Light-triggered site-directed RNA editing by endogenous ADAR1 with photolabile guide RNA

RNA A-to-I editing is a post-transcriptional modification pervasively occurring in cells. Artificial intervention of A-to-I editing at specific sites of RNA could also be achieved with guide RNA and exogenous ADAR en-zymes. In contrast to previous fused SNAP-ADAR enzymes for light-driven RNA A-to-I editing, we developed photo-caged antisense guide RNA oligonucleotides with simple 30-terminal cholesterol modification, and successfully achieved light-triggered site-specific RNA A-to-I editing for the first time utilizing endogenous ADAR enzymes. Our caged A-to-I editing system effectively implemented light-dependent point mutation of mRNA transcripts of both exogenous and endogenous genes in living cells and 3D tumorspheres, as well as spatial regulation of EGFP expression, which provides a new approach for precise manipulation of RNA editing.