Necroptosis is preceded by nuclear translocation of the signaling proteins that induce it

A signaling pathway that induces programmed necrotic cell death (necroptosis) was reported to be activated in cells by several cytokines and various pathogen components. The major proteins participating in that pathway are the protein kinases RIPK1 and RIPK3 and the pseudokinase mixed lineage kinase domain-like protein (MLKL). Recent studies have suggested that MLKL, once activated, mediates necroptosis by binding to cellular membranes, thereby triggering ion fluxes. However, our knowledge of both the sequence of molecular events leading to MLKL activation and the subcellular sites of these events is fragmentary. Here we report that the association of MLKL with the cell membrane in necroptotic death is preceded by the translocation of phosphorylated MLKL, along with RIPK1 and RIPK3, to the nucleus.Apart from the apoptotic cell death pathway that ligands of the tumor necrosis factor (TNF) family can activate, these ligands and various other inducers, including the interferons and various pathogen components, have in recent years been found also to trigger a signaling cascade that induces programmed necrotic death (necroptosis). This cascade encompasses sequential activation of the protein kinases RIPK1 and RIPK3 and the pseudokinase mixed lineage kinase domain-like protein (MLKL).^{1, 2, 3, 4, 5} RIPK3-mediated phosphorylation of MLKL triggers its oligomerization, which is necessary and sufficient for the induction of cell death,^{6, 7, 8} and can also trigger some non-deadly functions.⁹ MLKL was recently suggested to trigger cell death by binding to cellular membranes and initiating ion fluxes through them.^{6, 7, 8, 10} However, its exact molecular target in death induction is contentious.^{6, 8, 10, 11, 12} Current knowledge of the subcellular sites of MLKL action is based mainly on determination of the location of this protein close to the time of cell death. Here we present a detailed assessment of the cellular location of MLKL at different times following its activation. Our findings indicate that before cell death, MLKL translocates to the nucleus along with RIPK1 and RIPK3.