A lysosomal NO-selective fluorescent probe enables bioorthogonal exploration of NO involved in autophagy and ferroptosis

Compartmentalized nitric oxide (NO) within specific organelles in cells has been associated with the selective expression of NO syntheses (NOSs) in these organelles. Moreover, it has been reported that lysosomal NOS expression contributes to autophagy and ferroptosis. Unfortunately, a method for detecting NO within lysosomes in live cells unavailable yet. In this study, we developed a lysosomal NO-selective chemosensor (Lyso-NO), composed of a 4-(4-nitrophenyl)thiosemicarbazide-linked naphthalimide as an NO-mediated fluorogenic reporter and a morpholine moiety as a lysosome-targeting unit. Upon selective reaction with NO in solution and within live cells, Lyso-NO showed a strong fluorescence intensity at 550 nm, and its predominant lysosomal location was revealed by confocal microscopy. Further, lysosomal NO increase was demonstrated to be deeply involved in autolysosome formation during autophagy, and in ferroptosis. Lyso-NO was considered to act as a lysosomal NO-selective sensor, allowing for the bioorthogonal analysis of lysosome-related biological processes.