LAUSR

In situ imaging the repair activity of 8-oxoguanine (8-OG) DNA glycosylase in living cells is important as it is associated with genetic mutation. However, the existing imaging methods confront the interference of intracellular nuclease and resulting in false positive signal. Here, a closing-upon-repair DNA tetrahedron nanoswitch (CRTN) was designed for FRET imaging the repair activity of 8-OG DNA glycosylase in living cells with high specificity and accuracy. CRTN comprised a DNA tetrahedron, a recognition strand modified with 8-OG bases, and a reporting strand designed as hairpin structure and labeled with Cy3/Cy5 dual fluorophores. Initially, the DNA tetrahedron was linked with the reporting strand hybridized to the recognition strand, separating the Cy3 donor and Cy5 acceptor into FRET-invalid distance. Upon repair the 8-OG bases by 8-OG DNA glycosylase, CRTN could undergo a structure change from the open to closed state. Specifically, the reporting strand was dissociated from the recognition strand under the action of 8-OG DNA glycosylase and folded into hairpin structure, bringing the Cy3 donor and Cy5 acceptor into FRET-valid proximity with the generation of FRET signal, which could prevent false positive signal arising from nuclease degradation. CRTN exhibited the feasibility for detecting 8-OG DNA glycosylase activity in vitro with good sensitivity and selectivity. More importantly, CRTN could enter cells without any transfection for FRET imaging the repair activity of intracellular 8-OG DNA glycosylase with high specificity and accuracy. This approach provided a promising tool for deeper understanding 8-OG DNA glycosylase function and further studying genetic mutation-related diseases.