“in plants”
Claim · model-written summary
Guide mismatches at positions 10 and 11 abolished guide-induced silencing.
- Taxon
- plantsViridiplantaee1
- Treatment
- mismatches in guide sequences at position 10 and 11e0
- Outcome
- GIGSe0
Not reportedComparatorDirection
Verbatim
e0“mismatches in guide sequences at position 10 and 11 abolish GIGS”
e1“in plants”
Claim · model-written summary
Guides without the Cas13 direct repeat elicited guide-induced silencing.
- Taxon
- plantsViridiplantaee1
- Treatment
- guides that lack the Cas13 direct repeate0
- Outcome
- GIGSe0
Not reportedComparatorDirection
Verbatim
e0“GIGS is elicited by guides that lack the Cas13 direct repeat”
e1“in plants”
Claim · model-written summary
Cas9-designed crRNA of at least 28 base pairs also elicited guide-induced silencing.
- Taxon
- plantsViridiplantaee1
- Treatment
- Cas9 designed crRNAe0
- Outcome
- Cas9 designed crRNAe0
- Value
- at least 28 base pairse0
Not reportedComparatorDirection
Verbatim
e0“can extend to Cas9 designed crRNA of at least 28 base pairs”
e1“in plants”
Claim · model-written summary
The authors infer that guide-induced silencing uses endogenous RNAi machinery.
- Taxon
- plantsViridiplantaee1
- Outcome
- GIGSe0
- Qualifier
- endogenous RNAi machinerye0
Verbatim
e0“GIGS utilizes endogenous RNAi machinery”
e1“in plants”
The same questions in other papers
loss of function · molecular: metabolites · Viridiplantae (plants)Positive and negative results
4 papers in the same direction as this paper · 8 in the opposite direction · 1 reporting no difference from an alternative
- negative
- negative
- negative
10 papers more in this group
See all 14 papers →Claims are grouped by organism, treatment and what was measured or targeted. A paper counts as positive when any of its claims in the group is positive.