Light‐Activated Control of Translation by Enzymatic Covalent mRNA Labeling

Activation of cellular protein expression upon visible‐light photocleavage of small‐molecule caging groups covalently attached to the 5′ untranslated region (5′ UTR) of an mRNA was achieved. These photocleavable caging groups are conjugated to in vitro transcribed mRNA (IVT‐mRNA) through RNA transglycosylation, an enzymatic process in which a bacterial tRNA guanine transglycosylase (TGT) exchanges a guanine nucleobase in a specific 17‐nucleotide motif (Tag) for synthetic pre‐queuosine1 (preQ1) derivatives. The caging groups severely reduce mRNA translation efficiency when strategically placed in the 5′ UTR. Using this method, we demonstrate the successful spatiotemporal photoregulation of gene expression with single‐cell precision. Our method can be applied to therapeutically relevant chemically modified mRNA (mod‐mRNA) transcripts. This strategy provides a modular and efficient approach for developing synthetic gene regulatory circuits, biotechnological applications, and therapeutic discovery. Freie Fahrt für die Translation: Die Gentranslation kann durch Abspaltung hemmender Gruppen (rot) vom untranslatierten 5′‐Ende von mRNA unter Bestrahlung mit sichtbarem Licht aktiviert werden. Die hemmenden Gruppen werden durch RNA‐Transglykosylierung angebracht und reduzieren die mRNA‐Translationseffizienz erheblich. Mit dieser Methode wird die räumliche und zeitliche Steuerung der Genexpression mithilfe von Licht auf der Ebene einzelner Zellen möglich.