Phosphatidylserine externalization, “necroptotic bodies” release, and phagocytosis during necroptosis

Necroptosis is a regulated, nonapoptotic form of cell death initiated by receptor-interacting protein kinase-3 (RIPK3) and mixed lineage kinase domain-like (MLKL) proteins.It is considered to be a form of regulated necrosis, and, by lacking the "find me" and "eat me" signals that are a feature of apoptosis, necroptosis is considered to be inflammatory.One such "eat me" signal observed during apoptosis is the exposure of phosphatidylserine (PS) on the outer plasma membrane.Here, we demonstrate that necroptotic cells also expose PS after phosphorylated mixed lineage kinase-like (pMLKL) translocation to the membrane.Necroptotic cells that expose PS release extracellular vesicles containing proteins and pMLKL to their surroundings.Furthermore, inhibition of pMLKL after PS exposure can reverse the process of necroptosis and restore cell viability.Finally, externalization of PS by necroptotic cells drives recognition and phagocytosis, and this may limit the inflammatory response to this nonapoptotic form of cell death.The exposure of PS to the outer membrane and to extracellular vesicles is therefore a feature of necroptotic cell death and may serve to provide an immunologically-silent window by generating specific "find me" and "eat me" signals. Author summaryNecroptosis, a recently discovered regulated form of cell death, is widely considered to be inflammatory due to the absence of specific "find me" and "eat me" signals prior to lytic death.Here, we demonstrate that necroptotic cells generate "find me" and "eat me" signals by exposure of phosphatidylserine on their outer plasma membrane.This was further associated with the release of extracellular vesicles ("necroptotic bodies") that contain phosphatidylserine, pMLKL (a key necroptotic marker), as well as other proteins.These signals drive recognition and phagocytosis of necroptotic cells to modulate the immune response.The exposure of phosphatidylserine and release of "necroptotic bodies" indicate