Development of a UV-Responsive Fusion Peptide Combining a Cell-Penetrating and a Mitochondria-Targeting Peptide to Suppress Intracellular Off-Target Effects

Modification of plant mitochondria has attracted increasing attention as a strategy for developing highly productive crops, driving efforts to achieve efficient mitochondrial delivery of cargos. Mitochondria-targeting gene carriers with a strong cell-penetrating ability have been shown to deliver plasmid DNA into plant mitochondria, but this ability can reduce mitochondrial selectivity through nonspecific interaction with intracellular membranes. Herein, we developed a fusion peptide in which a mitochondria-targeting peptide was linked to a cell-penetrating peptide via an ultraviolet (UV)-cleavable linker, enabling regulation of cell-penetrating function after cellular uptake by UV irradiation. As a result, mitochondrial localization of the fusion peptide was significantly enhanced in Arabidopsis thaliana. Furthermore, UV irradiation improved the mitochondrial gene delivery efficiency and selectivity of gene carriers modified with the fusion peptide compared with nonirradiated conditions. Overall, these findings demonstrate that controlled release of the cell-penetrating peptide suppresses off-target effects and enables selective mitochondrial gene delivery in plant cells.