RNA editing

Circular RNA

ADAR

Biology

Guide RNA

Gene

Mutant

Genome editing

Cas9

Point mutation

Bystander effect

In vitro

Immunology

Competing endogenous RNA

Computational biology

Cell biology

CRISPR

Genetics

Long non-coding RNA

Nature Biotechnology

Current methods for programmed RNA editing using endogenous ADAR enzymes and engineered ADAR-recruiting RNAs (arRNAs) suffer from low efficiency and bystander off-target editing. Here, we describe LEAPER 2.0, an updated version of LEAPER that uses covalently closed circular arRNAs, termed circ-arRNAs. We demonstrate on average ~3.1-fold higher editing efficiency than their linear counterparts when expressed in cells or delivered as in vitro-transcribed circular RNA oligonucleotides. To lower off-target editing we deleted pairings of uridines with off-target adenosines, which almost completely eliminated bystander off-target adenosine editing. Engineered circ-arRNAs enhanced the efficiency and fidelity of editing endogenous CTNNB1 and mutant TP53 transcripts in cell culture. Delivery of circ-arRNAs using adeno-associated virus in a mouse model of Hurler syndrome corrected the pathogenic point mutation and restored α-L-iduronidase catalytic activity, lowering glycosaminoglycan accumulation in the liver. LEAPER 2.0 provides a new design of arRNA that enables more precise, efficient RNA editing with broad applicability for therapy and basic research. Circular ADAR-recruiting RNAs improve RNA editing in vitro and in a mouse disease model.

Engineered circular ADAR-recruiting RNAs increase the efficiency and fidelity of RNA editing in vitro and in vivo