Photocaged Dinucleoside Tetraphosphates for Light‐Mediated Activation of 5′ Cap Interactions and Translation In Vitro

Dinucleoside tetraphosphates (Np 4 N) have been reported as 5′ caps of RNAs in bacteria and mammalian cells. The guanosine‐based version (Gp 4 N) resembles the canonical eukaryotic 5′ cap (m 7 Gp 3 N), which is a central platform for cap‐interacting proteins. Herein, photocaged versions of m 7 Gp 4 G are reported. The photocleavable protecting group at the N 2 position of m 7 G prevents further methylation and decapping, but not incorporation of these 5′ caps into RNA by in vitro transcription. The resulting caged‐m 7 Gp 4 G‐RNA is not translated. Upon irradiation, all interactions are restored, and methylation, decapping, and translation are observed in vitro. Photocaged dinucleoside tetraphosphates serve as valuable tools for precisely investigating the interactions of Np 4 Ns and Np 4 N‐capped RNAs with cap‐binding and cap‐modifying enzymes. Additionally, they enable temporal control over the activation of these interactions and functions.