Tensing Flipper: Photosensitized\nManipulation of Membrane\nTension, Lipid Phase Separation, and Raft Protein Sorting in Biological Membranes

The lateral organization of proteins and lipids in the\nplasma membrane\nis fundamental to regulating a wide range of cellular processes. Compartmentalized\nordered membrane domains enriched with specific lipids, often termed\nlipid rafts, have been shown to modulate the physicochemical and mechanical\nproperties of membranes and to drive protein sorting. Novel methods\nand tools enabling the visualization, characterization, and/or manipulation\nof membrane compartmentalization are crucial to link the properties\nof the membrane with cell functions. Flipper, a commercially available\nfluorescent membrane tension probe, has become a reference tool for\nquantitative membrane tension studies in living cells. Here, we report\non a so far unidentified property of Flipper, namely, its ability\nto photosensitize singlet oxygen (<sup>1</sup>O<sub>2</sub>) under\nblue light when embedded into lipid membranes. This in turn results\nin the production of lipid hydroperoxides that increase membrane tension\nand trigger phase separation. In biological membranes, the photoinduced\nsegregated domains retain the sorting ability of intact phase-separated\nmembranes, directing raft and nonraft proteins into ordered and disordered\nregions, respectively, in contrast to radical-based photo-oxidation\nreactions that disrupt raft protein partitioning. The dual tension\nreporting and photosensitizing abilities of Flipper enable simultaneous\nvisualization and manipulation of the mechanical properties and lateral\norganization of membranes, providing a powerful tool to optically\ncontrol lipid raft formation and to explore the interplay between\nmembrane biophysics and cell function.