An Endogenously Gated DNA Nanodevice Enables Mitochondria-Specific FEN1 Monitoring in Response to Therapeutic Challenge

Monitoring therapy-induced molecular changes within subcellular compartments is crucial for understanding cellular stress and providing clues to treatment response. Mitochondrial base excision repair (BER) enzymes have emerged as promising indicators of therapeutic response, yet effective tools for their on-site and dynamic monitoring remain scarce. Here, we report an endogenously gated DNA nanodevice (EGDN) capable of real-time, mitochondria-specific monitoring of flap endonuclease 1 (FEN1), a key BER enzyme, under therapeutic challenge. The EGDN remains silent during transport and is sequentially activated by the mitochondria-encoded ribosomal RNA and FEN1, thus achieving spatial control over FEN1 imaging. This conditional regulation pattern reduces nonspecific signals and sharpens the spatial resolution of FEN1 visualization within mitochondria. We further demonstrate that EGDN dynamically tracks mitochondrial FEN1 fluctuations in tumor cells undergoing chemotherapy or radiation therapy, providing a molecular readout of treatment-associated alterations. Collectively, this work establishes a robust biosensing toolbox for dissecting the BER process at the subcellular level, with broad implications for cancer early diagnosis and targeted therapy.