Engineered TALEDs Improve Targeted Base Editing of Mitochondrial DNA

Targeted base editing of mammalian mtDNA is a powerful tool for modeling mitochondrial genetic diseases and developing potential therapies. Programmable deaminases, such as cytosine and adenine base editors, enable precise mtDNA editing by binding to specific DNA sites in the mitochondrial genome and converting bases. However, current approaches have limitations, including off-target edits.

Now, researchers from Korea University have engineered transcription activator-like effector-linked deaminases (TALEDs) for improved precision editing, resulting in an animal model with A-to-G mtDNA edits. The engineered TALEDs decreased off-target RNA edits and minimized off-target mutations in mtDNA, without exhibiting toxicity or causing developmental issues in mouse embryos.

“To address limitations of the conventionally used TALEDs, we created and assessed 209 TALED variants. Each variant replaced one of the 11 amino acid residues near the substrate-binding pocket in the protein of interest with one of the other 19 amino acid residues,” explained study leader Hyunji Lee.

The results  of the study, published in Cell, showed that the engineered TALEDs significantly decreased off-target RNA edits by over 99%. The engineered TALEDs also minimized off-target mutations in mtDNA and bystander edits—unintended changes that occur at locations in the genome near the targeted site during editing processes—at a specific target site. In contrast to the original TALEDs, these engineered TALEDs did not exhibit toxicity. They also did not lead to developmental arrest in mouse embryos.

The ability to edit mtDNA with improved precision opens avenues for studying and treating mitochondrial genetic disorders. The research aligns with the broader trend of personalized medicine, offering hope for individuals with specific mitochondrial genetic variations.