Conventional photodynamic therapy is severely constrained by the limited light-penetration depth in tissue. Here, we show efficient photodynamic therapy (PDT) mediated by bioluminescence resonance energy transfer (BRET) that overcomes the light-penetration limitation. The photosensitizer Rose Bengal (RB) was loaded in biodegradable poly(lactic-co-glycolic acid) (PLGA) nanoparticles, which were then conjugated with firefly luciferase. Spectroscopic characterizations indicated that BRET effectively activated RB to generate reactive oxygen species (ROS). In vitro studies of the cellular cytotoxicity and photodynamic effect indicated that cancer cells were effectively destroyed by BRET-PDT treatment. In vivo studies in a tumor-bearing mouse model demonstrated that tumor growth was significantly inhibited by BRET-PDT in the absence of external light irradiation. The BRET-mediated phototherapy provides a promising approach to overcome the light-penetration limitation in photodynamic treatment of deep-seated tumors.