Lysosome-Associated MiniSOG as a Photosensitizer for Mammalian Cells

Abstract

Genetically encoded photosensitizers represent a promising optogenetic tool for the induction of light-controlled oxidative stress strictly localized to a selected intracellular compartment. Here we tested the phototoxic effects of the flavin-containing phototoxic protein miniSOG targeted to the cytoplasmic surfaces of late endosomes and lysosomes by fusion with Rab7. In HeLa Kyoto cells stably expressing miniSOG-Rab7, we demonstrated a high level of cell death upon blue-light illumination. Pepstatin A completely abolished phototoxicity of miniSOG-Rab7, showing a key role for cathepsin D in this model. Using a far-red fluorescence sensor for caspase-3, we observed caspase-3 activation during miniSOG-Rab7–mediated cell death. We conclude that upon illumination, miniSOG-Rab7 induces lysosomal membrane permeabilization (LMP) and leakage of cathepsins into the cytosol, resulting in caspase-dependent apoptosis.

Natural and Designed Toxins for Precise Therapy: Modern Approaches in Experimental Oncology

“…This new type of biological PSs is considered as a promising substitute for current synthetic photosensitizes used in PDT, and as it was shown in a series of in vitro investigations, miniSOG and KillerRed possess phototoxicity equal or exceeding that of commonly used PSs or other fluorescent chromoproteins [ 8 , 9 , 156 ]. Remarkable phototoxicity, in addition to water solubility and biocompatibility, has placed genetically encoded PSs among the top ideal hydrophilic candidates for PDT, which has been successfully proven for photoablation in cell models [ 9 , 157 , 158 , 159 , 160 ].…”

Subcellular Singlet Oxygen and Cell Death: Location Matters

“…Other investigators have found both similar and different orders of efficacy in cell ablation when targeting subcellular organelles. Using miniSOG, Ryumina et al (2013Ryumina et al ( , 2016 found that membrane targeting was more effective than mitochondrial, lysosomal, or nuclear targeting. Xu and Chisolm (Xu and Chisholm, 2016), using miniSOG2, also found membrane targeting more effective than cytoplasmic or mitochondrial targeting.…”

Photodynamic Physiology—Photonanomanipulations in Cellular Physiology with Protein Photosensitizers

“…For lysosomal targeting, the C-terminal cytosolic tail sequence of the lysosomal-associated membrane protein II (LIMP II) could be used (Tabuchi et al, 2000). GTPase Rab7A sequence has been used to target KillerRed to lysosomes (Serebrovskaya et al, 2011; Ryumina et al, 2016). ER targeting could use the ER localization sequence (MLLSVPLLLGLLGLAVA) of calreticulin (Fliegel et al, 1989) and the ER retaining sequence KDEL (Munro and Pelham, 1987).…”