Degranulation-enhanced presynaptic membrane packing protects NK cells from perforin-mediated autolysis

Abstract

Abstract NK cells are resistant to autolysis when they kill a target cell using perforin secretion into the lytic immunological synapse. Perforin makes pores in target cell membranes allowing delivery of pro-apoptotic enzymes. Despite the fact that the perforin-containing organelles (lytic granules) are released in close range to both the NK and target cell, the NK cell membrane is protected. How NK cells avoid autolysis during degranulation is perplexing. We demonstrate that NK cells are protected from perforin by their densely packed presynaptic membranes. When treated with 7-ketocholesterol lipid packing is reduced in NK cells, making them susceptible to perforin mediated autolysis after degranulation. Using advanced imaging, we showed that lytic granules themselves have endogenously densely packed membranes. Importantly, during degranulation lytic granule-cell membrane fusion further reinforces local presynaptic membrane packing. This provides enhanced membrane protection at the specific sites of degranulation where NK cells face maximum local concentrations of secreted perforin. Additionally, we found that an aggressive breast cancer cell line utilizes the same strategy to evade NK cellmediated killing. These cells are perforin-resistant owing to a densely packed postsynaptic membrane. By disrupting membrane packing, we effectively switched them to an NK-susceptible state. Overall, we reveal an unexpected role for lipid membranes in NK cell functionality and that degranulation uses lytic granule membrane packing to create local “shields” against autolysis. Furthermore, lipid membrane modulation could alter the susceptibility of perforin-resistant cancer cells and should be considered in cytotoxic cell therapies.